Design of antiviral AGO2-dependent short hairpin RNAs

The increasing emergence and re-emergence of RNA virus outbreaks underlines the urgent need to develop effective antivirals.RNA interference(RNAi)is a sequence-specific gene silencing mechanism that is triggered by small interfering RNAs(siRNAs)or short hairpin RNAs(shRNAs),which exhibits significant promise for antiviral therapy.AGO2-dependent shRNA(agshRNA)generates a single-stranded guide RNA and presents significant advantages over traditional siRNA and shRNA.In this study,we applied a logistic regression algorithm to a previously published chemically siRNA efficacy dataset and built a machine learning-based model with high predictive power.Using this model,we designed siRNA sequences targeting diverse RNA viruses,including human enterovirus A71(EV71),Zika virus(ZIKV),dengue virus 2(DENV2),mouse hepatitis virus(MHV)and severe acute respiratory syndrome coronavirus 2(SARS-CoV-2),and transformed them into agshRNAs.We validated the performance of our agshRNA design by evaluating antiviral efficacies of agshRNAs in cells infected with different viruses.Using the agshRNA targeting EV71 as an example,we showed that the anti-EV71 effect of agshRNA was more potent compared with the corresponding siRNA and shRNA.Moreover,the antiviral effect of agshRNA is dependent on AGO2-processed guide RNA,which can load into the RNA-induced silencing complex(RISC).We also confirmed the antiviral effect of agshRNA in vivo.Together,this work develops a novel antiviral strategy that combines machine learning-based algorithm with agshRNA design to custom design antiviral agshRNAs with high efficiency.

Omics study reveals abnormal alterations of breastmilk proteins and metabolites in puerperant women with COVID-19

Dear Editor,The nutrition contents of breastmilk directly participate in neonatal immune response.The alternations of the components of breastmilk under the context of viral infection not only reflect the physiological changes in mothers but also affect neonatal immunity and metabolism via breastfeeding.Herein,we attempted to answer the important questions whether breastmilk production is affected by COVID-19 and whether breastfeeding is still a safe or recommended operation for COVID-19 puerperant women.

STUB1 regulates antiviral RNAi through inducing ubiquitination and degradation of Dicer and AGO2 in mammals

RNA interference(RNAi)is an intrinsic antiviral immune mechanism conserved in diverse eukaryotic organisms.However,the mechanism by which antiviral RNAi in mammals is regulated is poorly understood.In this study,we uncovered that the E3 ubiquitin ligase STIP1 homology and U-box-containing protein 1(STUB1)was a new regulator of the RNAi machinery in mammals.We found that STUB1 interacted with and ubiquitinated AGO2,and targeted it for degradation in a chaperon-dependent manner.STUB1 promoted the formation of Lys48(K48)-linked polyubiquitin chains on AGO2,and facilitated AGO2 degradation through ubiquitin-proteasome system.In addition to AGO2,STUB1 also induced the protein degradation of AGO1,AGO3 and AGO4.Further investigation revealed that STUB1 also regulated Dicer's ubiquitination via K48-linked polyubiquitin and induced the degradation of Dicer as well as its specialized form,termed antiviral Dicer(avi Dicer)that expresses in mammalian stem cells.Moreover,we found that STUB1 deficiency up-regulated Dicer and AGO2,thereby enhancing the RNAi response and efficiently inhibiting viral replication in mammalian cells.Using the newborn mouse model of Enterovirus A71(EV-A71),we confirmed that STUB1 deficiency enhanced the virus-derived si RNAs production and antiviral RNAi,which elicited a potent antiviral effect against EV-A71 infection in vivo.In summary,our findings uncovered that the E3 ubiquitin ligase STUB1 was a general regulator of the RNAi machinery by targeting Dicer,avi Dicer and AGO1–4.Moreover,STUB1 regulated the RNAi response through mediating the abundance of Dicer and AGO2 during viral infection,thereby providing novel insights into the regulation of antiviral RNAi in mammals.