Cdh1 regulates cell cycle through modulating the claspin/Chk1 and the Rb/E2F1 pathways

Area of Science:

• Cell Biology
• Molecular Biology
• Cancer Research

Background:

• APC/Cdh1 is a key regulator of the cell cycle, with roles in DNA damage repair that are not fully understood.
• Previous work established Cdh1's interaction with Rb for Skp2 degradation.

Purpose of the Study:

• To elucidate the precise role of APC/Cdh1 in cell cycle regulation and its interaction with the Rb pathway.
• To identify novel APC/Cdh1 interacting proteins and substrates.

Main Methods:

• Affinity purification coupled with mass spectrometry to identify interacting proteins.
• Ubiquitination assays to determine Cdh1's substrate.
• Cell-based assays in HeLa and primary human fibroblasts to assess cell cycle progression and senescence.

Main Results:

• Claspin identified as a novel Cdh1-interacting protein and ubiquitin substrate.
• Cdh1 inactivation activates the Claspin/Chk1 pathway.
• Cdh1 reciprocally regulates the Rb pathway by competing with E2F1 for hypophosphorylated Rb.
• Cdh1 depletion causes premature S phase entry in p53/Rb-defective cells but premature senescence in normal fibroblasts.
• Loss of other tumor suppressors like PTEN and VHL also induces p53/Rb-dependent senescence.
• SV40 LT-antigen partially reverses Cdh1 depletion-induced growth arrest.

Conclusions:

• Loss of APC/Cdh1 function has differential effects on cell cycle progression depending on the integrity of p53 and Rb pathways.
• Cdh1 loss is oncogenic only in cells with defective p53/Rb pathways, explaining its infrequent mutation in tumors.