A nematode-inducible promoter can effectively drives RNAi construct to confer Meloidogyne incognita resistance in tomato.

Abstract

Heterologous expression of a nematode-responsive promoter in tomato successfully driven the RNAi constructs to impart root-knot nematode resistance. The root-knot nematode Meloidogyne incognita seriously afflicts the global productivity of tomatoes. Nematode management options are extremely reliant on chemical methods, however, only a handful of nematicides are commercially available. Additionally, nematodes have developed resistance-breaking phenotypes against the commercially available Mi gene-expressing tomatoes. Nematode resistance in crop plants can be enhanced using the bio-safe RNAi technology, in which plants are genetically modified to express nematode gene-specific dsRNA/siRNA molecules. However, the majority of the RNAi crops conferring nematode tolerance have used constitutive promoters, which have many limitations. In the present study, using promoter-GUS fusion, we functionally validated two nematode-inducible root-specific promoters (pAt1g74770 and pAt2g18140, identified from Arabidopsis thaliana) in the Solanum lycopersicum-M. incognita pathosystem. pAt2g18140 was found to be nematode-responsive during 10-21days post-inoculation (dpi) and became non-responsive during the late infection stage (28 dpi). In contrast, pAt1g74770 remained nematode-responsive for a longer duration (10-28 dpi). Next, a number of transgenic lines were developed that expressed RNAi constructs (independently targeting the M. incognita integrase and splicing factor genes) driven by the pAt1g74770 promoter. M. incognita parasitic success (measured by multiplication factor ratio) in pAt1g74770:integrase and pAt1g74770:splicing factor RNAi lines were significantly reduced by 60.83-74.93% and 69.34-75.31%, respectively, compared to the control. These data were comparable with the RNAi lines having CaMV35S as the promoter. Further, a long-term RNAi effect was evident, because females extracted from transgenic lines were of deformed shape with depleted transcripts of integrase and splicing factor genes. We conclude that pAt1g74770 can be an attractive alternative to drive localized expression of RNAi constructs rather than using a constitutive promoter. The pAt1g74770-driven gene silencing system can be expanded into different plant-nematode interaction models.