Exploring the genetic variability and evolution of Bowman-Birk inhibitors in Vigna unguiculata (L.) Walp

Domesticated plants possess only a small fraction of the genetic variability found in their wild relatives, and this variability represents an important resource. One of these plants is Vigna unguiculata (L.) Walp., or cowpea, an important crop which has several features suitable for sustainable agriculture. Bowman-Birk protease inhibitors (BBI) are useful molecules produced by plants, especially Fabaceae and Poaceae, as defensive mechanism against insects attack. Nutraceutical effects of these proteins on humans have been demonstrated, including chemopreventive and anticarcinogenic ones. In this study we analyzed the natural genetic variability of two BBI isoforms of Vigna unguiculata in both wild and domesticated accessions. In particular the two isoforms encode for a trypsin-trypsin BBI, characterized by two trypsin-specific domains (BBI-TT) and the second one, a trypsin-chymotrypsin BBI, characterized by one trypsin and one chymotrypsin-specific domain (BBI-TC). For this purpose, both genes were sequenced from about 450 individuals belonging to 124 wild and 67 domesticated accessions, obtaining 24 haplotypes corresponding to 13 isoforms of BBI-TT and 29 haplotypes corresponding to 20 isoforms of BBI-TC. Most of the haplotypes found belonged exclusively to wild accessions plants. For both genes, haplotypes and isoforms distributions were mapped, and network analyses were performed to understand haplotypes relationships. Furthermore, sites under selective pressure were identified with different algorithm approaches. This work allowed the detection of several natural forms of BBI, found mostly in wild plants, that could potentially be used to improve the defenses of cultivated plants or for pharmaceutical and nutritional purposes.