Related Experiment Videos
A complex between E2F and the pRb-related protein p130 is specifically targeted by the simian virus 40 large T
D A Wolf1, H Hermeking, T Albert
1Institute für Klinische Molekularbiologie und Tumorgenetik, GSF Forschungszentrum für Umwelt und Gesundheit, München, Germany.
Abstract:
The p130 protein is a recently cloned member of the retinoblastoma protein family. We show here that transformation of NIH3T3-L1 fibroblasts (L1 cells) by the simian virus 40 large T antigen (LTAg) depends on the disruption of DNA binding complexes between transcription factor E2F and p130. LTAg binds to the pocket region of p130 in vivo and disrupts the E2F-p130 complexes. E2F-p130 complexes are present only in quiescent L1 cells and disappear at the G1/S phase boundary concomitantly to induction of DNA synthesis and expression of the E2F-regulated cdc2 gene. p130 is a substrate of cyclin-dependent kinase 2 (Cdk2) in vitro and associates with a Cdk in vivo which is activated upon serum stimulation in late G1. Overexpression of p130 inhibits cdc2 promoter activity and entry of quiescent L1 cells into S phase. The results demonstrate that p130 is negative regulator of cell cycle progression which is specifically targeted by LTAg during cell transformation.
Insights
Simian virus 40 large T antigen (LTAg) disrupts cell cycle regulation by targeting the p130 protein. This interaction inhibits DNA binding complexes, preventing cell cycle progression and transformation.
Area of Science:
- Molecular Biology
- Cell Biology
- Virology
Background:
- p130 is a member of the retinoblastoma protein family.
- Retinoblastoma proteins play crucial roles in cell cycle regulation.
Purpose of the Study:
- To investigate the role of p130 in simian virus 40 large T antigen (LTAg)-mediated cell transformation.
- To elucidate the mechanism by which LTAg affects p130 function and cell cycle control.
Main Methods:
- NIH3T3-L1 fibroblasts (L1 cells) were transformed with LTAg.
- DNA binding complexes involving p130 and transcription factor E2F were analyzed.
- In vivo and in vitro binding assays were performed to study LTAg-p130 interactions.
- Cell cycle progression and gene expression (cdc2) were monitored.
- Overexpression studies of p130 were conducted.
Main Results:
- LTAg disrupts DNA binding complexes between E2F and p130 in L1 cells.
- LTAg directly binds to the pocket region of p130, leading to complex dissociation.
- E2F-p130 complexes are present in quiescent cells and disappear at the G1/S phase boundary.
- p130 is phosphorylated by cyclin-dependent kinase 2 (Cdk2) and associates with an activated Cdk in G1.
- Overexpression of p130 inhibits cdc2 promoter activity and S-phase entry in quiescent cells.
Conclusions:
- p130 acts as a negative regulator of cell cycle progression.
- LTAg targets p130 to disrupt cell cycle control and promote cell transformation.
- The interaction between LTAg and p130 is critical for LTAg-induced cell transformation.