CqStathmin negatively regulates white spot syndrome virus infection by destabilizing microtubules in Cherax quadricarinatus

Abstract

Stathmin is a microtubule-destabilizing protein that disrupts microtubule dynamics by promoting the depolymerization of microtubules or preventing the polymerization of tubulin heterodimers, thereby performing various cellular functions. In this study, a CqStathmin in the invertebrate crayfish was found to be involved in the infection of white spot syndrome virus (WSSV), a DNA virus susceptible to shrimp. The CqStathmin was identified from the red claw crayfish Cherax quadricarinatus, sharing the conserved domain and phylogenetic relationship with stathmin homologs of other species. The expression of CqStathmin was significantly upregulated in crayfish hematopoietic tissue (Hpt) cells after WSSV infection. CqStathmin silencing promoted WSSV infection in Hpt cells. Further evidence indicated that silencing CqStathmin promoted the polymerization of crayfish microtubule structural proteins Cqα-Tubulin and Cqβ-Tubulin, whereas overexpression of CqStathmin facilitated their depolymerization. Silencing of CqStathmin increased the acetylation of Cqα-tubulin in Hpt cells, a post-translational modification that maintains microtubule integrity, thereby promoting microtubule polymerization. In contrast, overexpression of CqStathmin reduced the acetylation level of α-tubulin in HEK293T cells. Then, the binding of Cqα-Tubulin to Cqβ-Tubulin, as well as the interaction of CqStathmin with Cqα-Tubulin and/or Cqβ-Tubulin, along with their co-localization, have been confirmed. These results indicated that CqStathmin has a negative regulatory role in microtubule polymerization in crayfish. Moreover, WSSV infection was significantly decreased by Cqα/β-Tubulin silencing, suggesting that WSSV infection depended on microtubules in Hpt cells. Collectively, our results revealed that CqStathmin inhibited WSSV infection by regulating microtubule depolymerization in the crayfish. This is the first study to report that CqStathmin from an aquatic crustacean was involved in WSSV infection, enhancing our understanding of the WSSV infection mechanism.