Objective:This study aimed to investigate the regulatory role and mechanism of myosin heavy chain 9 (MYH9) in the invasion of Zika virus (ZIKV) into human glioma cells (U251).Methods:Utilizing CRISPR/Cas9 technology, MYH9-knockout U251 cells (U251-MYH9 KD) were constructed. Following ZIKV infection, the protein expression levels, RNA load, and viral titer of ZIKV were detected through western blot (WB), Real-time fluorescence quantitative polymerase chain reaction (qPCR), and plaque formation assays, respectively. The infection efficiency of ZIKV in U251 cells treated with the MYH9 inhibitor blebbistatin was assessed. The binding and internalization efficiency of ZIKV were measured in U251-MYH9 KD cells. The interaction between MYH9 and the ZIKV envelope protein (E) was studied using co-immunoprecipitation (Co-IP). The effects of soluble MYH9 recombinant protein and anti-human MYH9 antibodies on ZIKV infection were evaluated by qPCR and plaque formation assays. Results:It was found that knockout or inhibition of MYH9 significantly suppressed ZIKV infection in U251 cells. MYH9 knockout notably inhibited the binding and internalization of ZIKV in U251 cells. MYH9 interacted with the ZIKV E protein, and both MYH9 recombinant protein and anti-human MYH9 antibodies, by blocking the binding of ZIKV E protein to cell surface MYH9, inhibited ZIKV infection in U251 cells in a dose-dependent manner.Conclusions:MYH9 facilitates ZIKV invasion into U251 cells through interaction with the ZIKV E protein.

Objective:This study aimed to investigate the regulatory role and mechanism of myosin heavy chain 9 (MYH9) in the invasion of Zika virus (ZIKV) into human glioma cells (U251).Methods:Utilizing CRISPR/Cas9 technology, MYH9-knockout U251 cells (U251-MYH9 KD) were constructed. Following ZIKV infection, the protein expression levels, RNA load, and viral titer of ZIKV were detected through western blot (WB), Real-time fluorescence quantitative polymerase chain reaction (qPCR), and plaque formation assays, respectively. The infection efficiency of ZIKV in U251 cells treated with the MYH9 inhibitor blebbistatin was assessed. The binding and internalization efficiency of ZIKV were measured in U251-MYH9 KD cells. The interaction between MYH9 and the ZIKV envelope protein (E) was studied using co-immunoprecipitation (Co-IP). The effects of soluble MYH9 recombinant protein and anti-human MYH9 antibodies on ZIKV infection were evaluated by qPCR and plaque formation assays. Results:It was found that knockout or inhibition of MYH9 significantly suppressed ZIKV infection in U251 cells. MYH9 knockout notably inhibited the binding and internalization of ZIKV in U251 cells. MYH9 interacted with the ZIKV E protein, and both MYH9 recombinant protein and anti-human MYH9 antibodies, by blocking the binding of ZIKV E protein to cell surface MYH9, inhibited ZIKV infection in U251 cells in a dose-dependent manner.Conclusions:MYH9 facilitates ZIKV invasion into U251 cells through interaction with the ZIKV E protein.