Regulation of cell cycle

in this presentation you will aware about regulation of cell cycle and the proteins involved in cancer study. How these protein regulated by cell cycle.

1. Regulation of Cell Cycle By – Mohit Kumar

2. overview Described by Howard and Pelc in 1953. From single cell, zygote to adult cell requires a series of cell division known as ‘cell cycle’.

3. Division cycles Somatic division cycle : used to fulfill specific requirements. Embryonic cycle: having lack of G1 and G2 phase. Meiotic cell cycle: allow formation of haploid gametes. Endoreduplication: S phase is not followed by mitosis.

4. Human cell cycle Length of cell cycle is highly variable, even within cells of a single organism. In human, frequency of cell turnover range few hours in early embryonic development to an average of two to five days for epithelial cells. In human, rapidly cell cycle length is about 24 hours. G1 phase lasts approximately nine hours . S phase lasts ten hours . G2 phase lasts about four and one-half hours . M phase lasts approximately one-half hour .

5. Factors that affects cell cycle regulation The death of nearby cells. As sweeping as the release of growth promoting hormones. Crowding of cells. Size of the cell during division (decreasing surface to volume ratio). Chromosomal mutation

6. Checkpoints A checkpoint is one of several points in the eukaryotic cell cycle at which the development 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 the metaphase.

7. G1 checkpoint Also known as restriction point in yeast. Check for genomic DNA damage. Plays crucial role at this check post due to proliferation of the cell. The cell can halt the cycle and waits, whether to moves G° phase or signal to improves condition .

8. G2 checkpoint Entry into mitotic phase. Whether chromosome have been replicated properly. No DNA damage occurs, if occurs than repair mechanism repair the mutation and completes proper replication.

9. M checkpoint Known as spindle checkpoint. At the end of metaphase in interphase. Sister chromatids are correctly attached to the microtubule or not. Cycle get arrested if spindle fiber attached chromatids doesn’t moves at poles.

10. cyclins and cyclin-dependent kinases (CDKs) CDKs Master regulator of cell cycle. Role is to phosphorylate protien on ‘Ser’ and ‘Thr’ amino acids. Yeast having 1 CDK while animals having 9 CDKs. Cyclins Regulatory subunits of complexes with kinase activity (with Cdks ). Their presence “oscillates” in the cell cycle. There are G-1cyclin, G1-S cyclin, S cyclin and M cyclin.

11. Cyclins and CDKs in vertebrates

12. Activation of CDKs-cyclin complex

13. Regulation of activity of cyclin-CDKs complex's Cyclins undergo a cycle of synthesis and degradation in each cell cycle, while CDKs level remains constant during the cell cycle. Cyclins-CDKs complex inactivated by regulated proteolysis of cyclin at specific cell cycle stage. Cyclin level in cells are controlled not only by changes in cyclin destructions but also by changes in cyclin synthesis.

14. M-CDKs They are mitosis promoting factor or maturation promoting factor (MPF). Synthesized during S and G-2 phase Include chromosomal condensation. Breakdown of nuclear envelop Assembly of mitotic spindle apparatus. Alignment of condensed chromosome at metaphase plate. It phosphorylate number of protein such as condensin, lamin etc.

15. Regulation of CDKs-cyclin in S. pombe

16. After the proper association of chromosomes with spindle microtubules, the M-CDKs complexes activates the anaphase promoting complex/cyclosome (APC/C). APC/C acts on different substrate at different time. It bears to different co activators i.e. cdc20 and cdh1.

17. Inactivation of cyclin-CDKs complex's

18. Inactivation of CDKs –cyclin complex Occurs by ubiquitin dependent proteolysis. Proteolysis of cyclin is catalyzed by ubiquitin lygase. Enzyme SCF(SKp1, cullin, F-box) & E3 ubiquitin lygase responsible for destruction of G1 phase cyclin. E-3 ubiquitin ligase- anaphase promoting complex/ cyclosome (APC/C) is responsible for S&M phase cyclin inactivation.

19. Role of Rb protein in cell cycle regulation RB1(retinoblastoma) gene encodes Rb protein of 110 KDa . It is a tumor suppressor protein and loss of function mutation in Rb1 gene leads tumor. It is present on 13q14.2 chromosome location.

20. Rb protein Its active, un or hypophosporylated form binds with E2-F transcription factor and suppress the gene transcription which is required for transition from G1 to S phase. The product of these genes are required for S phase of the cell cycle. Cyclin-D – CDKs 4/6 phosphorylate the Rb protein results losses its affinity with EF-2.

21. Rb protein Restriction point: This site ensure that the transition is responsive to extracellular signals. Then cyclin E level is increase, cyclin-E +CDK-2 complex drives fully Phosphorylated form of Rb protein due to that E2F is released. E2F binds with DP(DNA binding protein) & transcription translation could occurs.

22. p53 protein Tp53 gene encodes for p53 protein of molecular mass of 53KDa. It is also a tumor suppressor gene encodes a polypeptide chain of 393 amino acid residue. It is described as Guardian of the genome & cellular gatekeeper because it involves in the complex network of cellular events It regulates the expression of several genes involves in DNA repair, apoptosis, cell growth, antioxidant defense etc .

23. p53 protein p53 is inactivated by its –ve regulator MDM2 (mouse double minute 2). MDM2 functions as E3 ubiquitin ligase & cause proteosomal degradation of p53 protein. Upon DNA damage or other stress leads to dissociation of p53 and MDM2 complex, activated p53 will induce a cell cycle arrest to either repair and survival of the cell or apoptosis.

24. pP53 protein In double stranded DNA break, protein kinase ATM is activated which in turn activates CHK 2 kinase. Both ATM &CHK2 phosphorylate p53 at distinct sites. At the same time ATM phosphorylate MDM2 in way that cause its functional inactivation. Phosphorylated p53 acts as transcription factor for p21 gene. p21 protein binds with G1/S-CDK and S CDK complexes and inhibit their activity, thereby helping to block entry into the cell cycle. Hence, p53 level is increase and arrest the cell in the G1 phase of the cell cycle.

25. Ubiquitin mediated protein degradation Mainly two type of proteolytic system are there: Lysosome mediated- membrane bound + acid hydrolysis including many proteolytic enzyme. Proteasome mediated- regulated breakdown of protein by energy dependent manner.

26. Proteasome Found in both prokaryotes and eukaryotes. In eukaryotes they present in both cytosol and nucleus. In eukaryotes they are of two types: 20S Proteasome- cylindrical barrel shaped structure arranged as four rings in heptamer form upon one another. 26S Proteasome- it catalysis the ATP-dependent degradation of polyubiquitinated proteins.

27. Ubiquitination Most well established means of targeting protein to proteasome is by addition of ubiquitin. Ubiquitin is highly conserved protein of 76 amino acid residue and covalently attached to a lysine residue on a target protein. Protein can be monoubiquitinated, multiubiquitinated or polyubiquitinated.

28. Activation of ubiquitin Activated by ubiquitin E-1 (ubiquitin activating enzyme), in energy dependent manner E-1 attach to ubiquitin at C- terminal. Then, E-1 transfer to a ‘cysteine residue’ present on an E-2 (ubiquitin conjugating enzyme). Finally E-3 (ubiquitin ligase)transferred the activated ubiquitin from E-2 to a lysine amino acid residue of its target protein, forming an isopeptide bond.

29. Attractions: Cel l cycle Checkpoints Activation/inactivation of cyclin- cdks complex Rb protein P53 protein Ubiquitin mediated proteasomal degradation Thanks Much…