Protein-protein Interactions Between NSP And Proteins Of Banana Bunchy Top Virus

Banana is a perennial herb. It has a wide range of banana planting area in tropical and subtropical regions. Banana bunchy top virus (BBTV) is one of the members of Nanaviridae Babuvirus. BBTV lead to the banana bunchy top disease (BBTD) which is considered to be a major threat to the banana industry in the world.BBTV genome consists of at least six circular single stranded DNA components (BBTV DNA-1to6) each of1.1Kb. Each of the six DNA components associated with BBTV encodes at least one gene. BBTV DNA component6(BBTV DNA6) encodes a nuclear shuttle protein (NSP) which is17.4kD. To employ nuclear system of host to achieve replication and transcription many DNA plant viruses can transport the virus genome to the nucleus by NSP protein which is encoded by virus genome. Therefore, we hypothesize that the NSP encoded by BBTV DNA6may play an important role in the pathogenesis of viral infection through interacting with proteins of Banana bunchy top virus. In this study, we investigate the protein-protein interactions between NSP and proteins of BBTV through the yeast two hybrid system (YTHS) and bimolecular fluorescence complementation technology (BIFC). This study would pave the way to understand the mechanism of BBTV infection and pathogenesis.The research results are as follows:(1) Construct6bait vectors and6activation domain vectors of BBTV respectively as following:pGBKT7-DNA1, pGBKT7-DNA2, pGBKT7-DNA3, pGBKT7-DNA4, pGBKT7-DNA5, pGBKT7-DNA6, PGADT7-DNA1, pGADT7-DNA2, pGADT7-DNA3, pGADT7-DNA4, PGADT7-DNA5, pGADT7-DNA6.(2) Self activation experiments show that there are no self-activation in all bait proteins and activation domain proteins. Toxicity assays show all those proteins are nontoxic to host cells.(3) The results of YTHS show NSP not only interacts with itself but also CP as well as Clink.(4) To confirm the results of YTHS we construct the BIFC vectors as following2YN-DNA3,2YN-DNA5,2YN-DNA6and2YC-DNA6. After transforming into Agrobacterium cells co-infiltrate2YC-DNA6with2YN-DNA3,2YN-DNA5, as well as2YN-DNA6respectively. The observation of confocal laser scanning microscope shows that2YC-DNA6interacts with2YN-DNA3,2YN-DNA5as well as2YN-DNA6 respectively which are consistent with the results of YTHS. This shows that NSP do interacts with itself,CP as well as Clink respectively.According to the results of this study and literature reports, we hypothesize that the NSP encoded by BBTV DNA6may play an important role in the pathogenesis of viral infection through interacting with proteins CP, Clink as well as itself to transport these proteins into nucleus where virus replication and transcription take place. Our study may contribute to understand the mechanism of BBTV infection and pathogenesis.