ABSTRACT
Since the outbreak of the SARS-CoV-2 pandemic, there have been intense structural studies on purified recombinant viral components and inactivated viruses. However, investigation of the SARS-CoV-2 infection in the native cellular context is scarce, and there is a lack of comprehensive knowledge on SARS-CoV-2 replicative cycle. Understanding the genome replication, assembly and egress of SARS-CoV-2, a multistage process that involves different cellular compartments and the activity of many viral and cellular proteins, is critically important as it bears the means of medical intervention to stop infection. Here, we investigated SARS-CoV-2 replication in Vero cells under the near-native frozen-hydrated condition using a unique correlative multi-modal, multi-scale cryo-imaging approach combining soft X-ray cryo-tomography and serial cryoFIB/SEM volume imaging of the entire SARS-CoV-2 infected cell with cryo-electron tomography (cryoET) of cellular lamellae and cell periphery, as well as structure determination of viral components by subtomogram averaging. Our results reveal at the whole cell level profound cytopathic effects of SARS-CoV-2 infection, exemplified by a large amount of heterogeneous vesicles in the cytoplasm for RNA synthesis and virus assembly, formation of membrane tunnels through which viruses exit, and drastic cytoplasm invasion into nucleus. Furthermore, cryoET of cell lamellae reveals how viral RNAs are transported from double-membrane vesicles where they are synthesized to viral assembly sites; how viral spikes and RNPs assist in virus assembly and budding; and how fully assembled virus particles exit the cell, thus stablishing a model of SARS-CoV-2 genome replication, virus assembly and egress pathways.
Subject(s)
COVID-19 , Severe Acute Respiratory SyndromeABSTRACT
Macrophages regulate protective immune responses to infectious microbes, but aberrant macrophage activation frequently drives pathological inflammation. To identify regulators of vigorous macrophage activation, we analyzed RNA-seq data from synovial macrophages and identified SLAMF7 as a receptor associated with a super-activated macrophage state in rheumatoid arthritis. We implicated IFN-gamma as a key regulator of SLAMF7 expression. Engaging this receptor drove an exuberant wave of inflammatory cytokine expression, and induction of TNF-alpha following SLAMF7 engagement amplified inflammation through an autocrine signaling loop. We observed SLAMF7-induced gene programs not only in macrophages from rheumatoid arthritis patients, but in gut macrophages from active Crohn's disease patients and lung macrophages from severe COVID-19 patients. This suggests a central role for SLAMF7 in macrophage super-activation with broad implications in pathology.