ligases Cell cycle activities of Cdks and the ubiquitin Activation and activities of G1/s-Cdk ·G1/s cyclins accumulate during G1 o Because their transcription is allowed to proceed · G1/S Cdk phosphorylates substrate (Retinoblastoma (Rb)): pocket protein (meaning it holds onto proteins) that sequesters) members of the E2F family of transcription factors (meaning holds E2F out of action · E2F stimulates the transcription of genes that promote entry into S-phase such as G1-cyclins, S-cyclins and DHFR (dihydrofolate reductase) o DHFR is key in making nucleotides that are needed in S-phase for DNA replication mitogen mitogen receptor 1 Ras MAP kinase activation of transcription regulatory protein CYTOSOL NUCLEUS immediate early gene expression transcription Myc regulatory protein - delayed-response gene expression active G1-Cdk active Rb protein inactivated E2F protein positive feedback active G1/S-Cdk active E2F protein S-phase gene transcription G,/S-cyclin (cyclin E) S-cyclin (cyclin A) active S-Cdk DNA SYNTHESIS positive feedback inactivated Rb protein G, S Mitogen= growth factors that stimulate cells to divide MAP kinase- phosphorylates transcription factors to activate them o In this case will transcribe ‘Immediate early genes' One of the immediate early genes is a Transcription regulatory protein called Myc o Myc is a proto-oncogene meaning it has a mutation in it which is oncogenic. This means that it can become things that become cancer o Myc turns on another wave of transcription. This includes promotion of transcription of cyclins. This speeds up the accumulation of cyclins. 8 Active Rb holds onto E2F G1-Cdk phosphorylates the Rb ® This changes the shape of Rb and it lets go of E2F ® E2F can now start transcribing genes o In a positive feedback loop, it can produce more E2F and therefore more transcription ® More cyclins produced ® End product is active S-Cdk and DNA synthesis Why might we want to prevent entry into S- phase? . The arrest of the cell cycle in G1 or G2 following DNA damage reduces the risk that a neoplastic change will be passed onto the daughter cells · DNA damage includes damage done by ultraviolet and x-irradiation and chemotherapeutic drugs How does the DNA damage checkpoint work? · Severe DNA damage in G1 increases the activity of p53 · P53 is a transcription factor · In undamaged cells, p53 is unstable and is present at low concentrations since it is bound by Mdm2 and ubiquitin ligase that targets p53 for degradation. · DNA damage activates kinases (ARM/ATR and Chk1/Chk2) that phosphorylate p53, releasing it from Mdm2 o Stabilises p53 (doesn't degrade) P53 transcribes CKIs
. Once stable and active, p53 turns on the expression of genes that encode CKIs (inhibitors of Cdk-cyclin complexes) such as p21 (p21 is similar to p27, a CKI normally present in early G1) · When p21 or p27 is bound to G1/S-Cdk or S-Cdk the complex is inactive x-rays DNA DNA damage ATM/ATR kinase activation 1 Chk1/Chk2 kinase activation Mdm2 PHOSPHORYLATION OF p53 p53 053 UBIQUITYLATION AND DEGRADATION IN PROTEASOMES a stable, active p53 ACTIVE p53 BINDS TO