Cloning, expression and subcellular distribution of a Rana grylio virus late gene encoding ERV1 homologue.
An essential for respiration and viability (ERV1) homologue, 88R, was cloned and characterized from Rana grylio virus (RGV). Database searches found its homologues in all sequenced iridoviruses, and sequence alignment revealed a highly conserved motif shared by all ERV1 family proteins: Cys-X-X-Cys. RT-PCR and western blot analysis revealed that 88R begins to transcribe and translate at 6 h postinfection (p.i.) and remains detectable at 48 h p.i. during RGV infection course. Furthermore, using drug inhibition analysis by a de novo protein synthesis inhibitor and a viral DNA replication inhibitor, RGV 88R was classified as a late (L) viral gene during the in vitro infection. 88R-EGFP fusion protein was observed in both the cytoplasm and nucleus of pEGFP-N3-88R transfected EPC cells. Although result of immunofluorescence is similar, 88R protein was not detected in viromatrix. Moreover, function of RGV 88R on virus replication were evaluated by RNAi assay. Nevertheless, effect of knockdown of RGV 88R expression on virus replication was not detected in cultured fish cell lines. Collectively, current data indicate that RGV 88R was a late gene of iridovirus encoding protein that distributed both the cytoplasm and nucleus.
Ke
Fei
F
State Key Laboratory of Freshwater Ecology and Biotechnology, Institute of Hydrobiology, Chinese Academy of Sciences, Graduate School of the Chinese Academy of Sciences, Wuhan, China.
Zhao
Zhe
Z
Zhang
Qiya
Q
D000595:Amino Acid Sequence; D000818:Animals; D015153:Blotting, Western; D002460:Cell Line; D003001:Cloning, Molecular; D015967:Gene Expression Regulation, Viral; D020868:Gene Silencing; D005814:Genes, Viral; D049452:Green Fluorescent Proteins / Q000378:metabolism; D017994:Iridovirus / Q000235:genetics* / Q000502:physiology; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D010802:Phylogeny; D021381:Protein Transport; D034741:RNA, Small Interfering / Q000378:metabolism; D011898:Ranidae / Q000821:virology*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D016415:Sequence Alignment; D020539:Sequence Analysis, Protein; D017386:Sequence Homology, Amino Acid; D013347:Subcellular Fractions / Q000378:metabolism; D014764:Viral Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D014779:Virus Replication
D034741:RNA, Small Interfering; D011993:Recombinant Fusion Proteins; D014764:Viral Proteins; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins