Autophagy delivers cytoplasmic constituents to autophagolysosomes and is linked to both innate and adaptive immunity. Toll-like receptor 4 (TLR4) signaling induces autophagy and recruits Beclin-1, the mammalian homolog of yeast Atg6, to the receptor complex. We found that tumor necrosis factor receptor (TNFR)-associated factor 6 (TRAF6)-mediated, Lys(63) (K63)-linked ubiquitination of Beclin-1 is critical for TLR4-triggered autophagy in macrophages. Two TRAF6-binding motifs in Beclin-1 facilitated the binding of TRAF6 and the ubiquitination of Beclin-1. Lys(117), which is strategically located in the Bcl-2 homology 3 (BH3) domain of Beclin-1, was a major site for K63-linked ubiquitination. The deubiquitinating enzyme A20 reduced the extent of K63-linked ubiquitination of Beclin-1 and limited the induction of autophagy in response to TLR signaling. Treatment of macrophages with either interferon-gamma or interleukin-1 also triggered the K63-linked ubiquitination of Beclin-1 and the formation of autophagosomes. These results indicate that the status of K63-linked ubiquitination of Beclin-1 plays a key role in regulating autophagy during inflammatory responses.
The SARS (severe acute respiratory syndrome) outbreak was caused by a coronavirus (CoV) named the SARS-CoV. SARS pathology is propagated both by direct cytotoxic effects of the virus and aberrant activation of the innate immune response. Here, we identify several mechanisms by which a SARS-CoV open reading frame (ORF) activates intracellular stress pathways and targets the innate immune response. We show that ORF8b forms insoluble intracellular aggregates dependent on a valine at residue 77. Aggregated ORF8b induces endoplasmic reticulum (ER) stress, lysosomal damage, and subsequent activation of the master regulator of the autophagy and lysosome machinery, Transcription factor EB (TFEB). ORF8b causes cell death in epithelial cells, which is partially rescued by reducing its ability to aggregate. In macrophages, ORF8b robustly activates the NLRP3 inflammasome by providing a potent signal 2 required for activation. Mechanistically, ORF8b interacts directly with the Leucine Rich Repeat domain of NLRP3 and localizes with NLRP3 and ASC in cytosolic dot-like structures. ORF8b triggers cell death consistent with pyroptotic cell death in macrophages. While in those cells lacking NLRP3 accumulating ORF8b cytosolic aggregates cause ER stress, mitochondrial dysfunction, and caspase-independent cell death.
The Toll-like receptors (TLR) play an instructive role in innate and adaptive immunity by recognizing specific molecular patterns from pathogens. Autophagy removes intracellular pathogens and participates in antigen presentation. Here, we demonstrate that not only TLR4, but also other TLR family members induce autophagy in macrophages, which is inhibited by MyD88, Trif, or Beclin 1 shRNA expression. MyD88 and Trif co-immunoprecipitate with Beclin 1, a key factor in autophagosome formation. TLR signaling enhances the interaction of MyD88 and Trif with Beclin 1, and reduces the binding of Beclin 1 to Bcl-2. These findings indicate TLR signaling via its adaptor proteins reduces the binding of Beclin 1 to Bcl-2 by recruiting Beclin 1 into the TLR-signaling complex leading to autophagy.One of the mechanisms by which the innate immune system senses the invasion of pathogenic microorganisms is through Toll-like receptors (TLRs), 3 which recognize specific molecular patterns that are present in microbial components. Stimulation of different TLRs induces distinct patterns of gene expression, which not only leads to the activation of innate immunity, but also instructs the development of antigen-specific acquired immunity (1, 2). TLR signaling predominantly induces interferon regulatory factors, and nuclear factor-B (NF-B) and AP1 activation. This elicits the production of type I interferon and inflammatory cytokines (3, 4). TLR signaling also facilitates the selection of microbial antigens for presentation by dendritic cells (5). Parasitic protozoans can also trigger TLRs enhancing resistance to them by triggering pro-inflammatory responses (6). Ten members of the TLR family have been identified in human, and 13 in mice (4). Signaling by TLRs involves five adaptor proteins known as MyD88, MyD88-adapter like (MAL), Toll/interleukin receptor (TIR) domain-containing adaptor protein inducing interferon  (TRIF), TRIF-related adaptor molecule (TRAM), and sterile ␣-and armadillo motifcontaining protein (SARM). MAL recruits MyD88 to TLR2 and TLR4, and TRAM recruits TRIF to TLR4 to allow interferon regulator factor 3 activation. SARM negatively regulates TRIF. MyD88 is the key signaling adaptor for all TLRs with the exception of TLR3 and a subset of TLR4 signals, which utilize Trif (7).Autophagy is a cellular response to starvation as well as a quality control system that can remove damaged organelles and long-lived proteins from cytoplasm. Autophagy plays a role in programmed cell death, the promotion and prevention of cancer, neurodegeneration, and cellular defense to remove invading pathogens (8). Recently, it has been recognized that autophagy is involved in both innate and adaptive immunity against intracellular protozoa, bacteria, and viruses (9 -11). Autophagy has been implicated in major histocompatibility complex class II antigen presentation. During autophagy cytoplasmic materials are sequestered into double membrane-coated autophagosomes, which fuse with endosomes and lysosomes to form autolysosomes where the contents...
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.