Anti-WRNIP1 polyclonal antibody

Wrnip1 (Werner helicase-interacting protein 1) has been implicated in the bypass of stalled replication forks in bakers' yeast. However, the function(s) of human Wrnip1 has remained elusive so far. Here we report that Wrnip1 is distributed inside heterogeneous structures detectable in nondamaged cells throughout the cell cycle. In an attempt to characterize these structures, we found that Wrnip1 resides in DNA replication factories. Upon treatments that stall replication forks, such as UVC light, the amount of chromatin-bound Wrnip1 and the number of foci significantly increase, further implicating Wrnip1 in DNA replication. Interestingly, the nuclear pattern of Wrnip1 appears to extend to a broader landscape, as it can be detected in promyelocytic leukemia nuclear bodies. The presence of Wrnip1 into these heterogeneous subnuclear structures requires its ubiquitin-binding zinc finger (UBZ) domain, which is able to interact with different ubiquitin (Ub) signals, including mono-Ub and chains linked via lysine 48 and 63. Moreover, the oligomerization of Wrnip1 mediated by its C terminus is also important for proper subnuclear localization. Our study is the first to reveal the composite and regulated topography of Wrnip1 in the human nucleus, highlighting its potential role in replication and other nuclear transactions.

Conflicts between replication and transcription are a common source of genomic instability, a characteristic of almost all human cancers. Aberrant R-loops can cause a block to replication fork progression. A growing number of factors are involved in the resolution of these harmful structures and many perhaps are still unknown. Here, we reveal that the Werner interacting protein 1 (WRNIP1)-mediated response is implicated in counteracting aberrant R-loop accumulation. Using human cellular models with compromised Ataxia-Telangiectasia and Rad3-Related (ATR)-dependent checkpoint activation, we show that WRNIP1 is stabilized in chromatin and is needed for maintaining genome integrity by mediating the Ataxia Telangiectasia Mutated (ATM)-dependent phosphorylation of Checkpoint kinase 1 (CHK1). Furthermore, we demonstrated that loss of Werner Syndrome protein (WRN) or ATR signaling leads to formation of R-loop-dependent parental ssDNA upon mild replication stress, which is covered by Radiorestistance protein 51 (RAD51). We prove that Werner helicase-interacting protein 1 (WRNIP1) chromatin retention is also required to stabilize the association of RAD51 with ssDNA in proximity of R-loops. Therefore, in these pathological contexts, ATM inhibition or WRNIP1 abrogation is accompanied by increased levels of genomic instability. Overall, our findings suggest a novel function for WRNIP1 in preventing R-loop-driven genome instability, providing new clues to understand the way replication-transcription conflicts are handled.