Coding Mutations in Vacuolar Protein-Sorting 4 AAA+ ATPase Endosomal Sorting Complexes Required for Transport Protein Homologs Underlie bc-2 and New bc-4 Gene Conferring Resistance to Bean Common Mosaic Virus in Common Bean

Bean common mosaic virus (BCMV) is a major disease in common bean (Phaseolus vulgaris L.). Host plant resistance is the most effective strategy to minimize crop damage against BCMV and the related Bean common mosaic necrosis virus (BCMNV). To facilitate breeding for resistance, we sought to identify candidate genes and develop markers for the bc-2 gene and the unknown gene with which it interacts. Genome-wide association study (GWAS) of the Durango Diversity Panel (DDP) identified a peak region for bc-2 on chromosome Pv11. Haplotype mapping narrowed the bc-2 genomic interval and identified Phvul.011G092700, a vacuolar protein-sorting 4 (Vps4) AAA+ ATPase endosomal sorting complexes required for transport (ESCRT) protein, as the bc-2 candidate gene. The race Durango Phvul.011G092700 gene model, bc-2[UI111], contains a 10-kb deletion, while the race Mesoamerican bc-2[Robust] consists of a single nucleotide polymorphism (SNP) deletion. Each mutation introduces a premature stop codon, and they exhibit the same interaction with the pathogroups (PGs) tested. Phvul.005G125100, another Vps4 AAA+ ATPase ESCRT protein, was identified as the candidate gene for the new recessive bc-4 gene, and the recessive allele is likely an amino acid substitution in the microtubule interacting and transport (MIT) domain. The two Vps4 AAA+ ATPase ESCRT proteins exhibit high similarity to the Zym Cucsa.385040 candidate gene associated with recessive resistance to Zucchini yellow mosaic virus in cucumber. bc-2 alone has no resistance effect but, when combined with bc-4, provides resistance to BCMV (except PG-V) but not BCMNV, and, when combined with bc-ud, provides resistance to BCMV (except BCMV PG-VII) and BCMNV. So instead of different resistance alleles (i.e., bc-2 and bc-22), there is only bc-2 with a differential reaction based on whether it is combined with bc-4 or bc-ud, which are tightly linked in repulsion. The new tools and enhanced understanding of this host-virus pathogen interaction will facilitate breeding common beans for resistance to BCMV and BCMNV.

Vps4-A (vacuolar protein sorting 4-A) is a binding partner for a novel Rho family GTPase, Rnd2

Rho family GTPases are implicated in a variety of biological activities, including endocytic vesicle trafficking. Rnd2 is a new member of Rho family GTPases, but its biological functions are not known. In the present study, we have performed a yeast two-hybrid screening using Rnd2 as bait and revealed that Rnd2 binds specifically to Vps4-A (where Vsp4-A is vacuolar protein sorting 4-A), a member of the AAA ATPase family and a central regulator for early endosome trafficking. This interaction was determined by the yeast two-hybrid system, in vitro binding and co-immunoprecipitation studies. Vps4-A associated with both guanosine 5′-[β-thio]triphosphate-bound active and guanosine 5′-[β-thio]diphosphate-bound inactive forms of Rnd2. An ATPase-defective Vps4-A mutant, Vps4-AE228Q, expressed in HeLa cells was accumulated in the early endosomes. When Rnd2 was co-expressed with Vps4-AE228Q, Rnd2 was recruited to the Vps4-A-bound early endosomes. These results suggest that Rnd2 is involved in the regulation of endosomal trafficking via direct binding to Vps4-A.