Silencing CsMAP65-2 and CsMAP65-3 in cucumber reduces susceptibility to Meloidogyne incognita.

Root knot nematodes (RKNs) induce hypertrophy and cell proliferation within the vascular cylinders of host plants, leading to the formation of giant cells (GCs) that are enlarged, multinucleate cells with high metabolic activity. These GCs are formed through repeated karyokinesis without cytokinesis and are accompanied by significant changes in cytoskeleton organization. In this study, two microtubule-binding protein genes, CsMAP65-2 and CsMAP65-3, are upregulated in cucumber roots upon RKNs infection, specifically at 3, 96, and 120 hpi. GUS expression analysis further confirmed the induction of CsMAP65-2 and CsMAP65-3 in both roots and nematode-induced galls. Silencing CsMAP65-2 or CsMAP65-3 using VIGS technology led to a reduction in gall size and number, as well as a decrease in GCs number (24.98% for CsMAP65-2; 19.48% for CsMAP65-3) and area (6% for CsMAP65-2; 4% for CsMAP65-3), compared to control plants. Furthermore, qRT-PCR analysis revealed upregulation of CsMYC2、CsPR1、CsPAD4, and CsPDF1 in CsMAP65-2 silenced lines and upregulation of CsFRK1 in CsMAP65-3 silenced lines, while CsJAZ1 was downregulated in both silenced lines. These findings suggest that CsMAP65-2 and CsMAP65-3 are critical for GCs development during RKN infection and provide a foundation for breeding nematode-resistant cucumber varieties. This research also offers insights for developing sustainable nematode management strategies in gourd crop cultivation.