Lumpy skin disease (LSD) is an emerging transboundary viral disease of livestock caused by the lumpy skin disease virus (LSDV), which has caused substantial economic losses and negatively impacted veterinary public health. The underlying pathogenic mechanisms remain poorly understood, limiting the effective prevention and control of LSD. Here, we identify LSDV-encoded protein LSDV001 as a positive regulator of IL-1β- and TNFα-triggered signaling. LSDV001 interacts with TAK1 and TAB2/3 and promotes assembly of the TAK1-TAB2/3 complex. This leads to IKK-dependent activation of the transcription factor NF-κB and induction of downstream inflammatory cytokines. LSDV001-deficient virus (LSDVΔ001) has attenuated the ability to activate NF-κB and induce the expression of inflammatory cytokines. Infection with LSDVΔ001 leads to smaller skin nodules and reduced inflammation compared to wild-type LSDV. Our findings suggest that LSDV001 acts as a key virulence factor of LSDV by promoting excessive inflammatory response upon infection. IMPORTANCE Lumpy skin disease is a current global concern caused by the lumpy skin disease virus (LSDV), for which there is a lack of safe and efficient vaccines. In this study, we report that LSDV001 protein potentiates IL-1β- and TNFα-triggered IKK-dependent activation of NF-κB and transcription of inflammatory cytokines. Mechanistically, LSDV001 enhances inflammatory response by interacting with TAK1 and TAB2/3 to promote TAK1-TAB2/3 complex formation. We further demonstrate that LSDV001 deficiency attenuates LSDV-triggered inflammatory response and pathogenesis. Our findings identify a new virulence factor and reveal a novel pathogenic mechanism of LSDV by which LSDV001 enhances inflammatory response.
Opportunistic pathogens can cause infections when host defenses are compromised. Among them, Streptococcus suis (S. suis) colonizes the upper respiratory tract of pigs and causes severe diseases in both swine and humans. Although the pathogenic mechanisms of these bacteria have been partially elucidated, the molecular processes that govern their adaptation, colonization, and pathogenesis remain incompletely understood. In this study, we identified PrlP as a transcriptional repressor in S. suis that responds to mildly acidic, oxidative, hyperosmotic, and thermal stresses, and regulates bacterial growth, chain morphology, nasal colonization, and virulence. The C-terminal S24 peptidase domain of PrlP mediates stress-induced self-cleavage to control protein stability, while the N-terminal helix-turn-helix (HTH) DNA-binding domain is essential for its transcriptional regulatory function. Combined ChIP-seq and RNA-seq analyses revealed its binding motif (5'-CCTGAAWCT-3') and identified B9H01_08740 as a direct target gene, as further validated by EMSA. Notably, deletion of B9H01_08740 in the prlP-deficient background restored the associated phenotypes. These findings highlight PrlP as a key regulator in S. suis that maintains cellular homeostasis in response to stress conditions and modulates target genes such as B9H01_08740 to promote nasal colonization and virulence. Therefore, this study provides new insights into the regulatory mechanisms of pathogenic bacteria and may aid in the development of targeted strategies against S. suis infections.
The molecular mechanisms underlying the transport of influenza A virus (IAV) membrane proteins to the cell surface remain largely unclear. In this study, siRNA screening identifies Rab27a as a critical host factor regulating this transport process. GTP-bound Rab27a operates via its effectors, synaptotagmin-like protein 1 (SYTL1) and SYTL4, to facilitate the transport of vesicles carrying viral membrane proteins to the plasma membrane. Absence of Rab27a or SYTL4 does not block the early stages of the IAV life cycle but restricts viral assembly and budding. Notably, silencing SYTL4 provides superior protection in the female mouse IAV infection model. This investigation elucidates the molecular mechanism by which Rab27a and its effectors modulate the transport of IAV membrane proteins, thereby bridging a critical gap in IAV life cycle research and presenting a potential target for the development of antiviral drugs.
Swine acute diarrhea syndrome coronavirus (SADS-CoV) is a novel coronavirus that causes acute diarrhea, vomiting, and high mortality in suckling piglets. Research has demonstrated that certain viruses enhance their replication by modulating intracellular cholesterol metabolism. However, the impact of SADS-CoV infection on cellular cholesterol synthesis remains unclear. Here, we found that SADS-CoV Spike (S) protein promoted syncytium formation by positively regulating cholesterol synthesis. Specifically, the virus upregulated the rate-limiting enzyme 3-hydroxy-3-methyl-glutaryl-CoA reductase through the inhibition of AMP-activated protein kinase (AMPK) activity. This inhibition was mediated by the activation of AKT-dependent phosphorylation of AMPKα at Ser485. Further investigation revealed that SADS-CoV S protein activated the PI3K/AKT pathway to promote cholesterol synthesis, a process that required the membrane protein integrin β1 (ITGB1). Importantly, we discovered that cholesterol facilitated cell-to-cell fusion mediated by the viral S protein, which enhanced syncytium formation. In summary, our findings demonstrate that the SADS-CoV S protein enhances cellular cholesterol accumulation by activating the PI3K/AKT/AMPK pathway through ITGB1, and that cholesterol facilitates syncytium formation mediated by the viral S protein. These insights contribute to a better understanding of SADS-CoV infection mechanisms and may inform future therapeutic strategies. IMPORTANCE Cholesterol, a vital component of cellular membranes, is crucial for maintaining cell structure and function. It also acts as an essential host factor for the entry, replication, and propagation of various viruses. In this study, we show that the Spike protein of swine acute diarrhea syndrome coronavirus (SADS-CoV) promotes syncytial formation by upregulating cellular cholesterol synthesis. The viral Spike protein activates the PI3K/AKT signaling pathway, leading to increased cholesterol production through the inhibition of AMP-activated protein kinase (AMPK). This upregulation of cholesterol facilitates cell-to-cell fusion, a process that enhances viral spread and pathogenesis. Moreover, we demonstrate that integrin β1 (ITGB1) acts as a critical host factor that links the viral Spike protein to the activation of the PI3K/AKT pathway. ITGB1 interacts with the S protein, playing a pivotal role in viral replication and cholesterol synthesis regulation. Our findings highlight the critical role of cholesterol in SADS-CoV infection and provide a deeper understanding of the molecular mechanisms behind viral replication. This research opens up potential therapeutic strategies targeting cholesterol metabolism to mitigate the effects of SADS-CoV and similar viral infections.
Classical swine fever virus (CSFV) infection induces complete mitophagy, which is essential for the clearance of damaged mitochondria. The endosomal sorting complex required for transport (ESCRT) machinery plays a vital role in mediating phagophore closure and autophagosome-lysosome fusion during starvation-induced autophagy. Nevertheless, its involvement in CSFV-induced mitophagy and the underlying mechanisms remain insufficiently understood. Here, we found that the ESCRT-III subunits including CHMP1A, CHMP1B, and CHMP4B, along with the AAA-ATPase VPS4, were actively recruited to autophagosomes during CSFV-induced mitophagy. Consistent with this, depletion of CHMP1A, CHMP1B, CHMP4B or VPS4A disrupted mitophagic flux, impairing both PINK1-PRKN-dependent and -independent pathways. Further investigations revealed that CSFV transiently recruited these subunits to nascent autophagosomes for phagophore sealing during mitophagy. Remarkably, multiple CSFV nonstructural proteins (NSPs) including NS3, NS4B, NS5A and NS5B interacted with these ESCRT key subunits and colocalized on mitophagosomes. Taken together, our study identifies CHMP1A, CHMP1B, CHMP4B, and VPS4A as pivotal regulators of phagophore closure in CSFV-induced mitophagy, unveiling novel mechanisms by which the virus manipulates host cellular pathways and highlighting potential therapeutic targets for infection control.Abbreviation: ATF4: activating transcription factor 4; ATG5: autophagy related 5; BafA1: bafilomycin A1; BFP: blue fluorescent protein; BNIP3L/NIX: BCL2 interacting protein 3like; BSA: bovine serum albumin; CALCOCO2/NDP52: calcium binding andcoiled-coil domain 2; CCCP: carbonyl cyanide 3-chlorophenylhydrazone; CHMP: charged multivesicular body protein; COX4: cytochrome c oxidase subunit 4; CSFV: classical swine fever virus; DAPI: 4',6-diamidino-2-phenylindole; DN: dominant-negative; ER: endoplasmic reticulum; ESCRT: endosomal sorting complex required for transport; FUNDC1: FUN14 domain containing 1; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; GFP: green fluorescent protein; hpt: hours post-transfection; HSPD1/HSP60: heat shock protein family D (Hsp60) member 1; IB: immunoblotting; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MFF: mitochondrial fission factor; MFN2: mitofusin 2; MITO: mitochondria; MOI: multiplicity of infection; mtDNA: mitochondrial DNA; OPTN: optineurin; PBS: phosphate-buffered saline; PINK1: PTEN induced kinase 1; PRKN: parkin RBR E3 ubiquitin protein ligase; RAPA: rapamycin; RFP: redfluorescent protein; RT-qPCR: reverse transcription-quantitativereal-time polymerase chain reaction; RT-PCR: real-time polymerasechain reaction; SD: standard deviation; siCtrl: negative control siRNA; siRNA: small interfering RNA; SQSTM1/p62: sequestosome 1; TOMM20: translocase of outer mitochondrial membrane 20; VDAC1: voltage dependent anion channel 1; VPS4A: vacuolar protein sorting 4 homolog A; WCL: whole-cell lysate; WT: wild-type.
