The dynamics of SARS-CoV-2 RNA structure and their functional relevance are largely unknown. Here we develop a simplified SPLASH assay and comprehensively map the in vivo RNA-RNA interactome of SARS-CoV-2 genome across viral life cycle. We report canonical and alternative structures including 5′-UTR and 3′-UTR, frameshifting element (FSE) pseudoknot and genome cyclization in both cells and virions. We provide direct evidence of interactions between Transcription Regulating Sequences, which facilitate discontinuous transcription. In addition, we reveal alternative short and long distance arches around FSE. More importantly, we find that within virions, while SARS-CoV-2 genome RNA undergoes intensive compaction, genome domains remain stable but with strengthened demarcation of local domains and weakened global cyclization. Taken together, our analysis reveals the structural basis for the regulation of replication, discontinuous transcription and translational frameshifting, the alternative conformations and the maintenance of global genome organization during the whole life cycle of SARS-CoV-2, which we anticipate will help develop better antiviral strategies.

Vaccines are a key weapon against the COVID-19 pandemic caused by SARS-CoV-2. However, there are inter-individual differences in immune response to SARS-CoV-2 vaccines and genetic contributions to these differences have barely been investigated. Here, we performed genome-wide association study (GWAS) of antibody levels in 168 inactivated SARS-CoV-2 vaccine recipients. A total of 177 SNPs, corresponding to 41 independent loci, were identified to be associated with IgG, total antibodies or neutral antibodies. Specifically, the rs4543780, the intronic variant of FAM89A gene, was associated with total antibodies level and was annotated as a potential regulatory variant affecting gene expression of FAM89A, a biomarker differentiating bacterial from viral infections in febrile children. These findings might advance our knowledge of the molecular mechanisms driving immunity to SARS-CoV-2 vaccine.

RNA species act as architectural scaffolds for nuclear structures including chromatin in eukaryotic cells. However, the composition and dynamics of tightly bound chromatin-associated RNAs during mitosis remains elusive. Here we report the identification of chromatin-enriched RNA (cheRNAs) by biochemical nuclear fractionation coupled with RNA sequencing in both interphase and mitotic phase of A549 and HeLa-S3 cell lines. We show that highly abundant cheRNAs, mostly small noncoding RNAs, are largely maintained in mitotic chromatin, and constitute a substantial part of chromatin RNA throughout cell cycle. We also show that the mitotic retained cheRNAs tend to be cell type nonspecific and might be involved in chromatin accessibility and epigenetic memory of gene expression control. Therefore, we reveal an unexpected set of cell type-nonspecific mitotic retained chromatin-enriched RNAs. We anticipate that the landscape of RNA composition of chromatin both in interphase and mitotic phase would help understanding structure and function of chromatin.

The COVID-19 pandemic has resulted in great morbidity and mortality worldwide and human genetic factors have been implicated in the susceptibility and severity of COVID-19. However, few replicate researches have been performed, and studies on associated genes mainly focused on genic regions while regulatory regions were a lack of in-depth dissection. Here, based on previously reported associated variants and genes, we designed a capture panel covering 1,238 candidate variants and 25 regulatory regions of 19 candidate genes and targeted-sequenced 96 mild and 145 severe COVID-19 patients. Genetic association analysis was conducted between mild and severe COVID-19 patients, between all COVID-19 patients and general population, or between severe COVID-19 patients and general population. A total of 49 variants were confirmed to be associated with susceptibility or severity of COVID-19 (p < 0.05), corresponding to 18 independent loci. Specifically, rs1799964 in the promoter of inflammation-related gene TNF, rs9975538 in the intron of interferon receptor gene IFNAR2, rs429358 in the exon of APOE, rs1886814 in the intron of FOXP4-AS1 and a list of variants in the widely reported 3p21.31 and ABO gene were confirmed. It is worth noting that, for the confirmed variants, the phenotypes of the cases and controls were highly consistent between our study and previous reports, and the confirmed variants identified between mild and severe patients were quite different from those identified between patients and general population, suggesting the genetic basis of susceptibility and severity of SARS-CoV-2 infection might be quite different. Moreover, we newly identified 67 significant associated variants in the 12 regulatory regions of 11 candidate genes (p < 0.05). Further annotation by RegulomeDB database and GTEx eQTL data filtered out two variants (rs11246060 and rs28655829) in the enhancer of broad-spectrum antiviral gene IFITM3 that might affect disease severity by regulating the gene expression. Collectively, we confirmed a list of previously reported variants and identified novel regulatory variants associated with susceptibility and severity of COVID-19, which might provide biological and clinical insights into COVID-19 pathogenesis and treatment.

Anthrax toxin causes anthrax pathogenesis and expression levels of ANTXR2 (anthrax toxin receptor 2) are strongly correlated with anthrax toxin susceptibility. Previous studies found that ANTXR2 transcript abundance varies considerably in individuals of different ethnic/geographical groups, but no eQTLs (expression quantitative trait loci) have been identified. By using 3C (chromatin conformation capture), CRISPR-mediated genomic deletion and dual-luciferase reporter assay, gene loci containing cis-regulatory elements of ANTXR2 were localized. Two SNPs (single nucleotide polymorphism) at the conserved CREB-binding motif, rs13140055 and rs80314910 in the promoter region of the gene, modulating ANTXR2 promoter activity were identified. Combining these two regulatory SNPs with a previously reported SNP, rs12647691, for the first time, a statistically significant correlation between human genetic variations and anthrax toxin sensitivity was observed. These findings further our understanding of human variability in ANTXR2 expression and anthrax toxin susceptibility.