Cell cycle Cell division Unicellular Cell growth and

Cell cycle Cell division Unicellular Cell growth and DNA replication throughout cell cycle Multicellular §Division through defined phases §Cell growth continuous §DNA synthesis is in only one phase §Cell growth and DNA synthesis highly coordinated and regulated §Cell cycle controlled by a series of protein kinases §Defects in cell cycle regulation-cancer

Why cell cycle? Ø Ø Cell division is regulated by highly conserved networks The comparative approach will illuminate the variation in the intrinsic stability of cell cycle controls in the different species Cell cycle progression regulating mechanisms in eukaryotes are largely conserved: cells as different as yeast and mammalian cells use the same kind of regulatory molecules to trigger cell cycle progression new insights in cell cycle regulation and the mechanisms that prevent uncontrolled proliferation of cells way to novel anti-tumor drugs cancer Infections fungal & viral Atheriosklerosis Restenosis Cell cycle Malaria Tissue engeneering Tissue repair & regeneration Yield Growth rate

INTERPHASE S (DNA synthesis) G 1 is ito MIT (M) OTIC PHA SE M to y C sis es n ki G 2

0. 5 µm Mitosis Chromosome duplication (including DNA synthesis) Centromere Sister chromatids Separation of sister chromatids Centromeres Sister chromatids

http: //www. biologymad. com/Cell. Division/Cell. Di 1. gif

11 hrs 1 hr 8 hrs 90 min. 4 hrs Human cells (24 hours) Mitosis Yeast cells (90 minutes) G 0 Synthesis Embryo cells (less than 30 min. ) Skin fibroblasts, liver cells

Cells at different stages of cell cycle can be distinguished by their DNA content

Regulation of cell cycle S. cerevisiae Nutrients Start Mating factors Cell size G 0 Animal cells Restriction point Growth factors

Regulation of cell cycle

Daughter chromosomes not aligned properly Spindle checkpoint S -checkpoint DNA damage DNA repair Cell cycle checkpoints?

Summary – Cells divide by following carefully scripted program of molecular events collectively called the cell cycle. – The cell cycle is subdivided into five phases named G 1, S, G 2, M, and G 0. Cells not actively dividing reside in G 1 or G 0 phase. – Several checkpoints define critical decision-making events in the cell cycle

Key molecule responsible for cell cycle regulation • Identification of MPF: Maturation Promoting Factor

Yeast Is a Model System for Analyzing Cell Cycle S. pombe lengthens and then divides with a septum, while S. cerevisiae buds during a cycle in which G 2 is absent and M occupies the greatest part.

Key molecule responsible for cell cycle Regulation ØS. cereviceae ØTemperature sensitive mutants (cell division cycle mutants) ØCell cycle arrested at START, (mutation in protein called Cdc 2) ØA similar protein identified in S. pombe ØProtein Cdc 2 controls both G 1 and G 2 checkpoints ØCharacterization- Cdc is a protein kinase START

Key molecule responsible for cell cycle Regulation • Accumulation and degradation of cyclins The cyclins were identified as proteins that accumulate throughout interphase and are rapidly degraded toward the end of mitosis.

Key molecule responsible for cell cycle Regulation MPF: M-phase Promoting Factor • MPF is composed of two key subunits: Cdc 2 and Cyclin B. – Cdc 2 is the protein that encoded by genes which are required for passage through START as well as for entry into mitosis. – Cyclin B is a regulatory Cyclin dependant Kinases subunit required for catalytic activity of the Cdc 2 protein (Cdks) kinase.

1970 s-80 s | 2001 Cyclins & Cdks • Interaction of Cdk’s & different cyclins triggers the stages of the cell cycle Leland H. Hartwell checkpoints Tim Hunt Cdks Sir Paul Nurse cyclins

Summary – Progression through the cell cycle is under the control of proteins that form checkpoints – These proteins are protein kinases – Protein kinases can function only with cyclins Cyclin Kinase X Cyclin Kinase √

Four molecular mechanisms that regulate Activities of Cdk's • Mechanisms of Cdk regulation

Families of Cyclins and Cyclin-Dependent Kinases The cell cycles of higher eukaryotes is controlled by multiple cyclins and cdc 2 Cyclin dependent kinases (cdks) Labelled numerically Cdc 2 - Cdk 1 -Cdk 8 Cyclins Labelled –alphabets Cyclin-B, cyclin-G, cyclin E

Control by cyclin/CDK complexes Distinct cyclin-cdk complexes control progression through cell cycle checkpoints.

Growth Factors and D-type Cyclins Growth factors D-type cyclin Restriction point

Control of the G 1 checkpoint expression of most CDKs does not vary much throughout the cycle, but without their corresponding cyclins, they are not functional

RB = tumour suppressor

Control of the G 1 checkpoint Cyclin A E 2 F turns on the expression of genes required for DNA synthesis

Regulation of Cell Cycle INK 4 (RB ) Cell 2004; 116: 221 -34

Cyclins • Cyclines and cyclin-dependent kinases (cdks) – The cyclines have oscillating levels during cell cycle – The cyclines are regulatory subunits of the CDK-kinases • cyclines+ cdk �cell cycle-dependent variations in the activity of the kinases Cyclin E Cyclin A Cyclin B determined by mitogenic growth factors Cyclin D G 0 G 1 S G 2 M

Control of the S checkpoint Assembly of the prereplication complex.

Control of the S checkpoint • 3 key steps Activation of the replication complex.

G 2 check point- check for DNA damage Cyclin B synthesized Cyclin B accumulates

G 2 - check point Wee 1 protein kinase Inactive Cdc 25 ADP Inactive CDK 1

Control of the G 2 checkpoint ATM/R P ATP ADP

CDK inhibitors

CDKs are regulated by Cyclin Dependent Kinase Inhibitors (CKIs) p 21 CDK Cyclin p 21 p 16 CDK 4 p 16 Cyclin active inactive CDK 4/6 Cyclin D + INK 4 + Cyclin D

DNA damage, other insults Growth Signal p 53 p 21 G 1 phase cyclin/CDK S phase

Inhibitor of Cell Cycle Progression Mdm 2 p 53 Normal Cells Mdm 2 stimulates degradation of p 53. Cellular levels of p 53 are low

P ATM/R p 53 X Mdm 2

Inhibitor of Cell Cycle Progression • Induction of p 21 by DNA damage P p 53 Conc. increase p 21 CDK Cyclin p 21

Overview of checkpoint controls in the cell cycle Lodish et al. , 2004

Overview of checkpoint controls in the cell cycle Lodish et al. , 2004