Abstract：
Porcine reproductive and respiratory syndrome virus (PRRSV) is a major swine pathogen, which can survive host antiviral immunity with various mechanisms. PRRSV infection induces macroautophagy/autophagy, facilitating virus replication. MALT1, a central immune regulator, was manipulated by PRRSV to optimize viral infection at different stages of the virus cycle. In this study, the key role of MALT1 in autophagy regulation during PRRSV infection was characterized, enlightening the role of autophagy flux in favor of virus spread and persistent infection. PRRSV-induced autophagy was confirmed to facilitate virus proliferation. Furthermore, autophagic fusion was dynamically regulated during PRRSV infection. Importantly, PRRSV-induced MALT1 facilitated autophagosome-lysosome fusion and autolysosome formation, thus contributing to autophagy flux and virus proliferation. Mechanically, MALT1 regulated autophagy via mediating MTOR-ULK1 and -TFEB signaling and affecting lysosomal homeostasis. MALT1 inhibition by inhibitor Mi-2 or RNAi induced lysosomal membrane permeabilization (LMP), leading to the block of autophagic fusion. Further, MALT1 overexpression alleviated PRRSV-induced LMP via inhibiting ROS generation. In addition, blocking autophagy flux suppressed virus release significantly, indicating that MALT1-maintained complete autophagy flux during PRRSV infection favors successful virus spread and its proliferation. In contrast, autophagosome accumulation upon MALT1 inhibition promoted PRRSV reserve for future virus proliferation once the autophagy flux recovers. Taken together, for the first time, these findings elucidate that MALT1 was utilized by PRRSV to regulate host autophagy flux, to determine the fate of virus for either proliferation or reserve.Abbreviations: 3-MA: 3-methyladenine; BafA1: bafilomycin A1; BFP/mBFP: monomeric blue fluorescent protein; CQ: chloroquine; DMSO: dimethyl sulfoxide; dsRNA: double-stranded RNA; GFP: green fluorescent protein; hpi: hours post infection; IFA: indirect immunofluorescence assay; LAMP1: lysosomal associated membrane protein 1; LGALS3: galectin 3; LLOMe: L-leucyl-L-leucine-methyl ester; LMP: lysosomal membrane permeabilization; mAb: monoclonal antibody; MALT1: MALT1 paracaspase; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MOI: multiplicity of infection; MTOR: mechanistic target of rapamycin kinase; NFKB/NF-κB: nuclear factor kappa B; nsp: nonstructural protein; ORF: open reading frame; pAb: polyclonal antibody; PRRSV: porcine reproductive and respiratory syndrome virus; PRRSV-N: PRRSV nucleocapsid protein; Rapa: rapamycin; RFP: red fluorescent protein; ROS: reactive oxygen species; SBI: SBI-0206965; siRNA: small interfering RNA; SQSTM1/p62: sequestosome 1; TCID50: 50% tissue culture infective dose; TFEB: transcription factor EB; ULK1: unc-51 like autophagy activating kinase 1.
摘要：
猪繁殖与呼吸综合征病毒(PRRSV)是猪的主要病原,它可以通过各种机制在宿主的抗病毒免疫中存活。PRRSV感染诱导巨自噬/自噬，促进病毒复制。MALT1是一种中枢免疫调节剂,由PRRSV操纵以在病毒周期的不同阶段优化病毒感染。在这项研究中,MALT1在PRRSV感染过程中自噬调节中的关键作用，启发自噬通量在病毒传播和持续感染中的作用。PRRSV诱导的自噬被证实促进病毒增殖。此外，自噬融合在PRRSV感染过程中受到动态调节。重要的是,PRRSV诱导的MALT1促进自噬体-溶酶体融合和自溶酶体形成，从而促进自噬通量和病毒增殖。机械上,MALT1通过介导MTOR-ULK1和-TFEB信号传导并影响溶酶体稳态来调节自噬。通过抑制剂Mi-2或RNAi诱导的溶酶体膜透化（LMP）抑制MALT1，导致自噬融合的阻滞。Further,MALT1过表达通过抑制ROS产生减轻PRRSV诱导的LMP。此外,阻断自噬通量显著抑制病毒释放，这表明在PRRSV感染期间MALT1维持的完全自噬通量有利于病毒的成功传播和增殖。相比之下,一旦自噬通量恢复，MALT1抑制后的自噬小体积累促进了PRRSV为未来病毒增殖的储备。一起来看,第一次,这些发现阐明PRRSV利用MALT1调节宿主自噬通量,确定病毒的增殖或储备的命运。
