Capns1 regulates usp1 stability and stem cells maintenance

Abstract

Calpains are a family of calcium-related cysteine-proteases that are involved in a wide number of cellular processes. The ubiquitous calpains, micro- and milli-calpain, are heterodimers composed of catalytic subunits and a common regulatory subunit, encoded by CAPNS1. We identified USP1 deubiquitinase as a CAPNS1-interacting protein. USP1 is a key modulator of DNA repair, partly through deubiquitination of its known targets FANCD2 and PCNA. Usp1 knockout mice have a severe phenotype and die soon after birth. Usp1−/− cells are defective in FANCD2 focus formation and are hypersensitive to DNA damage. PCNA ubiquitination is higher in USP1-depleted cells than in control cells, thus leading to recruitment of error-prone, translesion DNA synthesis (TLS) polymerases and the consequent increase in mutation rate. USP1 promotes inhibitor of DNA binding (ID) protein stability and stem cell-like characteristics in osteosarcoma and is required for normal skeletogenesis. We found that the ubiquitinated form of the USP1 substrate PCNA is stabilized in CAPNS1-depleted U2OS cells and mouse embryonic fibroblasts (MEFs), favoring polymerase-η loading on chromatin and increased mutagenesis. USP1 degradation directed by the cell cycle regulator APC/Ccdh1, which marks USP1 for destruction in the G1 phase, is upregulated in CAPNS1-depleted cells. USP1 stability can be rescued upon forced expression of calpain-activated Cdk5/p25, previously reported as a cdh1 repressor. Our data suggest a connection between the calpain system and the ubiquitin pathway in the regulation of DNA damage response and place calpain at the interface between cell cycle modulation and DNA repair.