WDR82在紫外線損傷下調控雙向啟動子區長鏈非編碼RNA表達及R環形成的作用機制

Abstract

DNA損傷會嚴重阻礙核糖核酸聚合酶II（RNAPII）的進程，並引發轉錄阻斷性損傷，從而導致 R-loop 的累積。R-loop 是一種包含 DNA:RNA 雜合體和被置換的單鏈去氧核糖核酸的三鏈結構。持續的 R-loops 會觸發基因組不穩定並導致細胞死亡。R-loops 可以通過轉錄耦合核苷酸切除修復 (TC-NER) 因子被加工成去氧核糖核酸雙鏈斷裂。在紫外線損傷的反應中，大多數 RNA表現量會減少，但長非編碼核糖核酸的比例卻顯著增加。有趣的是，某些反義股長非編碼核糖核酸的轉錄水平上升，而鄰近的編碼基因在紫外線暴露後下調。值得注意的是，這些反義長非編碼核糖核酸基因和編碼基因是以頭對頭的方式排列的。驅動這些長非編碼核糖核酸上調的機制及其在紫外線壓力下的功能作用仍然知之甚少。在這篇研究裡，我發現紫外線損傷會導致WDR82 與核糖核酸聚合酶II 的分離，進而促進長非編碼核糖核酸的表達。此外，這一現象伴隨著紫外線誘導的長非編碼核糖核酸雙向啟動子處的 R-loop 形成。在紫外線損傷後，核糖核酸聚合酶 II的聚集會從編碼基因的啟動子轉移到了反義長非編碼核糖核酸基因的啟動子。抑制WDR82的表現後，紫外線誘導的長非編碼核糖核酸會增加。這些結果表明，紫外線損傷會在雙向啟動子處引發核糖核酸聚合酶 II的轉移，並且WDR82與核糖核酸聚合酶II 的分離是紫外線誘導的長非編碼核糖核酸表達的關鍵促進因子。

DNA damage can severely block RNA polymerase II (RNAPII) progression, leading to transcription-blocking lesions and the accumulation of R-loops, a triple-stranded structure containing a DNA:RNA hybrid and a displaced single-stranded DNA. Persistent R-loops trigger genome instability and lead to cell death. R-loops can be processed into DNA double-strand breaks by transcription-coupled nucleotide excision repair (TC-NER) factors. In response to UV damage, while the levels of most RNA are reduced, a substantial fraction of long non-coding RNAs (lncRNAs) are increased. Interestingly, the transcription levels of certain antisense lncRNAs are elevated, while the nearby coding genes are downregulated following exposure to UV. Notably, these antisense lncRNA genes and coding genes are arranged in a head-to-head orientation. The mechanism underlying the upregulation of these non-coding RNAs and their functions they undertake in response to UV stress remain poorly understood. This study reveals that the dissociation of WDR82 from RNAPII triggers the expression of lncRNAs upon UV damage. Moreover, this phenomenon is accompanied by R-loop formation at the bidirectional promoters of UV-induced lncRNAs. The RNAPII occupancy shifts from the promoters of coding genes to the promoters of antisense lncRNA genes upon UV damage. Furthermore, CUT&RUN-seq analysis reveals that the RNAPII-S5 occupancy at TSS is increased in lncRNA genes with bidirectional promoters upon UV stress, while the occupancy of WDR82 is lost. Depletion of WDR82 results in elevated expression of UV-induced lncRNAs, which is associated with increased R-loop accumulation. These results suggest that WDR82 suppresses the expression of UV-induced lncRNAs, and that the dissociation of WDR82 from RNAPII triggers shifts in RNAPII occupancy at bidirectional promoters, thereby promoting lncRNA expression upon UV damage.