Preparation of BFV Gag antiserum and preliminary study on cellular distribution of BFV.
Viruses (e.g. Human immunodeficiency virus, Human simplex virus and Prototype foamy virus) are obligate intracellular parasites and therefore depend on the cellular machinery for cellular trafficking. Bovine foamy virus (BFV) is a member of the Spumaretrovirinae subfamily of Retroviruses, however, details of its cellular trafficking remain unknown. In this study, we cloned the BFV gag gene into prokaryotic expression vector pET28a and purified the denaturalized Gag protein. The protein was used to immunize BALB/c mouse to produce antiserum, which could specifically recognize the BFV Gag protein in BFV-infected cells through western blot assay. Additionally, these results demonstrated that both the optimal and suboptimal cleavage of Gag protein occur in BFV-infected cells. Subsequently, the Gag antiserum was used to investigate subcellular localization of BFV. In immunofluorescence microscopy assays, colocalization microtubules (MTs) and assembling viral particles were clearly observed, which implied that BFV may transport along cellular MTs in host cells. Furthermore, MTs-depolymerizing assay indicated MTs were required for the efficient replication of BFV. In conclusion, our study suggests that BFV has evolved the mechanism to hijack the cellular cytoskeleton for its replication.
Key Laboratory of Molecular Microbiology and Biotechnology, Ministry of Education, College of Life Sciences, Nankai University, Tianjin 300071, China.
Jian
J
Wang
Hong-yan
HY
Guo
Rui
R
Jia
Xuan
X
Xu
Juan
J
Tan
Yun-qi
YQ
Geng
Wen-tao
WT
Qiao
D000818:Animals; D000914:Antibodies, Viral / Q000302:isolation & purification; D002417:Cattle; D002478:Cells, Cultured; D003001:Cloning, Molecular; D019085:Fluorescent Antibody Technique, Direct; D015870:Gene Expression; D015683:Gene Products, gag / Q000235:genetics / Q000276:immunology* / Q000302:isolation & purification; D051379:Mice; D008807:Mice, Inbred BALB C; D008870:Microtubules / Q000821:virology; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification; D016092:Spumavirus / Q000502:physiology*; D014779:Virus Replication
D000914:Antibodies, Viral; D015683:Gene Products, gag; D011994:Recombinant Proteins