TRIM7/RNF90 promotes autophagy via regulation of ATG7 ubiquitination during <i>L. monocytogenes</i> infection

Abstract

<i>L. monocytogenes</i> is a widely used infection model for the research on pathogenesis and host defense against gram-positive intracellular bacteria. Emerging evidence indicates that posttranslational modifications play a critical role in the regulation of macroautophagy/autophagy. However, little is known about the posttranslational modifications of ATG7, the essential protein in the autophagy process. In this study, we demonstrated that the RING-type E3 ligase TRIM7/RNF90 positively regulated autophagosome accumulation by promoting the ubiquitination of ATG7 at K413, thereby affecting <i>L. monocytogenes</i> infection. TRIM7 expression was induced by a variety range of conditions, including starvation, rapamycin stimulation, and <i>L. monocytogenes</i> infection. TRIM7 deficiency in mice or cells resulted in elevated innate immune responses and increased <i>L. monocytogenes</i> infection. ATG7 was associated with TRIM7 and the positive regulatory role of TRIM7 in <i>L. monocytogenes</i> infection-, starvation- or rapamycin-induced autophagosome accumulation was suggested by TRIM7 deficiency, TRIM7 overexpression, and TRIM7 knockdown. Further mechanistic investigation indicated that TRIM7 promoted the K63-linked ubiquitination of ATG7 at K413 and ubiquitination at this site was required for the function of ATG7 in autophagy and <i>L. monocytogenes</i> infection. Thus, our findings suggested a new regulator in intracellular bacterial infection and autophagy, with a novel posttranslational modification targeting ATG7. This research may expand our understanding of host anti-bacterial defense and the role of autophagy in intracellular bacterial infection.<b>Abbreviations:</b> ATG3: autophagy related 3; ATG5: autophagy related 5; ATG7: autophagy related 7; ATG10: autophagy related 10; ATG12: autophagy related 12; ATG16L1: autophagy related 16 like 1; Baf A1: bafilomycin A₁; CQ: chloroquine; BMDC: bone marrow-derived dendritic cell; BMDM: bone marrow-derived macrophage; CFUs: colony-forming units; CXCL10/IP-10: C-X-C motif chemokine ligand 10; EBSS: Earle's balanced salt solution; ELISA: enzyme-linked immunosorbent assay; IFIT1/ISG56: interferon induced protein with tetratricopeptide repeats 1; IFNB/IFN-β: interferon beta; IL6: interleukin 6; IRF3, interferon regulatory factor 3; <i>Lm: L. monocytogenes</i>; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MEF: mouse embryonic fibroblast; MOI: multiplicity of infection; PLA: proximity ligation assay; PMA: phorbol myristate acetate; PMA-THP1, PMA-differentiated THP1; PMs: peritoneal macrophages; PTMs: posttranslational modifications; STING1, stimulator of interferon response cGAMP interactor 1; TBK1, TANK binding kinase 1; TNF/TNF-α: tumor necrosis factor; TRIM7/RNF90: tripartite motif containing; Hainan Provincial Natural Science Foundation of China.