Recent advances in functional genomics for sugar beet (Beta vulgaris L.) improvement: progress in determining the role of BvSTI in pest resistance in roots

Abstract

To gain knowledge of root resistance mechanisms in sugar beet, Beta vulgaris L., our laboratory has been studying the interaction of sugar beet with its most devastating insect pest, the sugar beet root maggot (SBRM; Tetanops myopaeformis Roder). Damage from SBRM infestations is a serious problem and current control measures rely on environmentally damaging insecticides. We recently reported root-specific gene expression incited by SBRM feeding in a moderately resistant F1016 and a susceptible parental F1010 line. AcDNA expressed sequence tag (EST) coding for a serine (trypsin-type) protease inhibitor (BvSTI) was identified and investigated further here. BvSTI shares sequence similarity with a root-specific tomato gene whose expression is induced by insect feeding. Since serine proteases comprise the major digestive enzymes in root maggot midguts, we hypothesize BvSTI may be involved in resistance. To elucidate the functional role of BvSTI, its coding region was fused to the CaMV 35S promoter and constitutively expressed in sugar beet hairy roots and N. benthamiana plants. In BvSTI-transformed F1010 hairy roots, trypsin inhibitory activity increased 2 to 4-fold. Using a polyacrylamide gel assay, new trypsin-like PI activity was detected in BvSTI-N. benthamiana plants. Since SBRM cannot be reared in vitro, two other insects that utilize serine digestive proteases, fall armyworm (Spodoptera frugiperda) and tobacco hornworm (Manduca sexta), were screened for resistance. To date, we demonstrated that 1) fall armyworm will feed on sugar beet hairy roots and 2) tobacco hornworm fed BvSTI-N. benthamiana leaves had reduced weights and pupal sizes. These results suggest that BvSTI may contribute to the moderate resistance of F1016 roots to SBRM. Functional analysis of additional ESTs will further support efforts to characterize the components of sugar beet root resistance mechanisms.