Long noncoding RNA AVAN promotes antiviral innate immunity by interacting with TRIM25 and enhancing the transcription of FOXO3a

Chengcai Lai; Lihui Liu; Qinghua Liu; Sijie Cheng; Keyu Wang; Lingna Zhao; Min Xia; Cheng Wang; Hongjing Gu; Yueqiang Duan; Zhongpeng Zhao; Lili Zhang; Ziyang Liu; Jianjun Luo; Jianxun Song; Penghui Yang; Runsheng Chen; Xiliang Wang

The copyright holder for this preprint (which was not peer-reviewed) is the author/funder. . https://doi.org/10.1101/623132 doi: bioRxiv preprint expression in nucleus by associating with the FOXO3a promoter and performing chromatin 240 remodeling, which additionally increased IL-8 and neutrophil chemotaxis. Previous experiments demonstrated that AVAN is located in the cytoplasm (Fig 2E, 2F ). 245 To investigate the molecular mechanism of AVAN in the cytoplasm, RNA pull-down assays 246 using biotin-labeled AVAN or AVAN antisense control followed by mass spectrometry (MS) 247 analysis were performed. The result of MS revealed that E3 ubiquitin ligase TRIM25, an 248 RNA-binding protein (48,49), was pulled down by AVAN in IAV-BJ501-infected A549 cells 249 but not by the AVAN antisense control (Fig 6A, Fig S7A) . This result was confirmed by AVAN 250 RNA pull-down western blot experiments ( Fig 6B) . Besides, ChIRP followed by western blot 251 revealed that AVAN-specific probes could pull down TRIM25 while LacZ couldn't ( Fig 6C) . 252 To validate the interaction between AVAN and TRIM25, we immunoprecipitated TRIM25 253 from IAV-BJ501-infected A549 cells and quantified the protein-bound AVAN. Significantly 254 higher levels of AVAN were detected with exogenous ( Fig 6D) and endogenous ( Fig 6E) 255 TRIM25 immunoprecipitation than with the isotype immunoglobulin G (IgG) control. Fig 6F) . Furthermore, to 260 explore the TRIM25 binding site on AVAN RNA, three truncated probes from AVAN were 261 All rights reserved. No reuse allowed without permission.