Functional analysis of proteins encoded by banana bunchy top virus DNA-4 to-6

Green fluorescent protein (GFP)-tagging was used to determine the intracell ular localization pattern of the proteins encoded by banana bunchy top viru s (BBTV) DNA-3, -4 and -6, The protein encoded by BBTV DNA-4, which possess es a hydrophobic N terminus, was found to localize exclusively to the cell periphery while the proteins encoded by BBTV DNA-3 and -6 were found in bot h the nucleus and the cytoplasm, Go-expression of the DNA-4 protein and the proteins encoded by BBTV DNA-3 and -6 revealed that the DNA-4 protein was able to re-locate the DNA-6 protein, but not the DNA-3 protein, to the cell periphery. The 29 amino acid N-terminal hydrophobic region of the DNA-4 ge ne product appeared to be essential for specific localization of this prote in since deletion of this region abolished its ability to localize to the c ell periphery. These results indicate that BBTV may utilize a system analog ous to that of the begomoviruses with the BBTV DNA-6 protein acting as a nu clear shuttle protein (NSP) while the DNA-4 protein transports the NSP-DNA complexes to the cell periphery for intercellular transport. The protein en coded by BBTV DNA-5 was found to contain an LXCXE motif and yeast two-hybri d analysis revealed that the DNA-5 protein has retinoblastoma (Rb)-binding activity. This activity was dependent on an intact LXCXE motif since specif ic mutations to either the C or E residue completely abolished Rb-binding a ctivity. These results indicate that the gene product of BBTV DNA-5 is an R b-binding-like protein and may play an important role in host-cell cycle ma nipulation.