부산대학교 도서관

Cell-cell fusion (abbreviated as cell fusion) is a characteristic pathology of medically important viruses, including varicella-zoster virus (VZV), the causative agent of chickenpox and shingles. Cell fusion is mediated by a complex of VZV glycoproteins, gB and gH-gL, and must be tightly regulated to enable skin pathogenesis based on studies with gB and gH hyperfusogenic VZV mutants. Although the function of gB and gH-gL in the regulation of cell fusion has been explored, whether host factors are directly involved in this regulation process is unknown. Here, we discovered host factors that modulated VZV gB/gH-gL mediated cell fusion via high-throughput screening of bioactive compounds with known cellular targets. Two structurally related non-antibiotic macrolides, tacrolimus and pimecrolimus, both significantly increased VZV gB/gH-gL mediated cell fusion. These compounds form a drug-protein complex with FKBP1A, which binds to calcineurin and specifically inhibits calcineurin phosphatase activity. Inhibition of calcineurin phosphatase activity also enhanced both herpes simplex virus-1 fusion complex and syncytin-1 mediated cell fusion, indicating a broad role of calcineurin in modulating this process. To characterize the role of calcineurin phosphatase activity in VZV gB/gH-gL mediated fusion, a series of biochemical, biological and infectivity assays was performed. Pimecrolimus-induced, enhanced cell fusion was significantly reduced by shRNA knockdown of FKBP1A, further supporting the role of calcineurin phosphatase activity in fusion regulation. Importantly, inhibition of calcineurin phosphatase activity during VZV infection caused exaggerated syncytia formation and suppressed virus propagation, which was consistent with the previously reported phenotypes of gB and gH hyperfusogenic VZV mutants. Seven host cell proteins that remained uniquely phosphorylated when calcineurin phosphatase activity was inhibited were identified as potential downstream factors involved in fusion regulation. These findings demonstrate that calcineurin is a critical host cell factor pivotal in the regulation of VZV induced cell fusion, which is essential for VZV pathogenesis. Author summary: VZV is a medically important human virus that causes varicella (chicken pox) and reactivates as zoster (shingles). Postherpetic neuralgia (PHN) is a common sequela of zoster, a persistent nerve pain that can last from months to years after zoster. Refractory to antiviral drugs and pain treatment, PHN significantly impacts quality of life. Despite the success of VZV live-attenuated vaccines in the healthy population, immunocompromised individuals are still vulnerable to significant morbidity. The recently approved subunit zoster vaccine is effective but whether it works against varicella is not known. Therefore, a better understanding of the molecular basis of VZV pathogenesis would provide significant advances towards alternative treatments. A hallmark of VZV pathogenesis is syncytia formation observed in infected skin and nerve ganglia. VZV-induced cell fusion is mediated by the viral fusion complex comprised of glycoproteins gB, gH, and gL. Dysregulation of this fusion process leads to impaired VZV infection in skin. Here, we report that inhibition of a cellular phosphatase, calcineurin, significantly increased gB/gH-gL mediated cell fusion, and intensified syncytia formation during VZV replication, but suppressed virus spread. Our study demonstrated that calcineurin phosphatase activity regulates VZV-induced cell fusion, providing a new perspective for potential antiviral strategies that target host factors. [ABSTRACT FROM AUTHOR]