Rtt105 regulates RPA function by configurationally stapling the flexible domains
Sahiti Kuppa, Jaigeeth Deveryshetty, Rahul Chadda, Jenna R. Mattice, Nilisha Pokhrel, Vikas Kaushik, Angela Patterson, Nalini Dhingra, Sushil Pangeni, Marisa K. Sadauskas, Sajad Shiekh, Hamza Balci, Taekjip Ha, Xiaolan Zhao, Brian Bothner, Edwin Antony
Abstract
Replication Protein A (RPA) is a heterotrimeric complex that binds to single-stranded DNA (ssDNA) and recruits over three dozen RPA-interacting proteins to coordinate multiple aspects of DNA metabolism including DNA replication, repair, and recombination. Rtt105 is a molecular chaperone that regulates nuclear localization of RPA. Here, we show that Rtt105 binds to multiple DNA binding and protein-interaction domains of RPA and configurationally staples the complex. In the absence of ssDNA, Rtt105 inhibits RPA binding to Rad52, thus preventing spurious binding to RPA-interacting proteins. When ssDNA is available, Rtt105 promotes formation of high-density RPA nucleoprotein filaments and dissociates during this process. Free Rtt105 further stabilizes the RPA-ssDNA filaments by inhibiting the facilitated exchange activity of RPA. Collectively, our data suggest that Rtt105 sequesters free RPA in the nucleus to prevent untimely binding to RPA-interacting proteins, while stabilizing RPA-ssDNA filaments at DNA lesion sites.