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CYCLINS

CELL DIVISION

     The complexity of the eukaryotic nucleus increased the complexity of the process of cell division and required regulation of the cell cycle.   The nuclei of an amoeba and a human lymphocyte are evident in the images below.

AMOEBA LYMPHOCYTE

Eukaryotes share a number of proteins which regulate the cell cycle, such as cyclins, kinases, phosphatases, growth factors, and their receptors.  Some proteins which are involved in the initiation of the eukaryotic cell cycle may have homologs in bacteria, such as Cdc6/Cdc18, MCMs, and Cdc45 (Leatherwood, 1998).  Many of the proteins which regulate the cell cycle evolved before the separation of coelomate lineages such homologs of AP-1, Fos, Jun, and members of the myc family of transcription factors (Perkins, 1988; Peyrefitte, 2001).

 

MITOSIS

CYCLINS

     Transcription initiation in eukaryotes requires a number of generalized transcription factors such as TFIIB.  GTF2B is part of the TFIIB complex which is involved in transcription in eukaryotes, eubacteria, and archebacteria.  It is homologous to cyclin A, indicating that cyclins may have evolved from proteins with a more generalized role in transcription in the ancestors of eukaryotes (OMIM; Bagby, 1995).  Archaeal homologs to TFIIB include regions homologous to the cyclin fold.  Thus, the evolution of the cyclin fold which eukaryotes use to regulate the cell cycle predated the evolution of the eukaryotic nucleus (Noble, 1997).  The following is a depiction of the generalized transcription factors which function in eukaryotic transcription, including TFIIB.

Label
transcription

After Brown, p. 261.

 

 

     Cyclins compose a gene family whose members share a cyclin box region of about 100 amino acids (Noble, 1997).  Sponges possess proteins which, when introduced into human cells, are capable of decreasing the amount cyclin B1, stopping the cell cycle before the Mitosis stage, and inducing apoptosis.  Thus, some of the regulatory mechanisms involving cyclin B and cell cycle control evolved before the first animals (Brown, 2004).   Plants may possess 7 cyclin D and multiple cyclin A proteins.

 

Cyclin A is required for two distinct points of the cell cycle: entry into M and the completion of S.

 

B cyclins are expressed in late G2 and early M (Potuschak, 2001).

 

Cyclins D1, D2, and D3 are expressed in specific tissues.  Cyclins D and E  inactivate RB in G1 (Vidal, 2000).

 

Cyclin I is similar in structure to cyclins G1 and G2 (Jensen, 2000).

 

 

CHECKPOINTS

     Cells must not only be able to promote cell division, they must also be able to stop cells from dividing when it is inappropriate.  This ability to check growth has obviously been lost in the cells of liver, breast, and uterine cancers depicted in the following images.

cancer cancer
cancer

     There are a number of proteins which interact to produce checkpoints in the cell cycle and prevent the transition to the following stage without the appropriate signals.  All eukaryotes share checkpoints preceding the beginning of the S and M phases of the cell cycle (Takeo, 2004). 

      In early G1, a restriction point is reached in which extracellular signals determine whether the cell progresses into late G1 and cell division or exits the cell cycle in G0 (Ho, 2002).  In order to continue with the cell cycle, two cyclins function in the G1 phase: cyclin D which functions in the middle of G1 and E which functions at the transition between G1 and S.  Cyclin D1, cyclin D2, Cdk4, and Cdk6 are protoncogenes whose misexpression can cause inappropriate progression through this checkpoint.  Although cyclin E is also expressed in G1, cyclin E and cdk2 mutations rarely promote cancer.

     The complexes formed by cyclins D, E, and their CDKs inactivate the RB family of tumor suppressor proteins (which include RB, p107, and p130).  D cyclins interact with growth factor pathways and determine response to mitogens.  D cyclins then activate cyclin E through the CIP/KIP family, particularly p21 and p27.  In response to DNA damage, p53 causes the accumulation of p21 which destroys cyclin D and prevents progression through G1.  The G1 checkpoint occurs through destruction of cyclin D (Agami, 2002; Ho, 2002).

     In animals, Forkhead is expressed in S and begins a cascade of transcription factors and the B cyclins which will regulate the transitions between G2 and M and M and G1.  Forkhead genes are not known in plants (Potuschak, 2001).  The G2-phase DNA-damage checkpoint is regulated by PI3K-like complexes (such as as ATM), checkpoint loading complexes (CLC), checkpoint sliding clamps (CSC), BRCT proteins (perhaps including BRCA) and effector kinases.  Yeast and human homologs of all these proteins exist  (O’Connell, 2000).

      It seems that the mutations which cause cancerous growth either effect the percentage of cells which proliferate or decrease the number which die but do not significantly effect the length of the cell cycle. 

 

p53

     Although cancer is often considered to be single disorder (for example, when some refer to a “cure for cancer”), it is not.  Different cancers in different individuals can have virtually nothing in common other than uncontrolled cell growth.  Cells from the blood of leukemia patients are depicted in the following images.

 

 

leukemia leukemia

     However, there is one feature that is shared by most human cancers.  The vast majority of human cancers have developed in part due to the loss of function of the p53 protein.  p53 is normally inactive in cells but in response to a variety of stressful stimuli, it will block progression of the cell cycle allowing for either repair of cell damage or apoptosis.  Upon activation, p53 travels to the nucleus where it functions as a transcription factor promoting the expression of genes which stop the cell cycle and/or promote apoptosis.  It also performs functions which are transcription-independent.   Flies possess p53 homologs which can bind to human p53 targets and overexpression of p53 causes apoptosis in flies (Sutcliffe, 2004).  Nematodes possess a p53 homolog which is expressed throughout the body and regulates cellular responses to stress and damage (Sutcliffe, 2004).

     The two human homologs of p53, p63 and p73, are involved in development, immunity, and neurogenesis.  p63 helps to maintain the epithelial lining and is required for proper development of the skin, limbs, breast, and prostate.  p73 functions in the development of neurons, inflammation, and pheromone detection.  There is some evidence that both are involved in some aspects of cell cycle control but they cannot be classified as tumor suppressors with p53.  Invertebrate p53 proteins are more similar to p63 and p73, suggesting that the tumor suppression of p53 is a later development in the evolution of this family (Yang, 2002a).

 

RETINOBLASTOMA

     The retinoblastoma (RB) protein is homologous to the cyclin fold and TFIIB.  RB binds to the protein E2F under resting cell conditions.  After being hyperphosphorylated, RB dissociates from E2F, allowing the cell to enter the S phase.  More than 20 proteins are known to interact with RB (Noble, 1997; Hagemeier, 1993).  Although retinoblastoma is a key inhibitor of this early G1 restriction point, its mutation is uncommon in all but a few cancers (retinoblastoma, osteosarcoma, and small cell lung carcinoma) (Ho, 2002). 

      The RB-E2F complex is dephosphorylated during mitosis (probably by PP-1), “resetting” it.  Cyclin A is broken down at this point.  The cyclin A-CDK2 complex can hyperphosphorylate RB during cell cycle progression.    Other members of the RB family, p130 and p107, also regulate E2F transcription factors (Vidal, 2000).

  The retinoblastoma protein is phosphorylated by cyclin-dependent kinases which causes it to dissociate from the transcription factor E2F, ending the inhibition of cell growth. In addition to a role in regulating the cell cycle, retinoblastoma functions in the differentiation of a number of cell types. The regions of the retinoblastoma protein are homologous (in structure but not function) to protein regions utilized by some archaea and poxviruses (Takemura, 2005).

 

CDK

     All eukaryotes use multiple kinases to regulate the cell cycle.  Although the cyclin-dependent kinases perhaps the best known, they do not appear to have been the first kinases to regulate the cell cycle.  In comparing the subfamilies of kinases involved in the regulation of the cell cycle in higher and lower eukaryotes, it is evident that the duplication of ancestral kinase genes involved in the cell cycle occurred in the evolution of the earliest eukaryotes.  The earliest branch contains kinases which function as a checkpoint of the cycle, followed by branches containing other kinases which regulate mitosis and meiosis.  The CDK family seems to have been the last group to evolve (Krylov, 2003).

 

CYCLIN DEPENDENT KINASES

CDK2 forms a complex with cyclin A and E and becomes activated at the end of G1 when cyclin E is available.  To be active, it must be bound to a cyclin and must be phosphorylated by CAK.  Its activity is also determined by the cleavage of the inhibitors CDKN1A and B by caspases.

 

CDK3 is absent in some mice species without any observed deficits; it may be a redundant signal.  Although the complete function of cdk3 is not known, it is an important rate-limiting step in the regulation of the cell cycle.  Most eukaryotic signaling pathways which affect the cell cycle control G1 (Matsuoka, 2000). 

 

CDK4 is activated (along with CDK6) early in G1 and blocks cell cycle repression by retinoblastoma.  Ras and CDK4 alter cyclin D and E so that they resist inhibition.  Mutations can contribute to some tumors such as melanoma.

 

CDK5 is expressed primarily in neurons where it phosphorylates neurofilaments and the microtubule-associated protein tau.   It is required in the development in the mammalian CNS and plays a role in cocaine addiction.  Null mutations in mice result in the absence of cerebellar foliation.

 

CDK5L is on the X chromosome and may cause X-linked infantile spasm syndrome ISSX.

 

CDK6 is active in mid-G1.  CDKs 6 and 4 link progression of the cell cycle to stimulation by growth factors.  CDK6 and 4 are very similar in structure and amino acid sequence.

 

CDK7 binds cyclin H and forms part of CAK, a multi-subunit complex which phosphorylates and activates CDKs.  CAK is a component of transcription factor TFIIH.

 

CDK8 interacts with cyclin C.  CDK8/cyclin C form part of RNA polymerase II holoenzyme and other transcription complexes.  CDK8 phosphorylates cyclin H to activate CDK7.

 

CDK9 can bind with cyclin T and act as a transcription elongation factor (P-TEFb).  Its activity is enhanced by UV radiation and chemical agents and inhibited by 7SK snRNA.

 

CDKI

     Cyclin dependent kinase inhibitors (CKIs) can be divided into two families: the INK4 family and the CIP/KIP family.  Ink4, Cip/Kip, and RB protein families prevent cells from entering S. 

     INK4 which interact with CDK 4/6, inhibiting their interaction with cyclin D.  The Ink4 family is composed of p15Ink4b, p18Ink4c, and p19Ink4d.  These proteins possess ankyrin repeats and inhibit cyclin D changes.

      The CIP/KIP family (with members such as p21, p27, and p57) which interact with CDK2 and CDK4/6.  The CIP/Kip family is composed of p21Cip1, p27Kip1, and p57Kip2.  The expression of p27 is needed to maintain G0 and low expression of p27 can contribute to cancer (Ho, 2002; Agami, 2002; Vidal, 2000).

 

 

Myb

Myb proteins are only known in coelomates where they seem to function in the S phase of the cell cycle.  There are three copies of Myb in vertebrates.

c-Myb is required for hematopoeisis and mutations can cause leukemia.

 

A-Myb functions in spermatogenesis and in growth of mammary glands.

 

B-Myb functions in early embryonic development (Manak, 2002).

 

 

OTHER PROTEINS

     In order for cells to complete a phase of mitosis and mitosis in general, cyclins are destroyed by ubiquitin-dependent proteolysis.  Human ubiquitin carrier protein E2-C/UbcH10 is homologous to, and can substitute for, that of clams (Townsley, 1997).