Ku70 Functions as an RNA Helicase to Regulate miR-124 Maturation and Neuronal Cell Differentiation / 生物化学与生物物理进展

ObjectiveHuman Ku70 protein mainly involves the non-homologous end joining (NHEJ) repair of double-stranded DNA breaks (DSB) through its DNA-binding properties, and it is recently reported having an RNA-binding ability. This paper is to explore whether Ku70 has RNA helicase activity and affects miRNA maturation. MethodsRNAs bound to Ku protein were analyzed by RNA immunoprecipitation sequencing (RIP-seq) and bioinfomatic anaylsis. The expression relationship between Ku protein and miRNAs was verified by Western blot (WB) and quantitative reverse transcriptase-polymerase chain reaction (qRT-PCR) assays. Binding ability of Ku protein to the RNAs was tested by biolayer interferometry (BLI) assay. RNA helicase activity of Ku protein was identified with EMSA assay. The effect of Ku70 regulated miR-124 on neuronal differentiation was performed by morphology analysis, WB and immunofluorescence assays with or without Zika virus (ZIKV) infection. ResultsWe revealed that the Ku70 protein had RNA helicase activity and affected miRNA maturation. Deficiency of Ku70 led to the up-regulation of a large number of mature miRNAs, especially neuronal specific miRNAs like miR-124. The knockdown of Ku70 promoted neuronal differentiation in human neural progenitor cells (hNPCs) and SH-SY5Y cells by boosting miR-124 maturation. Importantly, ZIKV infection reduced the expression of Ku70 whereas increased expression of miR-124 in hNPCs, and led to morphologically neuronal differentiation. ConclusionOur study revealed a novel function of Ku70 as an RNA helicase and regulating miRNA maturation. The reduced expression of Ku70 with ZIKV infection increased the expression of miR-124 and led to the premature differentiation of embryonic neural progenitor cells, which might be one of the causes of microcephaly.

Safety and superior immunogenicity of heterologous boosting with an RBD-based SARS-CoV-2 mRNA vaccine in Chinese adults

Homologous and heterologous booster with COVID-19 mRNA vaccines represent the most effective strategy to prevent the ongoing Omicron pandemic. The additional protection from these prototype SARS-CoV-2 S-targeting vaccine was attributed to the increased RBD-specific memory B cells with expanded potency and breadth. Herein, we show the safety and immunogenicity of heterologous boosting with the RBD-targeting mRNA vaccine AWcorna (also term ARCoV) in Chinese adults who have received two doses inactivated vaccine. The superiority over inactivated vaccine in neutralization antibodies, as well as the safety profile, support the use of AWcorna as heterologous booster in China.

SARS-CoV-2 infection causes transient olfactory dysfunction in mice

Olfactory dysfunction caused by SARS-CoV-2 infection represents as one of the most predictive and common symptoms in COVID-19 patients. However, the causal link between SARS-CoV-2 infection and olfactory disorders remains lacking. Herein we demonstrate intranasal inoculation of SARS-CoV-2 induces robust viral replication in the olfactory epithelium (OE), resulting in transient olfactory dysfunction in humanized ACE2 mice. The sustentacular cells and Bowmans gland cells in OE were identified as the major targets of SARS-CoV-2 before the invasion into olfactory sensory neurons. Remarkably, SARS-CoV-2 infection triggers cell death and immune cell infiltration, and impairs the uniformity of OE structure. Combined transcriptomic and proteomic analyses reveal the induction of antiviral and inflammatory responses, as well as the downregulation of olfactory receptors in OE from the infected animals. Overall, our mouse model recapitulates the olfactory dysfunction in COVID-19 patients, and provides critical clues to understand the physiological basis for extrapulmonary manifestations of COVID-19.

A proof of concept for neutralizing antibody-guided vaccine design against SARS-CoV-2

Mutations and transient conformational movements of receptor binding domain (RBD) that make neutralizing epitopes momentarily unavailable, present immune escape routes to SARS-CoV-2. To mitigate viral escape, we developed a cocktail of neutralizing antibodies (NAbs) targeting epitopes located on different domains of spike (S) protein. Screening of a library of monoclonal antibodies generated from peripheral blood mononuclear cells of COVID-19 convalescent patients yielded potent NAbs, targeting N-terminal domain (NTD) and RBD domain of S, effective at nM concentrations. Remarkably, combination of RBD-targeting NAbs and NTD-binding NAb, FC05, dramatically enhanced the neutralization potency in cell-based assays and animal model. Results of competitive SPR assays and cryo-EM structures of Fabs bound to S unveil determinants of immunogenicity. Combinations of immunogens, identified in NTD and RBD of S, when immunized in rabbits elicited potent protective immune responses against SARS-CoV-2. These results provide a proof-of-concept for neutralization-based immunogen design targeting SARS-CoV-2 NTD and RBD. One sentence summaryImmunogens identified in the NTD and RBD of the SARS-CoV-2 spike protein using a cocktail of non-competing NAbs when injected in rabbits elicited a potent protective immune response against SARS-CoV-2.

Impaired cellular immunity to SARS-CoV-2 in severe COVID-19 patients

The World Health Organization has declared SARS-CoV-2 virus outbreak a world-wide pandemic. Individuals infected by the virus exhibited different degrees of symptoms, the basis of which remains largely unclear. Currently, though convalescent individuals have been shown with both cellular and humoral immune responses, there is very limited understanding on the immune responses, especially adaptive immune responses, in patients with severe COVID-19. Here, we examined 10 blood samples from COVID-19 patients with acute respiratory distress syndrome (ARDS). The majority of them (70%) mounted SARS-CoV-2-specific humoral immunity with production of neutralizing antibodies. However, compared to healthy controls, the percentages and absolute numbers of both NK cells and CD8+ T cells were significantly reduced, accompanied with decreased IFN{gamma} expression in CD4+ T cells in peripheral blood from severe patients. Most notably, we failed in detecting SARS-CoV-2-specific IFN{gamma} production by peripheral blood lymphocytes from these patients. Our work thus indicates that COVID-19 patients with severe symptoms are associated with defective cellular immunity, which not only provides insights on understanding the pathogenesis of COVID-19, but also has implications in developing an effective vaccine to SARS-CoV-2.

Structural basis for neutralization of SARS-CoV-2 and SARS-CoV by a potent therapeutic antibody

The COVID-19 pandemic caused by the SARS-CoV-2 virus has resulted in an unprecedented public health crisis. There are no approved vaccines or therapeutics for treating COVID-19. Here we reported a humanized monoclonal antibody, H014, efficiently neutralizes SARS-CoV-2 and SARS-CoV pseudoviruses as well as authentic SARS-CoV-2 at nM level by engaging the S receptor binding domain (RBD). Importantly, H014 administration reduced SARS-CoV-2 titers in the infected lungs and prevented pulmonary pathology in hACE2 mouse model. Cryo-EM characterization of the SARS-CoV-2 S trimer in complex with the H014 Fab fragment unveiled a novel conformational epitope, which is only accessible when the RBD is in open conformation. Biochemical, cellular, virological and structural studies demonstrated that H014 prevents attachment of SARS-CoV-2 to its host cell receptors. Epitope analysis of available neutralizing antibodies against SARS-CoV and SARS-CoV-2 uncover broad cross-protective epitopes. Our results highlight a key role for antibody-based therapeutic interventions in the treatment of COVID-19. One sentence summaryA potent neutralizing antibody conferred protection against SARS-CoV-2 in an hACE2 humanized mouse model by sterically blocking the interaction of the virus with its receptor.

Rapid adaptation of SARS-CoV-2 in BALB/c mice: Novel mouse model for vaccine efficacy

Coronavirus disease 2019 (COVID-19) threatens global public health and economy. In order to develop safe and effective vaccines, suitable animal models must be established. Here we report the rapid adaption of SARS-CoV-2 in BALB/c mice, based on which a convenient, economical and effective animal model was developed. Specifically, we found that mouse-adapted SARS-CoV-2 at passage 6 (MACSp6) efficiently infected both aged and young wild-type BALB/c mice, resulting in moderate pneumonia as well as inflammatory responses. The elevated infectivity of MACSp6 in mice could be attributed to the substitution of a key residue (N501Y) in the receptorbinding domain (RBD). Using this novel animal model, we further evaluated the in vivo protective efficacy of an RBD-based SARS-CoV-2 subunit vaccine, which elicited highly potent neutralizing antibodies and conferred full protection against SARS-CoV-2 MACSp6 challenge. This novel mouse model is convenient and effective in evaluating the in vivo protective efficacy of SARS-CoV-2 vaccine. SummaryThis study describes a unique mouse model for SARS-CoV-2 infection and confirms protective efficacy of a SARS-CoV-2 RBD subunit vaccine.

Characterization of anti-viral immunity in recovered individuals infected by SARS-CoV-2

The WHO has declared SARS-CoV-2 outbreak a public health emergency of international concern. However, to date, there was hardly any study in characterizing the immune responses, especially adaptive immune responses to SARS-CoV-2 infection. In this study, we collected blood from COVID-19 patients who have recently become virus-free and therefore were discharged, and analyzed their SARS-CoV-2-specific antibody and T cell responses. We observed SARS-CoV-2-specific humoral and cellular immunity in the patients. Both were detected in newly discharged patients, suggesting both participate in immune-mediated protection to viral infection. However, follow-up patients (2 weeks post discharge) exhibited high titers of IgG antibodies, but with low levels of virus-specific T cells, suggesting that they may enter a quiescent state. Our work has thus provided a basis for further analysis of protective immunity to SARS-CoV-2, and understanding the pathogenesis of COVID-19, especially in the severe cases. It has also implications in designing an effective vaccine to protect and treat SARS-CoV-2 infection.

Laboratory confirmation of the first influenza A (H1N1) imported case in Mainland China / 病毒学报

The clinical throat swab specimen of an imported suspected case of influenza A (H1N1) was detec ted with real-time PCR, RT-PCR and subsequently confirmed by gene sequencing. The presence of influ enza A (H1N1) virus confirmed the first case with A (H1N1) infection in Mainland China.

Preparation and identification of dengue virus with deletion mutation of capsid protein / 南方医科大学学报

<p><b>OBJECTIVE</b>To generate rescued viruses with deletion mutation of capsid protein from dengue virus type 2 isolated in China (DEN2-43).</p><p><b>METHODS</b>On the basis of infectious full-length cDNA clone pD212 of DEN2-43 strain virus, the deletion mutants were constructed by fusion PCR, from which the rescued viruses with deletion mutation of capsid protein were generated by transcription in vitro and electroporation.</p><p><b>RESULT AND CONCLUSION</b>Sequence analysis demonstrated that the deletion mutations had been successfully inserted into the rescued viruses obtained. These mutant viruses may hold the key for elucidating the effects of deletion mutation of capsid protein on the biological characteristics of dengue virus.</p>