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Flavonoids are secondary metabolites associated with plant seed coat and flower color. These compounds provide health benefits to humans as anti-inflammatory and antioxidant compounds. The expression of the late biosynthetic genes in the flavonoid pathway is controlled by a ternary MBW protein complex consisting of interfacing MYB, beta-helix-loop-helix (bHLH), and WD40 Repeat (WDR) proteins. P, the master regulator gene of the flavonoid expression in common bean (Phaseolus vulgaris L.), was recently determined to encode a bHLH protein. The T and Z genes control the distribution of color in bean seeds and flowers and have historically been considered regulators of the flavonoid gene expression. T and Z candidates were identified using reverse genetics based on genetic mapping, phylogenetic analysis, and mutant analysis. Domain and AlphaFold2 structure analyses determined that T encodes a seven-bladed β-propeller WDR protein, while Z encodes a R2R3 MYB protein. Deletions and SNPs in T and Z mutants, respectively, altered the 3D structure of these proteins. Modeling of the Z MYB/P bHLH/T WDR MBW complex identified interfacing sequence domains and motifs in all three genes that are conserved in dicots. One Z MYB motif is a possible beta-molecular recognition feature (β-MoRF) that only appears in a structured state when Z MYB is modeled as a component of a MBW complex. Complexes containing mutant T and Z proteins changed the interaction of members of the complex in ways that would alter their role in regulating the expression of genes in the flavonoid pathway., Competing Interests: Conflicts of interest The authors declare no conflict of interest., (© The Author(s) 2024. Published by Oxford University Press on behalf of The Genetics Society of America.)

Background: Common bean was one of the first crops that benefited from the development and utilization of molecular marker-assisted selection (MAS) for major disease resistance genes. Efficiency of MAS for breeding common bean is still hampered, however, due to the dominance, linkage phase, and loose linkage of previously developed markers. Here we applied in silico bulked segregant analysis (BSA) to the BeanCAP diversity panel, composed of over 500 lines and genotyped with the BARCBEAN_3 6K SNP BeadChip, to develop codominant and tightly linked markers to the I gene controlling resistance to Bean common mosaic virus (BCMV)., Results: We physically mapped the genomic region underlying the I gene. This locus, in the distal arm of chromosome Pv02, contains seven putative NBS-LRR-type disease resistance genes. Two contrasting bulks, containing BCMV host differentials and ten BeanCAP lines with known disease reaction to BCMV, were subjected to in silico BSA for targeting the I gene and flanking sequences. Two distinct haplotypes, containing a cluster of six single nucleotide polymorphisms (SNP), were associated with resistance or susceptibility to BCMV. One-hundred and twenty-two lines, including 115 of the BeanCAP panel, were screened for BCMV resistance in the greenhouse, and all of the resistant or susceptible plants displayed distinct SNP haplotypes as those found in the two bulks. The resistant/susceptible haplotypes were validated in 98 recombinant inbred lines segregating for BCMV resistance. The closest SNP (~25-32 kb) to the distal NBS-LRR gene model for the I gene locus was targeted for conversion to codominant KASP (Kompetitive Allele Specific PCR) and CAPS (Cleaved Amplified Polymorphic Sequence) markers. Both marker systems accurately predicted the disease reaction to BCMV conferred by the I gene in all screened lines of this study., Conclusions: We demonstrated the utility of the in silico BSA approach using genetically diverse germplasm, genotyped with a high-density SNP chip array, to discover SNP variation at a specific targeted genomic region. In common bean, many disease resistance genes are mapped and their physical genomic position can now be determined, thus the application of this approach will facilitate further development of codominant and tightly linked markers for use in MAS.