Gene Hsc70-4 promotes the replication and proliferation of Bombyx mori nucleopolyhedrovirus

Abstract:

[Objective] Heat shock response is an important cellular defense strategy to environmental stresses. Identification of the function of heat shock protein in the process of baculovirus infection and revealing the molecular mechanism of its role will provide a theoretical basis for understanding the molecular basis of host-virus interaction. [Methods] Bmhsc70-4 gene was cloned by molecular cloning, and multiple sequence alignment was performed with BioEdit and GeneDoc. The Bmhsc70-4 gene was overexpressed by eukaryotic expression and knocked out by CRISPR/Cas9 gene editing system. The expression of the corresponding genes was detected by quantitative real-time PCR. The effect of Bmhsc70-4 gene on apoptosis was determined by detecting the activity of Caspase-9 and Caspase-3/7. The co-localization of BmHSC70-4 and BmIAP was verified by immunofluorescence, and their interaction was further verified by immunoprecipitation. [Results] The open reading frame of Bmhsc70-4 gene was 1950 bp, encoding 649 amino acids and it was highly conserved among insects. Bmhsc70-4 gene was up-regulated during BmNPV infection. Overexpression of Bmhsc70-4 gene could promote the proliferation of BmNPV, and knockout of Bmhsc70-4 gene inhibited the proliferation of BmNPV, indicating that the expression of Bmhsc70-4 gene was conducive to the proliferation of BmNPV. We confirmed that Bmhsc70-4 gene can inhibit the apoptosis of silkworm cells. Immunofluorescence assay showed that BmHSC70-4 and BmIAP were co-localized in the cytoplasm and immunocoprecipitation showed that they could interact with each other. Bmhsc70-4 gene could promote the expression of Bmiap gene during BmNPV infection. [Conclusion] Bmhsc70-4 gene had the function of inhibiting apoptosis of silkworm cells, and it could interact with BmIAP and promote BmNPV replication and proliferation during the process of BmNPV infecting silkworm cells.

Key words:Bombyx mori;BmNPV;heat shock response;apoptosis;Bmhsc70-4;Bmiap