Structure and function of S9 segment of grass carp reovirus Anhui strain

A highly virulent grass carp reovirus (GCRV) strain, named GCRV-AH528, was recently purified from a diseased grass carp with hemorrhage disease in Anhui, China. GCRV-AH528 S9 segment was 1320 nucleotides in length and encoded a 418 amino acid VP6 protein. BLAST search showed that the VP6 protein owned a conserved domain belonging to the reoviral σ2 family. Phylogenetic analysis of VP6 presented that GCRV-AH528 belonged to GCRV genotype II, which was more closely related to Orthoreovirus than GCRV genotype I and genotype III. Further analysis revealed that GCRV-AH528 S9 and mammalian orthoreovirus S8 might have evolved from a common ancestral precursor and have identical mechanism in virus assembly. The expression level of vp6 gene was detected by quantitative real-time PCR (qRT-PCR). Over time, the expression level of vp6 gradually increased in Ctenopharyngodon idellus kidney cells. However, the level of vp6 expression in blood sharply increased at 4-6 days, and then decreased to a low level after GCRV-AH528 challenge (P < 0.05). The vp6 gene was detected in all tissues examined, whereas at relatively higher levels in blood, kidney, and liver (P < 0.05). The yeast two-hybrid (Y2H) system was used to identify VP6 self-interaction, while no interaction was detected in VP6-VP6. This study not only revealed the S9 segment structure and expression pattern but also analyzed the VP6 mechanism by yeast hybridization method. The present study provides valuable informations for further experimental design and investigation of VP6 functions.