Although ACE2 is the primary receptor for Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) infection, a systematic assessment of host factors that regulate binding to SARS-CoV-2 spike protein has not been described. Here, we use whole-genome CRISPR activation to identify host factors controlling cellular interactions with SARS-CoV-2. Our top hit was a TLR-related cell surface receptor called leucine-rich repeat-containing protein 15 (LRRC15). LRRC15 expression was sufficient to promote SARS-CoV-2 spike binding where they form a cell surface complex. LRRC15 mRNA is expressed in human collagen-producing lung myofibroblasts and LRRC15 protein is induced in severe Coronavirus Disease 2019 (COVID-19) infection where it can be found lining the airways. Mechanistically, LRRC15 does not itself support SARS-CoV-2 infection, but fibroblasts expressing LRRC15 can suppress both pseudotyped and authentic SARS-CoV-2 infection in trans. Moreover, LRRC15 expression in fibroblasts suppresses collagen production and promotes expression of IFIT, OAS, and MX-family antiviral factors. Overall, LRRC15 is a novel SARS-CoV-2 spike-binding receptor that can help control viral load and regulate antiviral and antifibrotic transcriptional programs in the context of COVID-19 infection.
Abstract Although the pathophysiology of neurodegenerative diseases is distinct for each disease, considerable evidence suggests that a single manipulation, dietary restriction, is strikingly protective against a wide range of such diseases. Thus pharmacological mimetics of dietary restrictions could prove widely protective across a range of neurodegenerative diseases. The PPAR transcription complex functions to re-program gene expression in response to nutritional deprivation as well as in response to a wide variety of lipophilic compounds. In mammals there are three PPAR homologs, which dimerize with RXR homologs and recruit coactivators Pgc1-alpha and Creb-binding protein (Cbp). PPARs are currently of clinical interest mainly because PPAR activators are approved for use in humans to reduce lipidemia and to improve glucose control in Type 2 diabetic patients. However, pharmacological enhancement of the activity of the PPAR complex is neuroprotective across a wide variety of models for neuropathological processes, including stroke, Alzheimer’s disease, Parkinson’s disease, and Huntington’s disease. Conversely activity of the PPAR transcriptional complex is reduced in a variety of neuropathological processes. The main mechanisms mediating the neuroprotective effects of the PPAR transcription complex appear to be re-routing metabolism away from glucose metabolism and toward alternative subtrates, and reduction in inflammatory processes. Recent evidence suggests that the PPAR transcriptional complex may also mediate protective effects of dietary restriction on neuropathological processes. Thus this complex represents one of the most promising for the development of pharmacological treatment of neurodegenerative diseases.
Abstract Directed evolution (DE) is a process of mutation and artificial selection to breed biomolecules with new or improved activity 1,2 . DE platforms are primarily prokaryotic or yeast-based, and stable mutagenic mammalian systems have been challenging to establish and apply 3 . To this end, we developed PROTein Evolution Using Selection (PROTEUS), a new platform that uses chimeric virus-like vesicles (VLVs) to enable extended mammalian DE campaigns without loss of system integrity. This platform is stable and can generate sufficient diversity for DE in mammalian systems. Using PROTEUS, we altered the doxycycline responsiveness of tetracycline-controlled transactivators, generating a more sensitive TetON-4G tool for gene regulation. PROTEUS is also compatible with intracellular nanobody evolution, and we use it to design a DNA damage-responsive anti-p53 nanobody. Overall, PROTEUS is an efficient and stable platform to direct evolution of biomolecules within mammalian cells.
ATP ‐sensitive K + ( K ATP ) channels are expressed ubiquitously, but have diverse roles in various organs and cells. Their diversity can partly be explained by distinct tissue‐specific compositions of four copies of the pore‐forming inward rectifier potassium channel subunits ( K ir6.1 and/or K ir6.2) and four regulatory sulfonylurea receptor subunits ( SUR 1 and/or SUR 2). Channel function and/or subcellular localization also can be modified by the proteins with which they transiently or permanently interact to generate even more diversity. We performed a quantitative proteomic analysis of K ATP channel complexes in the heart, endothelium, insulin‐secreting min6 cells (pancreatic β‐cell like), and the hypothalamus to identify proteins with which they interact in different tissues. Glycolysis is an overrepresented pathway in identified proteins of the heart, min6 cells, and the endothelium. Proteins with other energy metabolic functions were identified in the hypothalamic samples. These data suggest that the metabolo‐electrical coupling conferred by K ATP channels is conferred partly by proteins with which they interact. A large number of identified cytoskeletal and trafficking proteins suggests endocytic recycling may help control K ATP channel surface density and/or subcellular localization. Overall, our data demonstrate that K ATP channels in different tissues may assemble with proteins having common functions, but that tissue‐specific complex organization also occurs.
Abstract Although ACE2 is the primary receptor for SARS-CoV-2 infection, a systematic assessment of factors controlling SARS-CoV-2 host interactions has not been described. Here we used whole genome CRISPR activation to identify host factors controlling SARS-CoV-2 Spike binding. The top hit was a Toll-like receptor-related cell surface receptor called leucine-rich repeat-containing protein 15 (LRRC15). LRRC15 expression was sufficient to promote SARS-CoV-2 Spike binding where it forms a cell surface complex with LRRC15 but does not support infection. Instead, LRRC15 functioned as a negative receptor suppressing both pseudotyped and live SARS-CoV-2 infection. LRRC15 is expressed in collagen-producing lung myofibroblasts where it can sequester virus and reduce infection in trans. Mechanistically LRRC15 is regulated by TGF-β, where moderate LRRC15 expression drives collagen production but high levels suppress it, revealing a novel lung fibrosis feedback circuit. Overall, LRRC15 is a master regulator of SARS-CoV-2, suppressing infection and controlling collagen production associated with “long-haul” COVID-19.
Abstract COVID-19 patients display a wide range of disease severity, ranging from asymptomatic to critical symptoms with high mortality risk. Our ability to understand the interaction of SARS-CoV-2 infected cells within the lung, and of protective or dysfunctional immune responses to the virus, is critical to effectively treat these patients. Currently, our understanding of cell-cell interactions across different disease states, and how such interactions may drive pathogenic outcomes, is incomplete. Here, we developed a generalizable workflow for identifying cells that are differentially interacting across COVID-19 patients with distinct disease outcomes and use it to examine five public single-cell RNA-seq datasets with a total of 85 individual samples. By characterizing the cell-cell interaction patterns across epithelial and immune cells in lung tissues for patients with varying disease severity, we illustrate diverse communication patterns across individuals, and discover heterogeneous communication patterns among moderate and severe patients. We further illustrate patterns derived from cell-cell interactions are potential signatures for discriminating between moderate and severe patients.
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