Summary Ku70 and Ku80 form Ku, a ring-shaped protein that initiates the non-homologous end-joining (NHEJ) DNA repair pathway. 1 Specifically, Ku binds to double-stranded DNA (dsDNA) ends and recruits other NHEJ factors ( e.g. , DNA-PKcs and LIG4). While Ku binds to double-stranded RNA (dsRNA) 2 and traps mutated-DNA-PKcs on ribosomal RNA in vivo, 3,4 the physiological significance of Ku-dsRNA interactions in otherwise wild-type cells remains elusive. Intriguingly, while dispensable for murine development, 5,6 Ku is essential in human cells. 7 Despite similar genome sizes, human cells express ∼100-fold more Ku than mouse cells, implying functions beyond NHEJ, possibly through a dose-sensitive interaction with dsRNA, which is ∼100 times weaker than with dsDNA. 2,8 While investigating the essentiality of Ku in human cells, we found that depletion of Ku - unlike LIG4 - induces profound interferon (IFN) and NF-kB responses reliant on the dsRNA-sensor MDA5/RIG-I and adaptor MAVS. Prolonged Ku-degradation also activates other dsRNA-sensors, e.g. PKR that suppresses protein translation, and OAS/RNaseL that cleaves rRNAs and eventually induces growth arrest and cell death. MAVS, RIG-I, or MDA5 knockouts suppressed IFN signaling and, together with PKR knockouts, partially rescued Ku-depleted human cells. Ku-irCLIP analyses revealed that Ku binds to diverse dsRNA, predominantly stem-loops in primate-specific Alu elements 9 at anti-sense orientation in introns and 3’-UTRs. Ku expression rose sharply in higher primates tightly correlating with Alu-expansion (r = 0.94/0.95). Together, our study identified a vital role of Ku in accommodating Alu-expansion in primates by mitigating a dsRNA-induced innate immune response, explaining the rise of Ku levels and its essentiality in human cells.
<div>Abstract<p>Subunits of SWI/SNF chromatin remodeling complexes are frequently mutated in human malignancies. The PBAF complex is composed of multiple subunits, including the tumor-suppressor protein PBRM1 (BAF180), as well as ARID2 (BAF200), that are unique to this SWI/SNF complex. <i>PBRM1</i> is mutated in various cancers, with a high mutation frequency in clear cell renal cell carcinoma (ccRCC). Here, we integrate RNA-seq, histone modification ChIP-seq, and ATAC-seq data to show that loss of PBRM1 results in <i>de novo</i> gains in H3K4me3 peaks throughout the epigenome, including activation of a retinoic acid biosynthesis and signaling gene signature. We show that one such target gene, <i>ALDH1A1</i>, which regulates a key step in retinoic acid biosynthesis, is consistently upregulated with PBRM1 loss in ccRCC cell lines and primary tumors, as well as non-malignant cells. We further find that ALDH1A1 increases the tumorigenic potential of ccRCC cells. Using biochemical methods, we show that ARID2 remains bound to other PBAF subunits after loss of PBRM1 and is essential for increased ALDH1A1 after loss of PBRM1, whereas other core SWI/SNF components are dispensable, including the ATPase subunit BRG1. In total, this study uses global epigenomic approaches to uncover novel mechanisms of PBRM1 tumor suppression in ccRCC.</p>Implications:<p>This study implicates the SWI/SNF subunit and tumor-suppressor PBRM1 in the regulation of promoter histone modifications and retinoic acid biosynthesis and signaling pathways in ccRCC and functionally validates one such target gene, the aldehyde dehydrogenase ALDH1A1.</p></div>
