Mechanisms of Cell Proliferation Cell Cycle G2 S G1 Multi cellular organisms depend on cell division proliferation Each organism has a developmental plan that determines its behavior and properties Differentiation gives rise to populations of cells which specialize in specific functions Almost every cell population in the adult multicellular organism is specified by its lineage and environment Within the mature organism cells refrain from exerting their intrinsic potential to grow and divide beyond territories and patterns laid down in the overall developmental plan Normal Growth factor dependence proliferation depends on availability of tissue type specific growth factors which are signals not nutrients In many cases factor withdrawal leads to apoptosis Anchorage dependence proliferation requires interaction of transmembrane proteins called integrins with components of the extracellular matrix ECM components Specific integrins recognize specific ECM molecules Contact inhibition contact with like cell types inhibits cell movement and proliferation Contact inhibition of growth limits division in culture when cells form a contiguous monolayer Contact inhibition of movement affects the cytoskeletal organization and motility of cells in a monolayer Contact with unlike cells allows motility and hence spontaneous cell sorting Limited proliferation capacity vertebrate somatic cells divide a limited number of times ca 50 70 divisions for human cells before the cells enter a senescent state that maintains metabolic activity but stops all further division Disruption of normal cell proliferation due to mutant alleles inherited from parents somatic mutations arising in the organism epigenetic changes which alter expression levels of key genes Immortalization and aneuploidy diploid cells grown to the point of senescence sometimes give rise to clonal lines that survive and grow continuously beyond normal limits Partial or complete loss of growth factor dependence transformed cells may gain the ability to grow on less rich serum or at lower initial cell density Loss of contact inhibition of growth Transformed cells may overgrow monolayers and pile up onto each other foci Loss of anchorage dependence Cells may grow on soft agar or in suspension rather attached to a substrate Loss of contact inhibition of movement Transformed cells maintain a motile phenotype which may be a consequence of failure to respond properly to cell cell adhesion signals For human fibroblasts after 50 70 divisions cells enter a state of replicative senescence in which cells are metabolically active but cease to proliferate The immediate cause is a strong block to cell cycle progression and entry to S phase mediated by cyclin kinase inhibitors CKI such as p16INK4A and p21CIP1 Cells can be forced to bypass senescence by suppression of the pRB and p53 replication regulators e g by the action of viral oncogenes such as SV40 large T or adenovirus E1A Cells thus forced to continue to divide reach a second proliferative block known as replicative crisis characterized by drastic chromosomal instability leading almost invariably to cell death The limit of some 50 70 division cycles for human diploid fibroblasts is mediated by telomere length The telomere is an extension of DNA at chromosome ends generated by the telomerase reverse transcriptase TERT which uses an internally bound RNA loop as a template The replication process terminates before the end of the lagging strand and telomeres thus shorten with each division unless maintained by telomerase Telomerase is active in germline cells but inactive in somatic cells Telomere length correlates with age of cells in culture and with age in the organism Cells reach senescence when the short telomeres trigger the protective mechanisms of p53 which stimulates the CKIs to halt further cell cycle progress Cells reach crisis when telomeres are lost exposing chromosome ends and provoking the double strand repair mechanism to make inappropriate attempts at recombination and ligation In some cases immortalized cells maintain telomeres by reactivating telomerase and maintain relatively stable chromosomes However a significant proportion of immortalized cells are viable in the absence of telomerase and use a less well characterized process alternative maintenance of telomeres ALT www chembio uoguelph ca CDKs and their role in cell cycle During G1 the levels of G1 cyclins rise and these cyclins associate with cyclin dependent kinases CDKs Activity of G1 CDKs promotes the passage of cells through START budding yeast also known as the restriction point R in fission yeast and higher eukaryotes After passing through this point a cell is committed to continue through the cell cycle Nature Reviews Molecular Cell Biology 2 815 826 2001 Major Cell Cycle Regulatory Proteins Function of p53 in response to DNA Damage Levels of cell cycle regulation oncogenic factors Activation of extracellular receptors Growth factors bind to extracellular domains of transmembrane receptors linked to cytoplasmic domains many of which have been found to be protein tyrosine kinases Examples of oncogenic transformation at this level include v sis which causes cells to express their own PDGF and erb B which expresses a truncated EGF receptor lacking an extracellular domain and which is active without a ligand Cells may start to secrete their own growth factors the autocrine effect instead of depending on an external source In other cases cell may lose dependence on growth factors by developing factor independent receptors with constitutively active kinase modules www chembio uoguelph ca Ligand induced dimerization of receptors stimulates the kinase Initially it was assumed that these tyrosine kinases would act through phosphorylation cascades much like Ser Thr kinases However in many cases the best substrate turned out to be the receptor itself Autophosphorylation or more correctly mutual phosphorylation is mediated by dimerization induced by ligand binding by subunit orientation modulated by ligand binding or by other ligand induced conformational changes Autophosphorylation generates key phosphotyrosine sites which are receptive to binding by signaling adapter proteins These proteins are known as grbs or growthfactor receptor bound The factor grb2 consists of a single SH2 domain flanked by two SH3 domains SH3 domains act as binding sites for proline rich domains in effector molecules acting downstream in the signaling