Effects of the splicing-specific novel proteins encoded by the 2.2 kb spliced variants of hepatitis B virus genome on gene expression profile in Huh7 hepatocytes

Objective To investigate the effects of the splicing-specific novel proteins encoded by the 2.2 kb singly(termed as TPss) and doubly(termed as TPds) spliced variants of hepatitis B virus genome on gene expression profile in Huh7 hepatocytes.Methods Coding region of TPss or TPds were amplified by PCR and cloned into vector pcDNA3.1/HisC.The recombinant vectors were separately transfected into Huh7 hepatocytes by FuGENE6 reagent.Total proteins and RNAs of transfected Huh7 cells were extracted,the expressed target proteins were detected by Western blot,and the gene expression alteration of the transfected Huh7 cells was evaluated by gene expression profile array assay followed by semi-quantitative RT-PCR.Results Recombinant vectors of pcDNA3.1/HisC-TPss and pcDNA3.1/HisC-TPds,which express well the TPss and TPds proteins respectively in Huh7 hepatocytes,were successfully constructed.Expression of TPss could up-regulate 4 genes including DGKβ,and down-regulate 5 genes including ZNF652,while the expression of TPds up-regulate 3 genes including MTSS1,and down-regulate WARS2 gene.Conclusion The splicing-specific novel proteins encoded by the 2.2 kb spliced variants of HBV may disturb cell function including the re-organization of the cystoskeleton and metabolism,which attribute to the abnormal cell proliferation,mobility and aberrant cellular metabolism.