Study On The Regulation Of Host Lipid Factors In CMV Replication

Cucumber mosaic virus(CMV)is a positive single-stranded RNA virus,being a typical member of the genus Cucumovirus,the family Bromoviridae.CMV genome consists of three single-stranded,positive-sense RNAs.Existed studies have shown that CMV replication locates on the host tonoplast.It is totally unknown whether the tonoplast lipids regulate CMV replication.Thus,identification and analysis of lipid components of vacuolar membrane(tonoplast)has an important relevance for investigation of the molecular mechanism of CMV replication.To examine the effect of CMV infection on the lipid components of the host tonoplast,lipdomic profiles of tonoplasts were deteremined by mass spectrometry.In the mock and CMV-infected plants,a total of 481 lipids were identified and divided into 16 categories.Comparison of lipid profies between mock and CMV-infected tonoplasts showed that the sphingolipid content was upregulated by the infection of CMV.Next we quantitatively analyzed the expression levels of the genes related to lipid metabolism in Nicotiana benthamiana plants with or without CMV infection by real-time PCR.The results showed that some of genes were up-regualted,especially 3-Ketoshpinganine reductase(TSC10)with a highest increase.Subsequently,we down-regulated the mRNA level of TSC10 in N.benthamiana by TRV-mediated gene silencing,and tested its effect on CMV accumulation.Interestingy,the down-regulation of TSC10 reduced the accumulation level of CMV.Moreover,there was no significant change in the number of CMV replication vesicles on the tonoplast.However,there was no effects on the accumulation levels of tobacco mosaic virus and potato virus X,demonstrating that TSC10 plays a specific role in the regulation of CMV replication.Salicylic acid treatment up-regulated the expression of TSC10 in N.benthamiana plants,indicating that CMV up-regulation of Nb-TSCIO may be related to SA signaling pathway.Taken together,CMV infection induces expression of some genes involved in sphingolipid synthsis,presumably leading to increased content of sphingolipid components in tonoplasts,which positively regulate CMV replication.Our findings suggest that sphingolipids play an important role in CMV replicatioin.