MSU Skol Tech Replication eukaryotes Replication

MSU & Skol. Tech Replication eukaryotes

Replication eukaryotes Cell cycle DNA synthesis: S-phase (synthesis) Cell division: M-phase (mitosis)

Replication eukaryotes Cell cycle control There are several decision making events

Replication eukaryotes Protein kinases Serine/threonine kinases Tyrosine kinases

Replication eukaryotes Cyclin dependent kinases Cyclin A Cdk 2(T 160 -P) Cyclin E Cdk 2(T 160 -P)

Replication eukaryotes Cell cycle control by cyclin concentration

Replication eukaryotes CDK activity regulation: phosphorylation and dephosphorylation

Replication eukaryotes CDK activity regulation: binding of inhibitor proteins

Replication eukaryotes Cyclins (and many other proteins) are degraded by ubiquitindependent pathway

Replication eukaryotes Ubiquitin-dependent degradation Ubiquitin ligases Proteasome

Replication eukaryotes G 1/S transition requires ubiquitin-dependent degradation of CDK inhibitor SCF: Skp 1 Cul 1 F-box protein (70! different) e. g. Skp 2 F-box protein Ub Cul 1 E 2

Replication eukaryotes Yet another ubiquitin ligase repress G 1/S transition

Replication eukaryotes Rb protein repress G 1/S transition

Replication eukaryotes G 1/S checkpoint

Replication eukaryotes Decision to enter S-phase Yeast cells: Cell size Nutrient availability Mammalian cells: Activation signals (growth factors) Inhibitory signals Adhesion to a proper surface Contact inhibition DNA damage

Replication eukaryotes Decision to enter S-phase A number of regulatory pathways operate through CKI inhibitors of cyclin dependent kinases

Replication eukaryotes Decision to enter S-phase

Replication eukaryotes Cyclin synthesis

Replication eukaryotes Initiation of replication Organism Genome Number of origins E. coli 4. 6 Mb 1 Yeast 12 Mb 350 Human 3 000 Mb 10 000 -1 000 ? Origins should fire once in an S-phase

Replication eukaryotes Origins

Replication eukaryotes ORC (Origin recognition complex) functional analog of bacterial Dna. A

Replication eukaryotes ORC (Origin recognition complex) functional analog of bacterial Dna. A

Replication eukaryotes Cdc 6 and Cdt 1 are necessary for helicase loading

Replication eukaryotes Helicase loading Cdc 6 MCM 2 -7 ORC Cdt 1

Replication eukaryotes Starting replication activation of helicase

Replication eukaryotes MCM-GINS-Cdc 45

Replication eukaryotes Further replication complex assembly

Replication eukaryotes Replicative fork

Replication eukaryotes DNA polymerases adegbzhi k - Replicative Primase subunit Replicative lagging strand Replicative leading strand Mitochondrial Reparative High fidelity, low processivity, BER Low fidelity, TT dimer reading Low fidelity, meiotic Low fidelity

Replication eukaryotes Prevention of origins re-replication

Replication eukaryotes Chromatin assembly during replication: modification is required

Replication eukaryotes Chromatin assembly during replication: separate carriers for H 2 A/H 2 B and H 3/H 4