Lectures 7 -10 Cell Cycle Regulation and Cancer. Introduction to the Cell Cycle. The Eukaryotic Cell Cycle and Chromosome Dynamics. Identification and Experimental Basis of Cell Cycle Regulatiory Factors. Major players: Cyclins , cdk’s (cyclin – dependant kinases)
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The Eukaryotic Cell Cycle and Chromosome Dynamics
Cell Cycle Regulatiory Factors
Also termed SCF
Cell Cycle Regulatory Factors contd…
Mutant studies in Saccharomyces cerevisiae (budding yeast) and Schizosaccharomyces pombe (fission yeast)
Complementation studies of the cdc (cell division cycle) temperature sensitive mutants using a plasmid library of wild type yeast DNA have enabled identification of numerous proteins involved in cell cycle regulation.
Identification and Experimental Basis of Cell Cycle Regulatory Factors contd…
Identification of MPF (Maturation Promoting Factor) and Cyclins in Xenopus Oocytes
Experimental Demonstration that the Synthesis & Degradation of Cyclin B are Required for the Cycling of MPF and Mitotic Events in Xenopus Egg Extracts
(mutation in destruction box sequence)
cyclin B degrades
Active Separase breaks
(Morgan, Nature 374 (1995) 131-134)
(b) Regulation of pre replication complexes:
cdk’ssimultaneously activate initiation of replication and prevent re-initiation at origin sites by phosphorylation & disassociation ofMcM’sfrom theORCs (origin recognition complex)which are permanently associated with replication origins.
For theMcM’sto reassembleat the origins they need to bedephosphorylated which does not occur until mitosis.
Multiple cdks and cyclins regulate passage of mammalian cells through the cell cycle.
Cyclin D is Required for Passage Through the Restriction Point in the Mammalian Cell: An In Vivo Depletion Experiment
Cells microinjected anti-cyclin D
8 h after growth factors
An in vivo
+ Cyclin A
release from RP
To ensure orderly passage of the cells through cell cycle there are four defined “checkpoints” where certain events need to occur before the cell will proceed in the cell cycle; otherwise the cell will be “arrested” at that checkpoint: G1 arrest, S arrest, G2 arrest and M arrest.
There are specific checkpoint protein kinases(chks) that are directly involved in arresting the cells.
Most commonly mutated tumorsuppressor gene associated with human cancers is p53 which encodes the transcription factor (TF) p53 protein.
Role of p53 in G1 Checkpoint Arrest
p53- mutant cells
(which activates a checkpoint kinase (phosphorylate p53)
Cancer and Growth Related Factors:Proto-Oncogenes and Tumor Supressor Genes
Intracellular transducers e.g., ras (a mutant form of an inner cell surface GTPase), whose function activates a number of other regulatory factors including another oncogene termedraf (a protein kinase)that work in concert to regulate cell proliferation via theMAP Kinase Pathwaywhich activates transcription factors
Oncogenes Contribute To Cancer By Over Expression Of Normal Regulatory Components
Why do patients who inherit one mutant allele of RB have a high probability of developing retinal tumors in childhood?
This is characteristic of cancers related to the loss of tumor suppressor genes and a likely explanation is the predisposition of such genes to a “lose of heterozygosity” or (LOH).
Cancer is characterized by many abnormalities in chromosome numbers(aneuploidy)and organization(translocations, inversions deletions, andamplifications).This is a direct result of the lack of checkpoint control during the cell cycle.
CANCER CELLS ARE CHARACTERIZED BY GLOBAL ABERRATIONS IN THE NUMBER SIZE AND BANDING PATTERNS OF CHROMOSOMES GIVING RISE TO THE IDEA OF A “CHAOTIC GENOME” IN CANCER CELLS.
Chromosome painting of an osteosarcoma cell line (MG-63)
Chromosome painting of normal diploid human cells