Resistance to cucumber mosaic virus (CMV) in melon (Cucumis melo L.) has been described in several exotic accessions and is controlled by a recessive resistance gene, cmv1, that encodes a vacuolar protein sorting 41 (CmVPS41). cmv1 prevents systemic infection by restricting the virus to the bundle sheath cells, preventing viral phloem entry. CmVPS41 from different resistant accessions carries two causal mutations, either a G85E change, found in Pat-81 and Freeman’s cucumber, or L348R, found in PI161375, cultivar Songwhan Charmi (SC). Here, we analyzed the subcellular localization of CmVPS41 in Nicotiana benthamiana and found differential structures in resistant and susceptible accessions. Susceptible accessions showed nuclear and membrane spots and many transvacuolar strands, whereas the resistant accessions showed many intravacuolar invaginations. These specific structures colocalized with late endosomes. Artificial CmVPS41 carrying individual mutations causing resistance in the genetic background of CmVPS41 from the susceptible variety Piel de Sapo (PS) revealed that the structure most correlated with resistance was the absence of transvacuolar strands. Coexpression of CmVPS41 with viral movement proteins, the determinant of virulence, did not change these localizations; however, infiltration of CmVPS41 from either SC or PS accessions in CMV-infected N. benthamiana leaves showed a localization pattern closer to each other, with up to 30% cells showing some membrane spots in the CmVPS41SC and fewer transvacuolar strands (reduced from a mean of 4 to 1–2) with CmVPS41PS. Our results suggest that the distribution of CmVPS41PS in late endosomes includes transvacuolar strands that facilitate CMV infection and that CmVPS41 re-localizes during viral infection.

Plant viruses need to use many host factors to establish infection. During the viral cycle, intracellular transport is fundamental to reach the plasmodesmata to enable cell-to-cell transport. Cucumovirus CMV (cucumber mosaic virus, CMV) can infect plants from most economically important crops. To identify additional host proteins involved in CMV movement in melon, we used the MP as a bait to screen a Yeast two-hybrid cDNA library from CMV-infected plants and identified a Niemann-Pick C1 (NPC1) protein as a novel MP interactor. NPC1 is a transmembrane protein involved in cholesterol transport in animal cells, but also in the infection by several viruses of different families. The identified clone from the melon NPC1 gene spans from exons 25 to 28 and includes two introns. Notably, deletion of the two introns and exon 28 does not impair the interaction capacity of the remaining peptide. The identified CmNPC1 gene maps to chromosome 11. In addition, the melon genome encodes a second copy of NPC1 in chromosome 7 (CmNPC1-C7), highly similar. Functional assays revealed that the interaction domain of CmNPC1-C7 also interacts with CMV MP, suggesting that both genes could have a role in CMV infection. This study represents the first report linking NPC1 to the infection process of a plant virus, expanding our understanding of plant–virus interactions.

Red leaf blotch of almond, a severe foliar disease caused by the hemibiotrophic ascomycete Polystigma amygdalinum, is an expanding threat to almond production. To understand the molecular basis of cultivar tolerance, we used dual RNA-seq and gene co-expression network analysis to compare the transcriptomic landscapes of ’Tarraco’ (susceptible) and ’Mardía’ (tolerant) almond cultivars. Our results reveal contrasting host transcriptomic architectures defining this plant-pathogen interaction. In ’Tarraco’, fungal colonization triggers a massive transcriptomic shift, characterized by the activation of a specialized functional module dominated by a strong upregulation of genes involved in cell wall degradation (e.g., pectin and xylan catabolic processes) and anthocyanin biosynthesis. In contrast, the tolerance response in ’Mardía’ is defined by a constitutively primed baseline and, with only a few genes upregulated upon infection, primarily linked to chitin and lignin metabolism. Comparative analysis of asymptomatic tissues identified intrinsic defense signatures in ’Mardía’ anchored by protein homeostasis and signaling machinery, while ’Tarraco’ prioritized translation-associated pathways. Cross-referencing these findings with a previously identified tolerance Quantitative Trait Locus on Linkage Group 3 identified a subset of high-priority positional candidate genes, including a receptor-like kinase FERONIA that was constitutively upregulated in the tolerant genotype. Furthermore, a de novo assembly of the fungal transcriptome revealed a candidate effector repertoire of 2,104 proteins, providing a first look into the pathogen’s molecular arsenal during colonization of the susceptible host.

Resistance to Cucumber mosaic virus (CMV) in melon has been described in several exotic accessions. It is controlled by a recessive resistance gene, cmv1 , which encodes a Vacuolar Protein Sorting 41 (CmVPS41). Cmv1 prevents systemic infection by restricting the virus to the bundle sheath cells, preventing viral phloem entry. CmVPS41 from different resistant accessions carried two causal mutations, either a G85E change, found in Pat-81 and Freeman’s Cucumber, or L348R found in PI161375 , cultivar Songwhan Charmi (SC). The analysis of the subcellular localization of CmVPS41 in N. benthamiana has revealed differential structures in resistant and susceptible accessions. Susceptible accessions showed nuclear and membrane spots and many transvacuolar strands, whereas the resistant accessions showed many intravacuolar invaginations. These specific structures colocalize with late endosomes. Artificial CmVPS41 carrying individual mutations causing resistance in the genetic background of CmVPS41 from the susceptible variety Piel de Sapo (PS), revealed that the structure most correlated with resistance was the absence of transvacuolar strands. Co-expression of CmVPS41 with the viral MPs, the determinant of virulence, did not change these localizations; however, infiltration of CmVPS41 from either SC or PS accessions in CMV-infected N. benthamiana leaves showed a localization pattern closer to each other, with up to 30% cells showing some membrane spots in the CmVPS41SC and fewer transvacuolar strands (from a mean of 4 to 1-2) with CmVPS41PS. Our results suggest that the distribution of CmVPS41PS in late endosomes includes transvacuolar strands that facilitate CMV infection and that CmVPS41 is re-localized during viral infection.