The RNA polymerase of cytoplasmically replicating Zika virus binds with chromatin DNA in nuclei and regulates host gene transcription.

Zika virus (ZIKV) targets the neural progenitor cells (NPCs) in brain during intrauterine infections and consequently causes severe neurological disorders, such as microcephaly in neonates. Although replicating in the cytoplasm, ZIKV dysregulates the expression of thousands of host genes, yet the detailed mechanism remains elusive. Herein, we report that ZIKV encodes a unique DNA-binding protein to regulate host gene transcription in the nucleus. We found that ZIKV NS5, the viral RNA polymerase, associates tightly with host chromatin DNA through its methyltransferase domain and this interaction could be specifically blocked by GTP. Further study showed that expression of ZIKV NS5 in human NPCs markedly suppressed the transcription of its target genes, especially the genes involved in neurogenesis. Mechanistically, ZIKV NS5 binds onto the gene body of its target genes and then blocks their transcriptional elongation. The utero electroporation in pregnant mice showed that NS5 expression significantly disrupts the neurogenesis by reducing the number of Sox2- and Tbr2-positive cells in the fetal cortex. Together, our findings demonstrate a molecular clue linking to the abnormal neurodevelopment caused by ZIKV infection and also provide intriguing insights into the interaction between the host cell and the pathogenic RNA virus, where the cytoplasmic RNA virus encodes a DNA-binding protein to control the transcription of host cell in the nuclei.

Non-Structural Protein 5 of Zika Virus Interacts with p53 in Human Neural Progenitor Cells and Induces p53-Mediated Apoptosis.

Zika virus (ZIKV) infection could disrupt neurogenesis and cause microcephaly in neonates by targeting neural progenitor cells (NPCs). The tumor suppressor p53-mediated cell cycle arrest and apoptotic cell death have been suggested to be activated upon ZIKV infection, yet the detailed mechanism is not well understood. In the present study, we investigated the effects of ZIKV-encoded proteins in the activation of p53 signaling pathway and found that, among the ten viral proteins, the nonstructural protein 5 (NS5) of ZIKV most significantly activated the transcription of p53 target genes. Using the immunoprecipitation-coupled mass spectrometry approach, we identified that ZIKV-NS5 interacted with p53 protein. The NS5-p53 interaction was further confirmed by co-immunoprecipitation and GST pull-down assays. In addition, the MTase domain of NS5 and the C-terminal domain of p53 were mapped to be responsible for the interaction between these two proteins. We further showed that ZIKV-NS5 was colocalized with p53 and increased its protein level in the nuclei and able to prolong the half-life of p53. Furthermore, lentivirus-mediated expression of ZIKV-NS5 in hNPCs led to an apparent cell death phenotype. ZIKV-NS5 promoted the cleavage of PARP1 and significantly increased the cell apoptosis of hNPCs. Taken together, these findings revealed that ZIKV-NS5 is a previously undiscovered regulator of p53-mediated apoptosis in hNPCs, which may contribute to the ZIKV-caused abnormal neurodevelopment.

Endogenous reverse transcriptase and RNase H-mediated antiviral mechanism in embryonic stem cells.

Nucleic acid-based systems play important roles in antiviral defense, including CRISPR/Cas that adopts RNA-guided DNA cleavage to prevent DNA phage infection and RNA interference (RNAi) that employs RNA-guided RNA cleavage to defend against RNA virus infection. Here, we report a novel type of nucleic acid-based antiviral system that exists in mouse embryonic stem cells (mESCs), which suppresses RNA virus infection by DNA-mediated RNA cleavage. We found that the viral RNA of encephalomyocarditis virus can be reverse transcribed into complementary DNA (vcDNA) by the reverse transcriptase (RTase) encoded by endogenous retrovirus-like elements in mESCs. The vcDNA is negative-sense single-stranded and forms DNA/RNA hybrid with viral RNA. The viral RNA in the heteroduplex is subsequently destroyed by cellular RNase H1, leading to robust suppression of viral growth. Furthermore, either inhibition of the RTase activity or depletion of endogenous RNase H1 results in the promotion of virus proliferation. Altogether, our results provide intriguing insights into the antiviral mechanism of mESCs and the antiviral function of endogenized retroviruses and cellular RNase H. Such a natural nucleic acid-based antiviral mechanism in mESCs is referred to as ERASE (endogenous RTase/RNase H-mediated antiviral system), which is an addition to the previously known nucleic acid-based antiviral mechanisms including CRISPR/Cas in bacteria and RNAi in plants and invertebrates.