title: Characterization of the polyadenylation activity in a replicase complex from Bamboo mosaic virus-infected Nicotiana benthamiana plants.
abstract: Bamboo mosaic virus (BaMV) has a positive-sense single-stranded RNA genome with a 5' cap and a 3' poly(A) tail. To characterize polyadenylation activity in the BaMV replicase complex, we performed the in vitro polyadenylation with various BaMV templates. We conducted a polyadenylation activity assay for BaMV RNA by using a partially purified BaMV replicase complex. The results showed that approximately 200 adenylates at the 3' end of the RNA were generated on the endogenous RNA templates. Specific fractions derived from uninfected Nicotiana benthamiana plants enhanced the polyadenylation activity, implying that host factors are involved in polyadenylation. Furthermore, polyadenylation can be detected in newly synthesized plus-strand RNA in vitro when using the exogenous BaMV minus-strand minigenome. For polyadenylation on the exogenous plus-strand minigenome, the 3' end requires at least 4A to reach 22% polyadenylation activity. The results indicate that the BaMV replicase complex recognizes the 3' end of BaMV for polyadenylation.
authors.affiliation: Graduate Institute of Biotechnology, National Chung Hsing University, Taichung, Taiwan.
authors.forename: I-Hsuan
authors.initials: IH
authors.lastname: Chen
authors.forename: Jai-Hong
authors.initials: JH
authors.lastname: Cheng
authors.forename: Ying-Wen
authors.initials: YW
authors.lastname: Huang
authors.forename: Na-Sheng
authors.initials: NS
authors.lastname: Lin
authors.forename: Yau-Heiu
authors.initials: YH
authors.lastname: Hsu
authors.forename: Ching-Hsiu
authors.initials: CH
authors.lastname: Tsai
mesh_terms: D046911:Macromolecular Substances / Q000302:isolation & purification / Q000378:metabolism; D010936:Plant Extracts / Q000302:isolation & purification / Q000378:metabolism; D026723:Polyadenylation; D017863:Potexvirus / Q000502:physiology*; D012367:RNA, Viral / Q000378:metabolism*; D014026:Nicotiana / Q000821:virology*; D014764:Viral Proteins / Q000302:isolation & purification / Q000378:metabolism; D014779:Virus Replication
chemical_list: D046911:Macromolecular Substances; D010936:Plant Extracts; D012367:RNA, Viral; D014764:Viral Proteins
keywords: BaMV; In vitro; In vitro polyadenylation; Poly(A) length; Polyadenylation signal; Replicase; Replication assay