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Names of different cyclins

Names of different cyclins


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Different types of cell cyclins are designated as a to y

Why are some letters like m, n, p, q… etc. skipped?

Source: https://en.wikipedia.org/wiki/Cyclin


Biological nomenclature can be impenetrable. Almost certainly, at some point in history, there were cyclins designated with these letters. Researchers would have discovered apparently novel cyclins which were subsequently determined to be part of an existing family and renamed. For example, this paper reports the discovery of:

a new cyclin, cyclin M, which appears to be most closely related to cyclin L. Its biological function is unknown, but it is related to cyclins that regulate transcription.

It is now called cyclin L2.

I hope this answer satisfies you to some extent. I doubt anyone is going to sift through the literature and try and determine why all cyclins are are named as they are; that is certainly a losing proposition. The naming of genes/proteins is rather trivial and doesn't generally follow any specific set of rules. Outside of curiosity, the knowledge of cyclin naming doesn't have much, if any, practical relevance.


Before directly answering your question, it's first worth mentioning that: within the table you provided, each cyclin protein is actually being referenced to by using the name of the gene that codes for it. That being said, in order to explain why the naming is the way it is, we need to consider the naming system for genes!


The institution that regulates gene naming is the HUGO Gene Nomenclature Committee, and within the HGNC website, they have a page that outlines the gene naming guidelines.

By considering the gene naming guideline page, and the cyclin-coding gene family, we can immediately say that the first three letters represent the CCN family, which stand for: Connective tissue growth factor, Cystein rich protein, and Nephroblastoma overexpressed gene. (source)

From there, the remaining letters in each acroynm represent the subfamily that the gene belongs to. These subfamilies are defined by a domain within the protein, and are abbreviated based on how that domain is named. For example, when considering CCNF, the F stands for "F-Box", which is a protein structural motif that, in this specific case, mediates protein-protein interactions.


So, to now answer your question as to why the letters are skipped: there is no specific reason (sorry to say it)! It really just comes down to who/when/how the protein motifs were discovered/named, and the fact that a single person/institution didn't discover them all, and that sometimes (quite often) biological naming isn't fixed after the fact to be so perfect.

A (somewhat) similiar situation of this can be seen with how vitamins are named… they were originally named using the (English) alphabet, in a sequential manner upon discovery, but then it was decided to stop this convention (I'm not sure why), and upon doing so went go back and declassified some vitamins because they no longer conformed to newer standards, which broke the (nice) sequential naming that was originally in place. So, it truely is a purely historical reason, and has no meaning beyond that.

And lastly, just to be inclusive, the number(s) at the end of each acronym are used to uniquely identify the multiple members within each family.


The regulation of the cell cycle

Pioneering work by Paul Nurse and Leland Hartwell identified genetic mutants that regulate cell cycle progression in Saccharomyces pombe and Saccharomyces cerevisiae respectively. Furthermore, the discovery of maturation-promoting factor in Xenopus laevis by Yoshio Masui together with the identification of cyclins in sea urchin eggs by Tim Hunt led to the discovery of key components that regulate the cell cycle. In 2001, Paul Nurse, Leland Hartwell, and Tim Hunt received the Nobel prize in physiology or medicine for their contribution to the understanding of cell cycle regulation (Pulverer, 2001) .

The mammalian cell cycle can be divided into the four distinct phases Gap 1 (G1), synthesis (S), Gap 2 (G2), and mitosis (M phase) which require three ‘switch-like’ transitions to regulate S phase entry, mitotic entry, and the exit of mitosis (Hochegger et al., 2008). Mitosis is subdivided into a further six phases including prophase, pro-metaphase, metaphase, anaphase, telophase, and cytokinesis which control nuclear envelope breakdown (NEBD), the attachment of chromosomes to spindle poles, the alignment of chromosomes upon the metaphase plate, the separation of sister chromatids and the separation of the cell into two daughter cells respectively.


Contents

Table 1: Known CDKs, their cyclin partners, and their functions in the human [3] and consequences of deletion in mice [4]
CDK ! Cyclin partner Function Deletion Phenotype in Mice
Cdk1 Cyclin B M phase None
Cdk2 Cyclin E G1/S transition Reduced size, imparted neural progenitor cell proliferation. Viable, but both males & females sterile.
Cdk2 Cyclin A S phase, G2 phase
Cdk3 Cyclin C G1 phase No defects. Viable, fertile.
Cdk4 Cyclin D G1 phase Reduced size, insulin deficient diabetes. Viable, but both male & female infertile.

Most of the known cyclin-CDK complexes regulate the progression through the cell cycle. Animal cells contain at least nine CDKs, four of which, CDK1, 2, 3, and 4, are directly involved in cell cycle regulation. [1] In mammalian cells, CDK1, with its partners cyclin A2 and B1, alone can drive the cell cycle. [4] Another one, CDK7, is involved indirectly as the CDK-activating kinase. [1] Cyclin-CDK complexes phosphorylate substrates appropriate for the particular cell cycle phase. [3] Cyclin-CDK complexes of earlier cell-cycle phase help activate cyclin-CDK complexes in later phase. [1]

Table 2: Cyclins and CDKs by Cell-Cycle Phase
Phase Cyclin CDK
G0 C Cdk3
G1 D, E Cdk4, Cdk2, Cdk6
S A, E Cdk2
G2 A Cdk2, Cdk1
M B Cdk1
Table 3: Cyclin-dependent kinases that control the cell cycle in model organisms [1]
Species Name Original name Size (amino acids) Function
Saccharomyces cerevisiae Cdk1 Cdc28 298 All cell-cycle stages
Schizosaccharomyces pombe Cdk1 Cdc2 297 All cell-cycle stages
Drosophila melanogaster Cdk1 Cdc2 297 M
Cdk2 Cdc2c 314 G1/S, S, possibly M
Cdk4 Cdk4/6 317 G1, promotes growth
Xenopus laevis Cdk1 Cdc2 301 M
Cdk2 297 S, possibly M
Homo sapiens Cdk1 Cdc2 297 M
Cdk2 298 G1, S, possibly M
Cdk4 301 G1
Cdk6 326 G1

A list of CDKs with their regulator protein, cyclin or other:

    cyclin A, cyclin B cyclin A, cyclin E
  • CDK3 cyclin C cyclin D1, cyclin D2, cyclin D3 CDK5R1, CDK5R2. See also CDKL5. cyclin D1, cyclin D2, cyclin D3 cyclin H
  • CDK8 cyclin C cyclin T1, cyclin T2a, cyclin T2b, cyclin K
  • CDK10
  • CDK11 (CDC2L2) cyclin L cyclin L
  • CDK13 (CDC2L5) cyclin L

CDK levels remain relatively constant throughout the cell cycle and most regulation is post-translational. Most knowledge of CDK structure and function is based on CDKs of S. pombe (Cdc2), S. cerevisiae (CDC28), and vertebrates (CDC2 and CDK2). The four major mechanisms of CDK regulation are cyclin binding, CAK phosphorylation, regulatory inhibitory phosphorylation, and binding of CDK inhibitory subunits (CKIs). [5]

Cyclin binding Edit

The active site, or ATP-binding site, of all kinases is a cleft between a small amino-terminal lobe and a larger carboxy-terminal lobe. [1] The structure of human Cdk2 revealed that CDKs have a modified ATP-binding site that can be regulated by cyclin binding. [1] Phosphorylation by CDK-activating kinase (CAK) at Thr 161 on the T-loop increases the complex activity. Without cyclin, a flexible loop called the activation loop or T-loop blocks the cleft, and the position of several key amino acid residues is not optimal for ATP-binding. [1] With cyclin, two alpha helices change position to permit ATP binding. One of them, the L12 helix that comes just before the T-loop in the primary sequence, becomes a beta strand and helps rearrange the T-loop, so it no longer blocks the active site. [1] The other alpha helix called the PSTAIRE helix rearranges and helps change the position of the key amino acid residues in the active site. [1]

There is considerable specificity in which cyclin binds with CDK. [3] Furthermore, cyclin binding determines the specificity of the cyclin-CDK complex for particular substrates. [3] Cyclins can directly bind the substrate or localize the CDK to a subcellular area where the substrate is found. Substrate specificity of S cyclins is imparted by the hydrophobic batch (centered on the MRAIL sequence), which has affinity for substrate proteins that contain a hydrophobic RXL (or Cy) motif. Cyclin B1 and B2 can localize Cdk1 to the nucleus and the Golgi, respectively, through a localization sequence outside the CDK-binding region. [1]

Phosphorylation Edit

Full kinase activity requires an activating phosphorylation on a threonine adjacent to the CDK's active site. [1] The identity of the CDK-activating kinase (CAK) that performs this phosphorylation varies across the model organisms. [1] The timing of this phosphorylation varies as well. In mammalian cells, the activating phosphorylation occurs after cyclin binding. [1] In yeast cells, it occurs before cyclin binding. [1] CAK activity is not regulated by known cell-cycle pathways and cyclin binding is the limiting step for CDK activation. [1]

Unlike activating phosphorylation, CDK inhibitory phosphorylation is vital for regulation of the cell cycle. Various kinases and phosphatases regulate their phosphorylation state. One of the kinases that place the tyrosine phosphate is Wee1, a kinase conserved in all eukaryotes. [1] Fission yeast also contains a second kinase Mik1 that can phosphorylate the tyrosine. [1] Vertebrates contain a different second kinase called Myt1 that is related to Wee1 but can phosphorylate both the threonine and the tyrosine. [1] Phosphatases from the Cdc25 family dephosphorylate both the threonine and the tyrosine. [1]

CDK inhibitors Edit

A cyclin-dependent kinase inhibitor (CKI) is a protein that interacts with a cyclin-CDK complex to block kinase activity, usually during G1 or in response to signals from the environment or from damaged DNA. [1] In animal cells, there are two major CKI families: the INK4 family and the CIP/KIP family. [1] The INK4 family proteins are strictly inhibitory and bind CDK monomers. Crystal structures of CDK6-INK4 complexes show that INK4 binding twists the CDK to distort cyclin binding and kinase activity. The CIP/KIP family proteins bind both the cyclin and the CDK of a complex and can be inhibitory or activating. CIP/KIP family proteins activate cyclin D and CDK4 or CDK6 complexes by enhancing complex formation. [1]

In yeast and Drosophila, CKIs are strong inhibitors of S- and M-CDK, but do not inhibit G1/S-CDKs. During G1, high levels of CKIs prevent cell cycle events from occurring out of order, but do not prevent transition through the Start checkpoint, which is initiated through G1/S-CDKs. Once the cell cycle is initiated, phosphorylation by early G1/S-CDKs leads to destruction of CKIs, relieving inhibition on later cell cycle transitions. In mammalian cells, the CKI regulation works differently. Mammalian protein p27 (Dacapo in Drosophila) inhibits G1/S- and S-CDKs, but does not inhibit S- and M-CDKs. [1]

Based on molecular docking results, Ligands-3, 5, 14, and 16 were screened among 17 different Pyrrolone-fused benzosuberene compounds as potent and specific inhibitors without any cross-reactivity against different CDK isoforms. Analysis of MD simulations and MM-PBSA studies, revealed the binding energy profiles of all the selected complexes. Selected ligands performed better than the experimental drug candidate (Roscovitine). Ligands-3 and 14 show specificity for CDK7 and Ligands-5 and 16 were specific against CDK9. These ligands are expected to possess lower risk of side effects due to their natural origin. [6]

Interpretation of dynamic simulations and binding free energy studies unveiled that Ligand2 (Out of 17 in-house synthesized pyrrolone-fused benzosuberene (PBS) compounds) has a stable and equivalent free energy to Flavopiridol, SU9516, and CVT-313 inhibitors. Ligand2 scrutinized as a selective inhibitor of CDK2 without off-target binding (CDK1 and CDK9) based on ligand efficiency and binding affinity. [7]

Suk1 or Cks Edit

The CDKs directly involved in the regulation of the cell cycle associate with small, 9- to 13-kiloDalton proteins called Suk1 or Cks. [3] These proteins are required for CDK function, but their precise role is unknown. [3] Cks1 binds the carboxy lobe of the CDK, and recognizes phosphorylated residues. It may help the cyclin-CDK complex with substrates that have multiple phosphorylation sites by increasing affinity for the substrate. [3]

Non-cyclin activators Edit

Viral cyclins Edit

Viruses can encode proteins with sequence homology to cyclins. One much-studied example is K-cyclin (or v-cyclin) from Kaposi sarcoma herpes virus (see Kaposi’s sarcoma), which activates CDK6. Viral cyclin-CDK complexes have different substrate specificities and regulation sensitivities. [8]

CDK5 activators Edit

The proteins p35 and p39 activate CDK5. Although they lack cyclin sequence homology, crystal structures show that p35 folds in a similar way as the cyclins. However, activation of CDK5 does not require activation loop phosphorylation. [8]

RINGO/Speedy Edit

Proteins with no homology to the cyclin family can be direct activators of CDKs. [9] One family of such activators is the RINGO/Speedy family, [9] which was originally discovered in Xenopus. All five members discovered so far directly activate Cdk1 and Cdk2, but the RINGO/Speedy-CDK2 complex recognizes different substrates than cyclin A-CDK2 complex. [8]

Leland H. Hartwell, R. Timothy Hunt, and Paul M. Nurse received the 2001 Nobel Prize in Physiology or Medicine for their complete description of cyclin and cyclin-dependent kinase mechanisms, which are central to the regulation of the cell cycle.

CDKs are considered a potential target for anti-cancer medication. If it is possible to selectively interrupt the cell cycle regulation in cancer cells by interfering with CDK action, the cell will die. At present, some CDK inhibitors such as seliciclib are undergoing clinical trials. Although it was originally developed as a potential anti-cancer drug, seliciclib has also proven to induce apoptosis in neutrophil granulocytes, which mediate inflammation. [10] This means that novel drugs for treatment of chronic inflammation diseases such as arthritis and cystic fibrosis could be developed.

Flavopiridol (alvocidib) is the first CDK inhibitor to be tested in clinical trials after being identified in an anti-cancer agent screen in 1992. It competes for the ATP site of the CDKs. [11] Palbociclib and abemaciclib have been approved for the management of hormone receptor (estrogen receptor/progestogen receptor) expressing metastatic breast cancer in combination with endocrine therapy. [12] [13]

More research is required, however, because disruption of the CDK-mediated pathway has potentially serious consequences while CDK inhibitors seem promising, it has to be determined how side-effects can be limited so that only target cells are affected. As such diseases are currently treated with glucocorticoids, which have often serious side-effects, even a minor success would be an improvement. [12]

Complications of developing a CDK drug include the fact that many CDKs are not involved in the cell cycle, but other processes such as transcription, neural physiology, and glucose homeostasis. [14]


Names of different cyclins - Biology

Cell division is mediated by a large number of discrete biochemical events, which must be highly coordinated. The mechanics and timing of cell division is described in terms of the cell cycle. The stages of the cell cycle are G1 (first period of growth), S (DNA replication), G2 (second period of growth), Mitosis (chromosome separation), and Cytokinesis (cell division). These timing of these events are orchestrated by the expression of cyclins. Cyclins bind and activate CDKs (cyclin dependent kinases) that phosphorylate several structural proteins and enzymes, triggering different cell cycle events. Checkpoints monitor the progress of cell cycle events and prevent entry into subsequent stages until the current stage is completed. The mitotic spindle ensures complete and accurate separation of chromosomes.

Summary

  • The cell cycle proceeds in 5 ordered stages, G1, S, G2, M, and cytokinesis, with critical cellular events triggered in each stage.
  • There are four classes of cyclins expressed at different stages of the cell cycle that activate cyclin-dependent kinases (CDKs) which drive cell cycle events.
  • Three checkpoints monitor the progress of the cell cycle and regulate the activity of cyclin-CDK complexes.
  • The cell ensures that chromosomes are replicated only once by destroying the disabling the proteins that initiate DNA replication.
  • Motor proteins and microtubules drive separation of chromosome through the mitotic spindle.

Introduction

To divide, cells must copy their cellular material (organelles, membranes, structural proteins, DNA) and then partition that material between the two daughter cells. Most material is present in enough copies that both cells are ensured of receiving an adequate supply. DNA is the exception, as only one copy of each chromosome is made. Because it is critical that both cells receive a complete set of chromosomes, the cell expends considerable energy partitioning the chromosomes.

The cell cycle is divided into stages based on the several critical cellular events. In G1 cells prepare for division by increasing in size. In S, cell replicate their DNA and centrosomes. G2 is another potential growth phase. During mitosis, the chromosomes separate, and in cytokinesis the plasma membrane constricts between the separated chromosomes, giving rise to two cells. G0 is a stage when cells exit the cycle and are considered quiescent. Differentiated cells are thought to exist in G0. To produce two viable cells, the stages of the cell cycle must proceed in order G1 -> S -> G2 -> M -> cytokineses.

Cyclins and Cyclin-Dependent Kinases (CDKs)

CDKs phosphorylate hundreds of proteins to initiate the different cell cycle events. Mammalian cells express several CDKs that function in different stages of the cell cycle: CDK4 and CDK6 (G1), CDK2 (G1/S, and S), CDK1 (mitosis). The activity of CDKs is regulated by several mechanism. Most important is the binding to cyclin. Cyclin binding is required to activate CDKs and different cyclins are expressed at different stages of the cell cycle: cyclin D (G1), cyclin E (G1/S), cyclin A (S), cyclin B (mitosis). The level of cyclin during each stage of the cell cycle is regulated through gene expression and proteolysis. Gene expression of a cyclin is turned just before its stage of the cell cycle, and once its stage is complete, the cyclin is ubiquitinated and targeted for degradation by the proteosome. In addition to activating CDKs, cyclins also target CDKs to their substrates.

Activation of CDKs

Cyclins activate CDKs by inducing a conformation change in CDKs that open an ATP-binding pocket. Peptide sequences in CDK substrates can then gain access to the pocket and be phosphorylated.

CDK activity is also regulated by phosphorylation. CDK-activating kinases phoshporylate CDKs and increase the kinase activity. In contrast, Wee1 phosphorylates CDKs in the ATP-binding pocket, reducing the activity of CDKs. The phosphatase Cdc25 removes the phosphate in the pocket restoring kinase activity of CDKs.

A positive feedback loop regulates the activation of cyclin-CDK. Active cyclin-CDK complexes activate Cdc25 and inactivate Wee1. Increased Cdc25 activity and decreased Wee1 activity generate large amounts of active cyclin-CDK. The positive feedback loop creates a switch-like change in cyclin-CDK activity in which cyclin-CDK activity is converted from low to high very quickly. Signals that initiate a stage in the cell cycle appear to activate Cdc25. Cdc25 turns on a small amount of cyclin-CDK that through the positive feedback loop activate a large amount of cyclin-CDK.

Ensuring a Single Round of DNA Replication

Cells must duplicate their DNA before dividing but they must do so only once because cells with multiple chromosomes are highly unstable and often become cancerous. Cells ensure that their DNA is replicated only once by destroying the machinery that initiates DNA replication after replication has started. DNA replication is initiated by a prereplication complex that assembles on chromosomes at the origins of replication. The prereplication complex consist of several sets of proteins including ORC and Cdc6. During S phase, cyclinA-CDK2 phosphorylate several proteins that bind the prereplication complex and start DNA replication. CyclinA-CDK2 also phoshporylate ORC to inactivate it and Cdc6 that targets it for degradation. Without these prereplication factors DNA replication cannot be initiated again.

After replication, the sister chromatids are held together by a complex of proteins called cohesins. Cohesins form a ring around the sister chromatids and prevent their premature separation. After all chromosomes are attached to the mitotic spindle, cohesins are degraded allowing the sister chromatids to separate.

Mitosis

During mitosis, the duplicated chromosomes are separated to opposite sides of the cell. A spindle composed of microtubules and molecular motors positions the chromosomes in the center of the cell. Microtubules attach to chromosomes at the centromere and then through a combination of microtubule depolymerization and motor protein activity, the chromosomes are pulled to opposite sides of the cell.

Stages of Mitosis

Mitosis is divided into several easily observable stages.

  • Prophase -> chomosome condensation, centrosomes separate to opposite sides of the cell.
  • Prometaphase -> nuclear envelope breaksdown, microtubules start to attach to chromosomes.
  • Metaphase -> Chromosomes aligned in the center of the spindle.
  • Anaphase -> Chromatids separate, spindle elongates.
  • Telophase -> Nucleus reforms around chromosomes.

Assembly of the Mitotic Spindle

As the centrosomes separate they nucleate the formation of two types of microtubules. Astral microtubules radiate toward the plasma membrane and dynein at the the plasma membrane pulls on these microtubules to separate the centrosomes and elongate the spindle during anaphase. Another set of microtubules radiates toward the opposite centrosome. Some microtubules will attach to the chromosomes to become kinetichore microtubules whereas others will overlap microtubules coming from the opposite centrosome to become interpolar microtubules. Kinesins on the microtubules help position the spindle and generate attachments between chromosomes and microtubules. During anaphase kinesins will push apart the spindle to elongate it.

Chromosome Separation

All chromosomes must attach to microtubules before separation begins. Separation is initiated when mitotic cyclin and the cohesins that surround the chromosomes is degraded. Cyclin and cohesin are marked for destruction by the anaphase promoting complex (APC). The spindle checkpoint ensures that the APC does not tag cyclin and cohesin until all chromosome are attached to microtubules. Cdc20 targets APC to mitotic cyclin and cohesin, and kinetichores on the chromosomes that are not attached to microtubules keep Cdc20 in an inactive state.

After all chromosomes are attached to microtubules, separation of the sister chromatids begins and is generated by two mechanisms. First, depolymerization of kinetichore microtubules generates force to separate the chromatids. Second, kinesins on interpolar microtubules start elongating the spindle which further separates the chromatids.

Cytokinesis

Two divide a single cell into two, the plasma membrane must be sealed between the two sets of chromosomes. Most animal cells accomplish this by constricting the plasma membrane between the chromosomes. The constriction is generated by actin and myosin filaments. Bipolar myosin filaments pull on actin filaments attached to the plasma membrane bringing the plasma membrane from opposite sides of the cell closer. Eventually the plasma membrane will fuse to separate the two cells.


Mitosis (With Diagram) | Cell Cycle

A proteinaceous factor termed as maturation promoting factor (MPF), subsequently renamed as mitosis promoting factor, is identified and purified (Masui, Markert, Mailer). MPF is a heterodimer complex between kinase – a catalytic sub- unit and cyclin – a regulatory subunit (Fig. 5.27A).

Kinase enzyme brings about phosphory­lation of specific target proteins at different stages of the cell cycle which is necessary for its pro­gression. Cyclin regulates the kinase activity by binding to it. In the absence of cyclin, the kinas­es are inactive and are, therefore, called cyclin dependent kinases (Cdks).

Cyclin confers basal kinase activity to the Cdk due to conformational changes. A number of Cdks are known — Cdk 1 to Cdk 7. Different members of the cyclin family appear at different points of the cell cycle — G1 cyclins, S phase cyclins and mitotic (M) cyclins.

Suppression of M-Cdk activity after mitosis causes the cell to enter into G1 phase for cell growth. Exit from mitosis is initiated by the inactivation of M-Cdk through ubiquitin dependent M-cyclin degradation. Proteolysis of cyclin through ubiquitination is triggered by APC which is activated by Cdc20 and Hct1 protein (Fig. 5.28A).

Accumulation of new M-cyclin for reactivation of Cdk to initiate mitosis requires a specific time and is thus delayed to cause the entry of cell into G1 phase. Suppression of Cdk activity also occurs through increased production of Cdk inhibitor protein (CKI) e.g. Sic1 (Fig. 5.28B) or by decreased tran­scription of M-cyclin gene.

Initiation of S Phase:

Escape from stable G1 occurs through the accumulation of G1-cyclins leading to the G1-Cdk activity. G1-Cdk triggers the transcription of S-cyclin genes mediated through E2F regulatory protein and synthesizes S-cyclin. Thus S-Cdk activity resumes to cause the cell to enter into S phase (Fig. 5.29).

S-Cdk initiates DNA replication at ORC, complexed with Cdc6 and MCM proteins. S-Cdk triggers origin firing, assembly of DNA polymerase and other replication proteins and activates the DNA helicases to initiate DNA replication. Disso­ciation of Cdc6 from ORC and export of Mcm from nucleus terminates replication.

Passage through G2 Phase:

After completion of DNA replication in S phase, accumulation of M-cyclins promotes gradual accumulation of M-Cdk complex. But the M-Cdk complex remains inactive due to kinase activity of Wee1 on tyro­sine residue where phosphorylation is inhibitory. At the end of G2, inactive M-Cdk is present in large amount.

At the late G1, the phos­phatase activity of Cdc25 to cause dephosphorylation of tyrosine residue and Cak mediated phosphorylation of threonine residue make the M-Cdk fully active. Cdc25 is activated by polo- kinase which results in partial activation of M- Cdk complex. M-Cdk inhibits Wee1 activity and activates Cdc25 in a positive feed-back manner. Thus by a chain of reactions, M-Cdk complexes are being fully activated and the cells enter into mitosis (Fig. 5.30).

Active MPF induces chromosome conden­sation, nuclear envelope breakdown and assembly of spindle. MPF also activates enzymes that conjugate ubiquitin to cyclin leading to cyclin degradation which renders MPF inactive.

Cyclin degradation and MPF inactivation lead to chromosome segregation, chromosome de-condensation, nuclear reforma­tion and cytokinesis. The number of Cdks and cyclins involved in cell cycle regulation varies from species to species. Moreover, other factors such as proteinaceous factors, exogenic and endogenic substances have been implicated in the control of cell cycle progression in higher plants and animals.


10.3 Control of the Cell Cycle

In this section, you will explore the following questions:

  • What are examples of internal and external mechanisms that control the cell cycle?
  • What molecules are involved in controlling the cell cycle through positive and negative regulation?

Connection for AP ® Courses

Each step of the cell cycle is closely monitored by external signals and internal controls called checkpoints. There are three major checkpoints in the cell cycle: one near the end of G1, a second at the G2/M transition, and the third during metaphase. Growth factor proteins arriving at the dividing cell’s plasma membrane can trigger the cell to begin dividing. Cyclins and cyclin-dependent kinases (Cdks) are internal molecular signals that regulate cell transitions through the various checkpoints. Passage through the G1 checkpoint makes sure that the cell is ready for DNA replication in the S stage of interphase passage through the G2 checkpoint triggers the separation of chromatids during mitosis. Positive regulator molecules like the cyclins and Cdks allow the cell cycle to advance to the next stage negative regulator molecules, such as tumor suppressor proteins, monitor cellular conditions and can halt the cycle until specific requirements are met. Errors in the regulation of the cell cycle can cause cancer, which is characterized by uncontrolled cell division.

Information presented and the examples highlighted in the section support concepts and Learning Objectives outlined in Big Idea 3 of the AP ® Biology Curriculum Framework, as shown in the tables. The Learning Objectives listed in the Curriculum Framework provide a transparent foundation for the AP ® Biology course, an inquiry-based laboratory experience, instructional activities, and AP ® exam questions. A Learning Objective merges required content with one or more of the seven Science Practices.

Big Idea 3 Living systems store, retrieve, transmit and respond to information essential to life processes.
Enduring Understanding 3.A Heritable information provides for continuity of life.
Essential Knowledge 3.A.2 In eukaryotes, heritable information is passed to the next generation via processes that include the cell cycle and mitosis or meiosis plus fertilization.
Science Practice 6.4 The student can make claims and predictions about natural phenomena based on scientific theories and models.
Learning Objective 3.7 The student can make predictions about natural phenomena occurring during the cell cycle.
Essential Knowledge 3.A.2 In eukaryotes, heritable information is passed to the next generation via processes that include the cell cycle and mitosis or meiosis plus fertilization.
Science Practice 1.2 The student can describe representations and models of natural or man-made phenomena and systems in the domain.
Learning Objective 3.8 The student can describe the events that occur in the cell cycle.

Teacher Support

Introduce the topic of control of the cell cycle using visuals such as this video.

The length of the cell cycle is highly variable, even within the cells of a single organism. In humans, the frequency of cell turnover ranges from a few hours in early embryonic development, to an average of two to five days for epithelial cells, and to an entire human lifetime spent in G0 by specialized cells, such as cortical neurons or cardiac muscle cells. There is also variation in the time that a cell spends in each phase of the cell cycle. When fast-dividing mammalian cells are grown in culture (outside the body under optimal growing conditions), the length of the cycle is about 24 hours. In rapidly dividing human cells with a 24-hour cell cycle, the G1 phase lasts approximately nine hours, the S phase lasts 10 hours, the G2 phase lasts about four and one-half hours, and the M phase lasts approximately one-half hour. In early embryos of fruit flies, the cell cycle is completed in about eight minutes. The timing of events in the cell cycle is controlled by mechanisms that are both internal and external to the cell.

Regulation of the Cell Cycle by External Events

Both the initiation and inhibition of cell division are triggered by events external to the cell when it is about to begin the replication process. An event may be as simple as the death of a nearby cell or as sweeping as the release of growth-promoting hormones, such as human growth hormone (HGH). A lack of HGH can inhibit cell division, resulting in dwarfism, whereas too much HGH can result in gigantism. Crowding of cells can also inhibit cell division. Another factor that can initiate cell division is the size of the cell as a cell grows, it becomes inefficient due to its decreasing surface-to-volume ratio. The solution to this problem is to divide.

Whatever the source of the message, the cell receives the signal, and a series of events within the cell allows it to proceed into interphase. Moving forward from this initiation point, every parameter required during each cell cycle phase must be met or the cycle cannot progress.

Regulation at Internal Checkpoints

It is essential that the daughter cells produced be exact duplicates of the parent cell. Mistakes in the duplication or distribution of the chromosomes lead to mutations that may be passed forward to every new cell produced from an abnormal cell. To prevent a compromised cell from continuing to divide, there are internal control mechanisms that operate at three main cell cycle checkpoints . A checkpoint is one of several points in the eukaryotic cell cycle at which the progression of a cell to the next stage in the cycle can be halted until conditions are favorable. These checkpoints occur near the end of G1, at the G2/M transition, and during metaphase (Figure 10.11).

The G1 Checkpoint

The G1 checkpoint determines whether all conditions are favorable for cell division to proceed. The G1 checkpoint, also called the restriction point (in yeast), is a point at which the cell commits to the cell division process. External influences, such as growth factors, play a large role in carrying the cell past the G1 checkpoint. In addition to adequate reserves and cell size, there is a check for genomic DNA damage at the G1 checkpoint. A cell that does not meet all the requirements will not be allowed to progress into the S phase. The cell can halt the cycle and attempt to remedy the problematic condition, or the cell can advance into G0 and await further signals when conditions improve.

The G2 Checkpoint

The G2 checkpoint bars entry into the mitotic phase if certain conditions are not met. As at the G1 checkpoint, cell size and protein reserves are assessed. However, the most important role of the G2 checkpoint is to ensure that all of the chromosomes have been replicated and that the replicated DNA is not damaged. If the checkpoint mechanisms detect problems with the DNA, the cell cycle is halted, and the cell attempts to either complete DNA replication or repair the damaged DNA.

The M Checkpoint

The M checkpoint occurs near the end of the metaphase stage of karyokinesis. The M checkpoint is also known as the spindle checkpoint, because it determines whether all the sister chromatids are correctly attached to the spindle microtubules. Because the separation of the sister chromatids during anaphase is an irreversible step, the cycle will not proceed until the kinetochores of each pair of sister chromatids are firmly anchored to at least two spindle fibers arising from opposite poles of the cell.

Link to Learning

Watch what occurs at the G1, G2, and M checkpoints by visiting this website to see an animation of the cell cycle.

  1. Failure in spindle checkpoint results in the formation of one gamete cell with two extra chromosomes and another gamete cell lacking chromosomes.
  2. Failure in spindle checkpoint yields the same number of chromosomes in each gamete cell.
  3. Failure in spindle checkpoint will form two gamete cells without any chromosomes.
  4. Failure in spindle checkpoint results in the formation of one gamete cell with an extra chromosome and another gamete cell lacking a chromosome.

Regulator Molecules of the Cell Cycle

In addition to the internally controlled checkpoints, there are two groups of intracellular molecules that regulate the cell cycle. These regulatory molecules either promote progress of the cell to the next phase (positive regulation) or halt the cycle (negative regulation). Regulator molecules may act individually, or they can influence the activity or production of other regulatory proteins. Therefore, the failure of a single regulator may have almost no effect on the cell cycle, especially if more than one mechanism controls the same event. Conversely, the effect of a deficient or non-functioning regulator can be wide-ranging and possibly fatal to the cell if multiple processes are affected.

Positive Regulation of the Cell Cycle

Two groups of proteins, called cyclins and cyclin-dependent kinases (Cdks), are responsible for the progress of the cell through the various checkpoints. The levels of the four cyclin proteins fluctuate throughout the cell cycle in a predictable pattern (Figure 10.12). Increases in the concentration of cyclin proteins are triggered by both external and internal signals. After the cell moves to the next stage of the cell cycle, the cyclins that were active in the previous stage are degraded.

Cyclins regulate the cell cycle only when they are tightly bound to Cdks. To be fully active, the Cdk/cyclin complex must also be phosphorylated in specific locations. Like all kinases, Cdks are enzymes (kinases) that phosphorylate other proteins. Phosphorylation activates the protein by changing its shape. The proteins phosphorylated by Cdks are involved in advancing the cell to the next phase. (Figure 10.13). The levels of Cdk proteins are relatively stable throughout the cell cycle however, the concentrations of cyclin fluctuate and determine when Cdk/cyclin complexes form. The different cyclins and Cdks bind at specific points in the cell cycle and thus regulate different checkpoints.

Since the cyclic fluctuations of cyclin levels are based on the timing of the cell cycle and not on specific events, regulation of the cell cycle usually occurs by either the Cdk molecules alone or the Cdk/cyclin complexes. Without a specific concentration of fully activated cyclin/Cdk complexes, the cell cycle cannot proceed through the checkpoints.

Although the cyclins are the main regulatory molecules that determine the forward momentum of the cell cycle, there are several other mechanisms that fine-tune the progress of the cycle with negative, rather than positive, effects. These mechanisms essentially block the progression of the cell cycle until problematic conditions are resolved. Molecules that prevent the full activation of Cdks are called Cdk inhibitors. Many of these inhibitor molecules directly or indirectly monitor a particular cell cycle event. The block placed on Cdks by inhibitor molecules will not be removed until the specific event that the inhibitor monitors is completed.

Negative Regulation of the Cell Cycle

The second group of cell cycle regulatory molecules are negative regulators. Negative regulators halt the cell cycle. Remember that in positive regulation, active molecules cause the cycle to progress.

The best understood negative regulatory molecules are retinoblastoma protein (Rb) , p53 , and p21 . Retinoblastoma proteins are a group of tumor-suppressor proteins common in many cells. The 53 and 21 designations refer to the functional molecular masses of the proteins (p) in kilodaltons. Much of what is known about cell cycle regulation comes from research conducted with cells that have lost regulatory control. All three of these regulatory proteins were discovered to be damaged or non-functional in cells that had begun to replicate uncontrollably (became cancerous). In each case, the main cause of the unchecked progress through the cell cycle was a faulty copy of the regulatory protein.

Rb, p53, and p21 act primarily at the G1 checkpoint. p53 is a multi-functional protein that has a major impact on the commitment of a cell to division because it acts when there is damaged DNA in cells that are undergoing the preparatory processes during G1. If damaged DNA is detected, p53 halts the cell cycle and recruits enzymes to repair the DNA. If the DNA cannot be repaired, p53 can trigger apoptosis, or cell death, to prevent the duplication of damaged chromosomes. As p53 levels rise, the production of p21 is triggered. p21 enforces the halt in the cycle dictated by p53 by binding to and inhibiting the activity of the Cdk/cyclin complexes. As a cell is exposed to more stress, higher levels of p53 and p21 accumulate, making it less likely that the cell will move into the S phase.

Rb exerts its regulatory influence on other positive regulator proteins. Chiefly, Rb monitors cell size. In the active, dephosphorylated state, Rb binds to proteins called transcription factors, most commonly, E2F (Figure 10.14). Transcription factors “turn on” specific genes, allowing the production of proteins encoded by that gene. When Rb is bound to E2F, production of proteins necessary for the G1/S transition is blocked. As the cell increases in size, Rb is slowly phosphorylated until it becomes inactivated. Rb releases E2F, which can now turn on the gene that produces the transition protein, and this particular block is removed. For the cell to move past each of the checkpoints, all positive regulators must be “turned on,” and all negative regulators must be “turned off.”


Regulation at Internal Checkpoints

It is essential that the daughter cells produced be exact duplicates of the parent cell. Mistakes in the duplication or distribution of the chromosomes lead to mutations that may be passed forward to every new cell produced from an abnormal cell. To prevent a compromised cell from continuing to divide, there are internal control mechanisms that operate at three main cell cycle checkpoints. A checkpoint is one of several points in the eukaryotic cell cycle at which the progression of a cell to the next stage in the cycle can be halted until conditions are favorable. These checkpoints occur near the end of G1, at the G2/M transition, and during metaphase (Figure 1).

Figure 1. The cell cycle is controlled at three checkpoints. The integrity of the DNA is assessed at the G1 checkpoint. Proper chromosome duplication is assessed at the G2 checkpoint. Attachment of each kinetochore to a spindle fiber is assessed at the M checkpoint.

The G1 Checkpoint

The G1 checkpoint determines whether all conditions are favorable for cell division to proceed. The G1 checkpoint, also called the restriction point (in yeast), is a point at which the cell irreversibly commits to the cell division process. External influences, such as growth factors, play a large role in carrying the cell past the G1 checkpoint. In addition to adequate reserves and cell size, there is a check for genomic DNA damage at the G1 checkpoint. A cell that does not meet all the requirements will not be allowed to progress into the S phase. The cell can halt the cycle and attempt to remedy the problematic condition, or the cell can advance into G0 and await further signals when conditions improve.

The G2 Checkpoint

The G2 checkpoint bars entry into the mitotic phase if certain conditions are not met. As at the G1 checkpoint, cell size and protein reserves are assessed. However, the most important role of the G2 checkpoint is to ensure that all of the chromosomes have been replicated and that the replicated DNA is not damaged. If the checkpoint mechanisms detect problems with the DNA, the cell cycle is halted, and the cell attempts to either complete DNA replication or repair the damaged DNA.

The M Checkpoint

The M checkpoint occurs near the end of the metaphase stage of karyokinesis. The M checkpoint is also known as the spindle checkpoint, because it determines whether all the sister chromatids are correctly attached to the spindle microtubules. Because the separation of the sister chromatids during anaphase is an irreversible step, the cycle will not proceed until the kinetochores of each pair of sister chromatids are firmly anchored to at least two spindle fibers arising from opposite poles of the cell.

Link to Learning


A. Discovery and Characterization of Maturation Promoting Factor (MPF)

Growing, dividing cells monitor their progress through the phases. Cells produce internal chemical signals that tell them when it&rsquos time to begin replication or mitosis, or even when to enter into G0 when they reach their terminally differentiated state. The experiment that first demonstrated a chemical regulator of the cell cycle involved fusing very large frog&rsquos eggs! The experiment is described below.

The hypothesis tested here was that frog oocyte cytoplasm from germinal vesicle stage oocytes (i.e., in mid-meiosis) contains a chemical that caused the cell to lose its nuclear membrane, condense its chromatin into chromosomes and enter meiosis. Cytoplasm was withdrawn from one of these mid-meiotic oocytes with a fine hypodermic needle, and then injected into a pre-meiotic oocyte. The mid-meiotic oocyte cytoplasm induced premature meiosis in the immature oocyte. A maturation promoting factor (MPF) could be isolated from the mid-meiotic cells and injected into pre-meiotic cells it caused them to enter meiosis. MPF turns out to be a protein kinase made up of two polypeptide subunits as shown below.

MPF was then also shown to stimulate somatic cells in G2to enter premature mitosis. So conveniently, MPF can also be Mitosis Promoting Factor! Hereafter we will discuss the effects of MPF as being equivalent in mitosis and meiosis. When active, MPF targets many cellular proteins.

Assays of MPF activity as well as the actual levels of the two subunits over time during the cell cycle are graphed below.

One subunit of MPF is cyclin, a regulatory polypeptide. The other subunit, cyclin-dependent kinase (cdk), contains the kinase enzyme active site. Both subunits must be bound to make an active kinase. Cyclin was so-named because its levels rise gradually after cytokinesis, peak at the next mitosis, and then fall. Levels of the cdk subunit do not change significantly during the life of the cell. Because the kinase activity of MPF requires cyclin, it tracks the rise in cyclin near the end of the G2, and its fall after mitosis. Cyclin begins to accumulate in G1, rising gradually and binding to more and more cdk subunits. MPF reaches a threshold concentration in G2 that triggers entry into mitosis. For their discovery of these central molecules Leland H. Hartwell, R. Timothy Hunt, and Paul M. Nurse won the 2001 Nobel Prize in Physiology or Medicine.


Biology 171

By the end of this section, you will be able to do the following:

  • Understand how the cell cycle is controlled by mechanisms that are both internal and external to the cell
  • Explain how the three internal “control checkpoints” occur at the end of G1, at the G2/M transition, and during metaphase
  • Describe the molecules that control the cell cycle through positive and negative regulation

The length of the cell cycle is highly variable, even within the cells of a single organism. In humans, the frequency of cell turnover ranges from a few hours in early embryonic development, to an average of two to five days for epithelial cells, and to an entire human lifetime spent in G0 by specialized cells, such as cortical neurons or cardiac muscle cells.

There is also variation in the time that a cell spends in each phase of the cell cycle. When rapidly dividing mammalian cells are grown in a culture (outside the body under optimal growing conditions), the length of the cell cycle is about 24 hours. In rapidly dividing human cells with a 24-hour cell cycle, the G1 phase lasts approximately nine hours, the S phase lasts 10 hours, the G2 phase lasts about four and one-half hours, and the M phase lasts approximately one-half hour. By comparison, in fertilized eggs (and early embryos) of fruit flies, the cell cycle is completed in about eight minutes. This is because the nucleus of the fertilized egg divides many times by mitosis but does not go through cytokinesis until a multinucleate “zygote” has been produced, with many nuclei located along the periphery of the cell membrane, thereby shortening the time of the cell division cycle. The timing of events in the cell cycle of both “invertebrates” and “vertebrates” is controlled by mechanisms that are both internal and external to the cell.

Regulation of the Cell Cycle by External Events

Both the initiation and inhibition of cell division are triggered by events external to the cell when it is about to begin the replication process. An event may be as simple as the death of nearby cells or as sweeping as the release of growth-promoting hormones, such as human growth hormone (HGH or hGH) . A lack of HGH can inhibit cell division, resulting in dwarfism, whereas too much HGH can result in gigantism. Crowding of cells can also inhibit cell division. In contrast, a factor that can initiate cell division is the size of the cell: As a cell grows, it becomes physiologically inefficient due to its decreasing surface-to-volume ratio. The solution to this problem is to divide.

Whatever the source of the message, the cell receives the signal, and a series of events within the cell allows it to proceed into interphase. Moving forward from this initiation point, every parameter required during each cell cycle phase must be met or the cycle cannot progress.

Regulation at Internal Checkpoints

It is essential that the daughter cells produced be exact duplicates of the parent cell. Mistakes in the duplication or distribution of the chromosomes lead to mutations that may be passed forward to every new cell produced from an abnormal cell. To prevent a compromised cell from continuing to divide, there are internal control mechanisms that operate at three main cell-cycle checkpoints : A checkpoint is one of several points in the eukaryotic cell cycle at which the progression of a cell to the next stage in the cycle can be halted until conditions are favorable. These checkpoints occur near the end of G1, at the G2/M transition, and during metaphase ((Figure)).


The G1 Checkpoint

The G1 checkpoint determines whether all conditions are favorable for cell division to proceed. The G1 checkpoint, also called the restriction point (in yeast), is a point at which the cell irreversibly commits to the cell division process. External influences, such as growth factors, play a large role in carrying the cell past the G1 checkpoint. In addition to adequate reserves and cell size, there is a check for genomic DNA damage at the G1 checkpoint. A cell that does not meet all the requirements will not be allowed to progress into the S phase. The cell can halt the cycle and attempt to remedy the problematic condition, or the cell can advance into G0 and await further signals when conditions improve.

The G2 Checkpoint

The G2 checkpoint bars entry into the mitotic phase if certain conditions are not met. As at the G1 checkpoint, cell size and protein reserves are assessed. However, the most important role of the G2 checkpoint is to ensure that all of the chromosomes have been replicated and that the replicated DNA is not damaged. If the checkpoint mechanisms detect problems with the DNA, the cell cycle is halted, and the cell attempts to either complete DNA replication or repair the damaged DNA.

The M Checkpoint

The M checkpoint occurs near the end of the metaphase stage of karyokinesis. The M checkpoint is also known as the spindle checkpoint, because it determines whether all the sister chromatids are correctly attached to the spindle microtubules. Because the separation of the sister chromatids during anaphase is an irreversible step, the cycle will not proceed until the kinetochores of each pair of sister chromatids are firmly anchored to at least two spindle fibers arising from opposite poles of the cell.

Watch what occurs at the G1, G2, and M checkpoints by visiting this website to see an animation of the cell cycle.

Regulator Molecules of the Cell Cycle

In addition to the internally controlled checkpoints, there are two groups of intracellular molecules that regulate the cell cycle. These regulatory molecules either promote progress of the cell to the next phase (positive regulation) or halt the cycle (negative regulation). Regulator molecules may act individually, or they can influence the activity or production of other regulatory proteins. Therefore, the failure of a single regulator may have almost no effect on the cell cycle, especially if more than one mechanism controls the same event. However, the effect of a deficient or non-functioning regulator can be wide-ranging and possibly fatal to the cell if multiple processes are affected.

Positive Regulation of the Cell Cycle

Two groups of proteins, called cyclins and cyclin-dependent kinases (Cdks), are termed positive regulators. They are responsible for the progress of the cell through the various checkpoints. The levels of the four cyclin proteins fluctuate throughout the cell cycle in a predictable pattern ((Figure)). Increases in the concentration of cyclin proteins are triggered by both external and internal signals. After the cell moves to the next stage of the cell cycle, the cyclins that were active in the previous stage are degraded by cytoplasmic enzymes, as shown in (Figure) below.


Cyclins regulate the cell cycle only when they are tightly bound to Cdks. To be fully active, the Cdk/cyclin complex must also be phosphorylated in specific locations to activate the complex. Like all kinases, Cdks are enzymes (kinases) that in turn phosphorylate other proteins. Phosphorylation activates the protein by changing its shape. The proteins phosphorylated by Cdks are involved in advancing the cell to the next phase. ((Figure)). The levels of Cdk proteins are relatively stable throughout the cell cycle however, the concentrations of cyclin fluctuate and determine when Cdk/cyclin complexes form. The different cyclins and Cdks bind at specific points in the cell cycle and thus regulate different checkpoints.


Because the cyclic fluctuations of cyclin levels are largely based on the timing of the cell cycle and not on specific events, regulation of the cell cycle usually occurs by either the Cdk molecules alone or the Cdk/cyclin complexes. Without a specific concentration of fully activated cyclin/Cdk complexes, the cell cycle cannot proceed through the checkpoints.

Although the cyclins are the main regulatory molecules that determine the forward momentum of the cell cycle, there are several other mechanisms that fine-tune the progress of the cycle with negative, rather than positive, effects. These mechanisms essentially block the progression of the cell cycle until problematic conditions are resolved. Molecules that prevent the full activation of Cdks are called Cdk inhibitors. Many of these inhibitor molecules directly or indirectly monitor a particular cell-cycle event. The block placed on Cdks by inhibitor molecules will not be removed until the specific event that the inhibitor monitors is completed.

Negative Regulation of the Cell Cycle

The second group of cell-cycle regulatory molecules are negative regulators, which stop the cell cycle. Remember that in positive regulation, active molecules cause the cycle to progress.

The best understood negative regulatory molecules are retinoblastoma protein (Rb) , p53 , and p21 . Retinoblastoma proteins are a group of tumor-suppressor proteins common in many cells. We should note here that the 53 and 21 designations refer to the functional molecular masses of the proteins (p) in kilodaltons (a dalton is equal to an atomic mass unit, which is equal to one proton or one neutron or 1 g/mol). Much of what is known about cell-cycle regulation comes from research conducted with cells that have lost regulatory control. All three of these regulatory proteins were discovered to be damaged or non-functional in cells that had begun to replicate uncontrollably (i.e., became cancerous). In each case, the main cause of the unchecked progress through the cell cycle was a faulty copy of the regulatory protein.

Rb, p53, and p21 act primarily at the G1 checkpoint. p53 is a multi-functional protein that has a major impact on the commitment of a cell to division because it acts when there is damaged DNA in cells that are undergoing the preparatory processes during G1. If damaged DNA is detected, p53 halts the cell cycle and then recruits specific enzymes to repair the DNA. If the DNA cannot be repaired, p53 can trigger apoptosis, or cell suicide, to prevent the duplication of damaged chromosomes. As p53 levels rise, the production of p21 is triggered. p21 enforces the halt in the cycle dictated by p53 by binding to and inhibiting the activity of the Cdk/cyclin complexes. As a cell is exposed to more stress, higher levels of p53 and p21 accumulate, making it less likely that the cell will move into the S phase.

Rb, which largely monitors cell size, exerts its regulatory influence on other positive regulator proteins. In the active, dephosphorylated state, Rb binds to proteins called transcription factors, most commonly, E2F ((Figure)). Transcription factors “turn on” specific genes, allowing the production of proteins encoded by that gene. When Rb is bound to E2F, production of proteins necessary for the G1/S transition is blocked. As the cell increases in size, Rb is slowly phosphorylated until it becomes inactivated. Rb releases E2F, which can now turn on the gene that produces the transition protein, and this particular block is removed. For the cell to move past each of the checkpoints, all positive regulators must be “turned on,” and all negative regulators must be “turned off.”


Rb and other proteins that negatively regulate the cell cycle are sometimes called tumor suppressors. Why do you think the name tumor suppressor might be appropriate for these proteins?

Section Summary

Each step of the cell cycle is monitored by internal controls called checkpoints. There are three major checkpoints in the cell cycle: one near the end of G1, a second at the G2/M transition, and the third during metaphase. Positive regulator molecules allow the cell cycle to advance to the next stage of cell division. Negative regulator molecules monitor cellular conditions and can halt the cycle until specific requirements are met.

Art Connections

(Figure) Rb and other proteins that negatively regulate the cell cycle are sometimes called tumor suppressors. Why do you think the name tumor suppressor might be appropriate for these proteins?

(Figure) Rb and other negative regulatory proteins control cell division and therefore prevent the formation of tumors. Mutations that prevent these proteins from carrying out their function can result in cancer.

Free Response

Describe the general conditions that must be met at each of the three main cell-cycle checkpoints.

The G1 checkpoint monitors adequate cell growth, the state of the genomic DNA, adequate stores of energy, and materials for S phase. At the G2 checkpoint, DNA is checked to ensure that all chromosomes were duplicated and that there are no mistakes in newly synthesized DNA. Additionally, cell size and energy reserves are evaluated. The M checkpoint confirms the correct attachment of the mitotic spindle fibers to the kinetochores.

Compare and contrast the roles of the positive cell-cycle regulators negative regulators.

Positive cell regulators such as cyclin and Cdk perform tasks that advance the cell cycle to the next stage. Negative regulators such as Rb, p53, and p21 block the progression of the cell cycle until certain events have occurred.

What steps are necessary for Cdk to become fully active?

Cdk must bind to a cyclin, and it must be phosphorylated in the correct position to become fully active.

Rb is a negative regulator that blocks the cell cycle at the G1 checkpoint until the cell achieves a requisite size. What molecular mechanism does Rb employ to halt the cell cycle?

Rb is active when it is dephosphorylated. In this state, Rb binds to E2F, which is a transcription factor required for the transcription and eventual translation of molecules required for the G1/S transition. E2F cannot transcribe certain genes when it is bound to Rb. As the cell increases in size, Rb becomes phosphorylated, inactivated, and releases E2F. E2F can then promote the transcription of the genes it controls, and the transition proteins will be produced.

Glossary


Results and discussion

Cyclin type influences interphase length more than cyclin level

RNAi of all three mitotic cyclins in the syncytial embryo arrests the cycle in an interphase that exhibits a prolonged S phase and, before cycle 13, uncoupled centrosome replication (McCleland and O�rrell, 2008 Farrell et al., 2012). In contrast, pairwise knockdown of cyclins allows progress to mitoses that exhibit distinctive defects depending on which cyclin remains (McCleland et al., 2009b). Here, we examine the consequence of pairwise knockdown of cyclins on interphase time.

Two of the three cyclins were removed by double-strand RNA (dsRNA) injection, and the cycle driven by the remaining cyclin was evaluated by live imaging. For example, CycB and CycB3 (abbreviated as CycB+B3 hereafter) were knocked down, and progress through the cycle with the remaining CycA ( Fig. 1 C ) was compared with control ( Fig. 1 B ). Transforming acidic coiled coil (TACC)–GFP and H2AvD-RFP were used to visualize the centrosome and nuclear cycles, respectively. Interphase duration was extended after knockdown of any pair of cyclins ( Fig. 1, C and D ). The centrosome cycle was extended in coordination with the cell cycle ( Fig. 1 C ). The results show that any single remaining cyclin is sufficient to promote mitosis 13 and a coordinated centrosome cycle but that no single cyclin can support normal interphase timing.

The prolongation of interphase upon pairwise knockdown suggested that cyclins have a role in defining interphase length. In previous work, we halved the level of the remaining cyclin by reducing gene dose in embryos with two knocked down cyclins ( Fig. 1 A ). This reduction of cyclin level had little effect on interphase timing, but it compromised execution of mitotic events (McCleland et al., 2009b). Thus, after pairwise cyclin knockdown, cyclin levels limit execution of mitosis but not the timing of mitotic entry.

In addition to the published reduction of function experiments, we wanted to test whether increasing the level of cyclin would reveal a cyclin level input into the timing of mitosis. To this end, we injected embryos with different pairwise combinations of cyclin dsRNA, and when they reached cycle 13, we injected them with a purified Drosophila GST-CycB fusion protein at one pole ( Fig. 2 A ). In embryos running on CycA alone (CycB+B3 RNAi) or embryos running on CycB3 alone (CycA+B RNAi), the injected GST-CycB accelerated progress to mitosis ( Fig. 2, C and D and Videos 2 and 3). However, in embryos running on CycB alone (CycA+B3 RNAi), the injected GST-CycB did not accelerate progress to mitosis ( Fig. 2, B and D and Video 1). Thus, injected CycB synergized with CycA or CycB3 to advance mitosis, but the same injection of CycB did not advance mitosis in embryos in which endogenous CycB was the remaining cyclin. Therefore, restoration of a second cyclin type, but not an increase in cyclin level, advanced mitosis. This experiment supports previous gene dose experiments in arguing that the level of the single remaining cyclin is not a major determinant of interphase length under the pairwise cyclin knockdown conditions. We propose that the different cyclin types have somewhat specialized activities and collaborate to promote rapid entry into mitosis. We have sought an explanation for this phenomenon and have uncovered an unexpected influence of cyclin type on cell cycle progress.

Diversity of cyclin types influences interphase length, whereas amount of a single cyclin has little effect. (A) Schematic of the experiment. dsRNA to two of the three cyclins was introduced throughout the embryo in cycle 10. Rhodamine-tagged GST-CycB protein was then injected at one pole during interphase 13. Embryos were imaged in regions 1 and 2, and the timing of mitosis in the two regions was determined. (B and C) Video frames of cycle 13 in which the remaining cyclin is CycB (B) or CycA (C). After injection of the GST-CycB protein, red fluorescence is seen in the injected pole (rhodamine). The absence of red at the other pole shows that significant GST-CycB does not reach the other pole in this time frame. Noting time stamps (minutes and seconds) on the images, it can be seen that GST-CycB does not advance mitosis when introduced into the CycB alone embryo but does advance mitosis when introduced into the CycA alone embryo. Bar, 5 µm. (D) A compilation of timing results from these experiments. Error bars represent SDs. n = 3.

Pairwise cyclin knockdown introduces a gap phase

Because S-phase duration governs interphase length in the early cycles (McCleland et al., 2009a) and we recently showed that cyclin�k1 activity shortens S phase (Farrell et al., 2012), we tested whether pairwise cyclin knockdown extends S phase. The changing distributions of proliferating cell nuclear antigen (PCNA)–GFP, which marks the sliding clamp of DNA polymerase, allowed us to visualize S phase (McCleland et al., 2009a Farrell et al., 2012). We injected pairwise combinations of cyclin dsRNA into embryos expressing H2AvD-RFP and then injected PCNA-GFP recombinant protein and imaged cells near the point of injection. In control embryos ( Fig. 3 A and Video 4), PCNA accumulated in the nucleus at the onset of interphase ( Fig. 3 A , 00:56) and then became increasingly restricted to foci that dimmed and dispersed at the end of S phase ( Fig. 3 A , 11:12). Dispersal of PCNA from the nucleus marked nuclear envelope breakdown ( Fig. 3 A , 13:37 and 15:03). Except for a delay in nuclear envelope breakdown, the PCNA dynamics showed little change upon pairwise cyclin knockdown ( Fig. 3, B𠄽 and Videos 5, 6, and 7), and the extension of S phase was small in comparison to the change in interphase ( Fig. 3 E ). Additionally, the S phase extension did not approach the near doubling of S phase caused by the triple cyclin knockdown (Farrell et al., 2012).

Interphase progression in cyclin RNAi-treated embryos. (A𠄽) Video frames of GFP-PCNA (top white) and histone-RFP (bottom red) during the thirteenth syncytial cycle. Time is given in minutes and seconds. (A) Control embryos had a short gap phase before mitotic entry (ς min, between 11:12 and 13:37 Video 1). (B𠄽) Pairwise knockdown of mitotic cyclins prolonged interphase, mainly by extending the G2 (e.g., 𢏆 min between 15:57 and 21:59 in B Video 2). (E) S-phase duration. Mild prolongations of S phase were observed in CycA+B and CycA+B3 RNAi-treated embryos as compared with the control, whereas treatment with CycB+B3 RNAi had no significant effect. (F) Gap-phase duration. All the combinations of cyclin RNAi treatment extended the gap phase. CycB+B3 RNAi was most dramatic. Error bars represent SDs. n > 3. (G and H) Alexa Fluor 546𠄽UTP (red) was incorporated into DNA when injected before but not after the dispersal of PCNA foci (green) in CycB+B3 RNAi-treated embryos. Bars, 5 µm.

Pairwise cyclin knockdown introduced a distinct pause between the completion of S phase and onset of chromosome condensation ( Fig. 3, B𠄽 and F ). To confirm that PCNA foci correctly marked S phase and that DNA replication did not extend into the “gap” phase, Alexa Fluor 546–labeled deoxy-UTP (dUTP) was injected into these cyclin RNAi-treated embryos before and after the dispersal of PCNA foci. For all knockdowns, nucleotide was incorporated before, but not after, PCNA foci dispersal (e.g., Fig. 3, G and H ), indicating that PCNA appropriately marks completion of active replication.

The finding that no pairwise cyclin knockdown gave the dramatic extension of S phase previously reported after triple cyclin knockdown (Farrell et al., 2012) leads us to conclude that each cyclin type is capable of accelerating S phase. Furthermore, we conclude that pairwise cyclin knockdown in the syncytial cycles generates a G2-like gap phase whose duration depends on which cyclins are knocked down ( Fig. 3 F ).

The induced G2 depends on the DNA replication checkpoint

Because cyclin levels are not limiting, we presumed that something else must be responsible for the G2 that is created upon knockdown of two of the three cyclins. How can this be?

We tested the contribution of S phase by deleting S phase to examine the consequence on the time of mitotic entry. Injection of the Cdt1 inhibitor Geminin blocks the licensing of replication origins, thereby deleting the subsequent S phase (McCleland et al., 2009a). As previously reported, Geminin injection into cycle 12 control embryos, which deletes S phase 13, shortened interphase 13. Geminin injection also shortened interphase 13 in the cyclin knockdown embryos, shortening it almost to the duration of interphase in embryos with a full complement of cyclins ( Fig. 4 A ). S phase deletion also attenuated the differences in interphase length among the different cyclin knockdowns. Thus, S phase influences interphase duration, even though it is ordinarily completed well before mitosis in the cyclin knockdown embryos. Furthermore, the different cyclin types drive mitosis at similar times in the absence of S phase, suggesting similar potencies to drive mitosis under these conditions.

The G2 and prolonged interphase after pairwise cyclin knockdown requires the replication checkpoint. (A) Inactivation of Chk1 (grp − ) or deletion of S phase (by Geminin injection) shortened interphase in control and cyclin RNAi-treated embryos and reduced the differences among cyclin types. Error bars represent SDs. Detailed measurements are listed in Table S1. Data for cyclin RNAi experiments in wild-type embryos are reproduced from Fig. 1 D for purpose of comparison. (B𠄾) Video frames of grp − mutant embryos in cycle 13 (GFP-PCNA is shown in white, and histone-RFP is shown in red) aligned at the start of DNA condensation (t = 00:00). Time is given in minutes and seconds. Bars, 5 µm. (F) A schematic model in which checkpoint inhibition of cyclin�k1 (gray) is reversed by compartment-specific action of cyclins plus a slow communication between compartments (arrows).

Pairwise knockdown of cyclins in embryos lacking Chk1/Grapes (embryos from grp mutant mothers) modestly extends interphase in the grp embryo ( Fig. 4 A ). This knockdown also substantially suppresses the mitosis 13 defects in grp embryos ( Figs. 4, B𠄾 and S2, A and B). The mitotic catastrophe in grp embryos has long been thought to be caused by entry into mitosis with incompletely replicated DNA. Indeed, our analysis of PCNA localization ( Figs. 4 and S2, C and D) supports a proposal that the small extension of interphase allows completion of S phase and, hence, suppression of the catastrophe. This suppression of the grp phenotype, which extends to partial restoration of gastrulation (Video 8), is reminiscent of suppression of hypomorphic mei-41 mutations when the maternal dose of CycA and/or CycB was reduced (Sibon et al., 1999). However, the most important feature of this analysis is that the extension of interphase by pairwise cyclin knockdown in grp embryos is so slight that interphase remains shorter than a wild-type interphase 13 ( Fig. 4 A ). Thus, each individual cyclin type can drive rapid advance to mitosis in the absence of functional Chk1. Furthermore, the G2 that was introduced by cyclin knockdown was absent in grp embryos ( Fig. 4, C𠄾 ), and cyclin type–specific differences in interphase length were minor ( Fig. 4 A ). These results demonstrate that mitotic entry is timed primarily by the Grapes-dependent checkpoint in cyclin knockdown embryos and, moreover, that the G2 induced in these embryos results from action of the checkpoint.

How might S phase govern the time of mitosis when mitosis begins well after completion of S phase? To test whether the S-phase checkpoint delayed accumulation of the remaining cyclin, we immunoblotted single knockdown embryos to follow accumulation in wild-type and grp embryos. Cyclin accumulated during S phase in both wild-type and grp mutant embryos (Fig. S3). Thus, Grapes function does not delay the production of cyclin. Instead, the difference between grp + and grp − embryos suggests that persistent activity of the checkpoint prevents the checkpoint-competent embryos from going into mitosis after pairwise cyclin knockdown. Because wild-type embryos enter mitosis immediately after S phase, the checkpoint is rapidly reversed when there is a full complement of cyclin types, but its reversal is delayed upon pairwise cyclin knockdown. Apparently, the different cyclin types ordinarily collaborate to rapidly reverse the checkpoint.

Compartments and cyclin specialization

Our findings show that a persistent action or consequence of the DNA replication checkpoint underlies a G2 phase that is introduced by pairwise knockdown of cyclins. One might propose that cyclin knockdown doesn’t really cause persistence of the checkpoint activity but simply makes a slowly decaying checkpoint function longer by compromising the cyclin�k1 activity that must be suppressed by the checkpoint. We disfavor such an interpretation because it is quantitative, and the data argue that neither reduction nor increase in the remaining cyclin affects the duration of interphase. Instead, we argue that cyclin�k1 contributes to shutting off the checkpoint and propose that efficient shutoff of the checkpoint requires multiple cyclin types. One way to explain this is based on the distinct subcellular localizations of mitotic cyclins (Jacobs et al., 1998 Stiffler et al., 1999). Once activated, the checkpoint can operate in multiple cellular compartments, such as the nucleus and the cytoplasm. Although signals coordinate entry into mitosis in the cytoplasm and nucleus (Gavet and Pines, 2010), persistent nuclear checkpoint activity can prevent mitotic entry despite cytoplasmic Cdk activity (Heald et al., 1993). Individual cyclins would not be able to act on their own to reverse the checkpoint in all compartments if each is excluded from one compartment. For example, cyclin B is efficiently excluded from the nucleus in cycle 13 embryos (Maldonado-Codina and Glover, 1992) and presumably would not contribute to checkpoint reversal in this compartment, whereas cyclin B3 is nuclear (Jacobs et al., 1998). In embryos with only CycB, the checkpoint should be reversed first in the cytoplasm however, progress to mitosis should depend on slower reversal in the nucleus, which might be based on communication between compartments ( Fig. 4 F ). Consistent with this proposal, injection of CycB protein preferentially drove cytoplasmic, but not nuclear, mitotic events (Royou et al., 2008). The full complement of cyclins with distinct localizations, however, appears to reverse the checkpoint promptly and coordinately in all the compartments.

Our data demonstrate a cyclin-type effect on reversal of the DNA replication checkpoint, which emphasizes the qualitatively distinct contributions among mitotic cyclins during mitotic entry. This study opens many further questions, such as what causes the checkpoint to inactivate? How do mitotic cyclins promote checkpoint reversal? We believe answers to these questions will help us fully understand the timing mechanism of the cell division cycle.


Press release

All organisms consist of cells that multiply through cell division. An adult human being has approximately 100 000 billion cells, all originating from a single cell, the fertilized egg cell. In adults there is also an enormous number of continuously dividing cells replacing those dying. Before a cell can divide it has to grow in size, duplicate its chromosomes and separate the chromosomes for exact distribution between the two daughter cells. These different processes are coordinated in the cell cycle.

This year’s Nobel Laureates in Physiology or Medicine have made seminal discoveries concerning the control of the cell cycle. They have identified key molecules that regulate the cell cycle in all eukaryotic organisms, including yeasts, plants, animals and human. These fundamental discoveries have a great impact on all aspects of cell growth. Defects in cell cycle control may lead to the type of chromosome alterations seen in cancer cells. This may in the long term open new possibilities for cancer treatment.

Leland Hartwell (born 1939), Fred Hutchinson Cancer Research Center, Seattle, USA, is awarded for his discoveries of a specific class of genes that control the cell cycle. One of these genes called “start” was found to have a central role in controlling the first step of each cell cycle. Hartwell also introduced the concept “checkpoint”, a valuable aid to understanding the cell cycle.

Paul Nurse (born 1949), Imperial Cancer Research Fund, London, identified, cloned and characterized with genetic and molecular methods, one of the key regulators of the cell cycle, CDK (cyclin dependent kinase). He showed that the function of CDK was highly conserved during evolution. CDK drives the cell through the cell cycle by chemical modification (phosphorylation) of other proteins.

Timothy Hunt (born 1943), Imperial Cancer Research Fund, London, is awarded for his discovery of cyclins, proteins that regulate the CDK function. He showed that cyclins are degraded periodically at each cell division, a mechanism proved to be of general importance for cell cycle control.

One billion cells per gram tissue

Cells having their chromosomes located in a nucleus and separated from the rest of the cell, so called eukaryotic cells, appeared on earth about two billion years ago. Organisms consisting of such cells can either be unicellular, such as yeasts and amoebas, or multi-cellular such as plants and animals. The human body consists of a huge number of cells, on the average about one billion cells per gram tissue. Each cell nucleus contains our entire hereditary material (DNA), located in 46 chromosomes (23 pairs of chromosomes).

It has been known for over one hundred years that cells multiply through division. It is however only during the last two decades that it has become possible to identify the molecular mechanisms that regulate the cell cycle and thereby cell division. These fundamental mechanisms are highly conserved through evolution and operate in the same manner in all eukaryotic organisms.

The phases of the cell cycle

The cell cycle consists of several phases (see figure). In the first phase (G1) the cell grows and becomes larger. When it has reached a certain size it enters the next phase (S), in which DNA-synthesis takes place. The cell duplicates its hereditary material (DNA-replication) and a copy of each chromosome is formed. During the next phase (G2) the cell checks that DNA-replication is completed and prepares for cell division. The chromosomes are separated (mitosis, M) and the cell divides into two daughter cells. Through this mechanism the daughter cells receive identical chromosome set ups. After division, the cells are back in G1 and the cell cycle is completed.

The duration of the cell cycle varies between different cell types. In most mammalian cells it lasts between 10 and 30 hours. Cells in the first cell cycle phase (G1) do not always continue through the cycle. Instead they can exit from the cell cycle and enter a resting stage (G0).

Cell cycle control

For all living eukaryotic organisms it is essential that the different phases of the cell cycle are precisely coordinated. The phases must follow in correct order, and one phase must be completed before the next phase can begin. Errors in this coordination may lead to chromosomal alterations. Chromosomes or parts of chromosomes may be lost, rearranged or distributed unequally between the two daughter cells. This type of chromosome alteration is often seen in cancer cells.

It is of central importance in the fields of biology and medicine to understand how the cell cycle is controlled. This year’s Nobel Laureates have made seminal discoveries at the molecular level of how the cell is driven from one phase to the next in the cell cycle.

Cell cycle genes in yeast cells

Leland Hartwell realized already at the end of the 1960s the possibility of studying the cell cycle with genetic methods. He used baker’s yeast, Saccharomyces cerevisiae, as a model system, which proved to be highly suitable for cell cycle studies. In an elegant series of experiments 1970-71, he isolated yeast cells in which genes controlling the cell cycle were altered (mutated). By this approach he succeeded to identify more than one hundred genes specifically involved in cell cycle control, so called CDC-genes (cell division cycle genes). One of these genes, designated CDC28 by Hartwell, controls the first step in the progression through the G1-phase of the cell cycle, and was therefore also called “start”.

In addition, Hartwell studied the sensitivity of yeast cells to irradiation. On the basis of his findings he introduced the concept checkpoint, which means that the cell cycle is arrested when DNA is damaged. The purpose of this is to allow time for DNA repair before the cell continues to the next phase of the cycle. Later Hartwell extended the checkpoint concept to include also controls ensuring a correct order between the cell cycle phases.

A general principle

Paul Nurse followed Hartwell’s approach in using genetic methods for cell cycle studies. He used a different type of yeast, Schizosaccharomyces pombe, as a model organism. This yeast is only distantly related to baker’s yeast, since they separated from each other during evolution more than one billion years ago.

In the middle of the 1970s, Paul Nurse discovered the gene cdc2 in S. pombe. He showed that this gene had a key function in the control of cell division (transition from G2 to mitosis, M). Later he found that cdc2 had a more general function. It was identical to the gene (“start”) that Hartwell earlier had identified in baker’s yeast, controlling the transition from G1 to S.

This gene (cdc2) was thus found to regulate different phases of the cell cycle. In 1987 Paul Nurse isolated the corresponding gene in humans, and it was later given the name CDK1 (cyclin dependent kinase 1). The gene encodes a protein that is a member of a family called cyclin dependent kinases, CDK. Nurse showed that activation of CDK is dependent on reversible phosphorylation, i.e. that phosphate groups are linked to or removed from proteins. On the basis of these findings, half a dozen different CDK molecules have been found in humans.

The discovery of the first cyclin

Tim Hunt discovered the first cyclin molecule in the early 1980s. Cyclins are proteins formed and degraded during each cell cycle. They were named cyclins because the levels of these proteins vary periodically during the cell cycle. The cyclins bind to the CDK molecules, thereby regulating the CDK activity and selecting the proteins to be phosphorylated.

The discovery of cyclin, which was made using sea urchins, Arbacia, as a model system, was the result of Hunt’s finding that this protein was degraded periodically in the cell cycle. Periodic protein degradation is an important general control mechanism of the cell cycle. Tim Hunt later discovered cyclins in other species and found that also the cyclins were conserved during evolution. Today around ten different cyclins have been found in humans.

The engine and the gear box of the cell cycle

The three Nobel Laureates have discovered molecular mechanisms that regulate the cell cycle. The amount of CDK-molecules is constant during the cell cycle, but their activities vary because of the regulatory function of the cyclins. CDK and cyclin together drive the cell from one cell cycle phase to the next. The CDK-molecules can be compared with an engine and the cyclins with a gear box controlling whether the engine will run in the idling state or drive the cell forward in the cell cycle.

A great impact of the discoveries

Most biomedical research areas will benefit from these basic discoveries, which may result in broad applications within many different fields. The discoveries are important in understanding how chromosomal instability develops in cancer cells, i.e. how parts of chromosomes are rearranged, lost or distributed unequally between daughter cells. It is likely that such chromosome alterations are the result of defective cell cycle control. It has been shown that genes for CDK-molecules and cyclins can function as oncogenes. CDK-molecules and cyclins also collaborate with the products of tumour suppressor genes (e.g. p53 and Rb) during the cell cycle.

The findings in the cell cycle field are about to be applied to tumour diagnostics. Increased levels of CDK-molecules and cyclins are sometimes found in human tumours, such as breast cancer and brain tumours. The discoveries may in the long term also open new principles for cancer therapy. Already now clinical trials are in progress using inhibitors of CDK-molecules.


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Nobel Prizes 2020

Twelve laureates were awarded a Nobel Prize in 2020, for achievements that have conferred the greatest benefit to humankind.

Their work and discoveries range from the formation of black holes and genetic scissors to efforts to combat hunger and develop new auction formats.