Identification and Characterization of the Defensin-Like Gene Family of Grapevine

Defensins are a class of small and diverse cysteine-rich pro- teins found in plants, insects, and vertebrates, which share a common tertiary structure and usually exert broad-spec- trum antimicrobial activities. We used a bioinformatic ap- proach to scan the Vitis vinifera genome and identified 79 defensin-like sequences (DEFL) corresponding to 46 genes and allelic variants, plus 33 pseudogenes and gene frag- ments. Expansion and diversification of grapevine DEFL has occurred after the split from the last common ancestor with the genera Medicago and Arabidopsis. Grapevine DEFL localization on the ‘Pinot Noir’ genome revealed the presence of several clusters likely evolved through local du- plications. By sequencing reverse-transcription polymerase chain reaction products, we could demonstrate the expres- sion of grapevine DEFL with no previously reported rec- ord of expression. Many of these genes are predominantly or exclusively expressed in tissues linked to plant reproduc- tion, consistent with findings in other plant species, and some of them accumulated at fruit ripening. The tran- scripts of five DEFL were also significantly upregulated in tissues infected with Botrytis cinerea, a necrotrophic mold, suggesting a role of these genes in defense against this pathogen. Finally, three novel defensins were discovered among the identified DEFL. They inhibit B. cinerea conidia germination when expressed as recombinant proteins