Study Of Proteins Related With ISKNV Infection

Infectious spleen and kidney necrosis virus(ISKNV) is a species of the genusMegalocytivirus from the family Iridoviridae. This virus is one of the major agentscausing mortality and economic losses to the freshwater fish culture industry in Asiancountries, particularly that of the mandarin fish (Siniperca chuatsi). In this study, theinteraction of the mandarin fish caveolin-1protein (mCav-1) with the major capsidprotein of ISKNV was detected via pulldown and co-immunoprecipitation. For the firsttime, the interaction between the viral major capsid protein and mandarin fish caveolin-1was clearly demonstrated using in vivo and in vitro assays. This interaction wasindependent of the classic caveolin-1scaffolding domain(CSD), in which manysignaling proteins interacted. Immunofluorescence confocal microscopy showed that themajor capsid protein and mandarin fish caveolin-1were both internalized at theperinuclear area in the virus-infected mandarin fish fry cells at0h to4h of infection.Moreover, the results of subcellular fractionation showed that the major capsid proteinwas associated with caveolae/lipid rafts in the early stages of viral infection. RNAinterference silencing of mandarin fish caveolin-1did not change the binding of the virusto the cell, but efficiently inhibited the viral infection. These results suggest that the direct interaction between the major capsid protein of the infectious spleen and kidney necrosisvirus and mandarin fish caveolin-1is important for the viral invasion and may play animportant role in the early stages of infection.Pathogens often utilize endocytic and cytoplasmic transport mechanisms to infect ahost cell. Understanding how these cellular functions are exploited by microorganismsallows us to better define the basic biology of pathogenesis. Our results demonstrate thatinfection by ISKNV is inhibited by treatment of mandarin fish fry1(MFF-1) cells withcytochalasin D, cytochalasin B or latrunculin A. Furthermore, intact actin microfilamentswere required early during virus internalization. Finally, Real-time quantitative PCRrevealed that microfilamentslates and microtubules play a role at the late stages ofISKNV infection. The results indicate that microtubules and actin microfilaments arevital for ISKNV infection. The findings presented here provide new fundamentalinformation on the specificity and potential role of the actin and microtubule network iniridovirus infection, helping us better understand how iridoviruses exploit thecytoskeleton to facilitate their infection.