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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.

Peroxiredoxins (Prx) catalyse the reduction of hydrogen peroxide (H2O2) and, in association with catalases and other peroxidases, may participate in signal transduction by regulating intercellular H2O2 concentration that in turn can control gene transcription and cell signaling. Using virus-induced-gene-silencing (VIGS), 2-Cys Peroxiredoxin (2CysPrx) family and type-II Peroxiredoxin B (PrxIIB) gene were silenced in Nicotiana benthamiana, to study the impact that the loss of function of each Prx would have in the antioxidant system under control (22 °C) and severe heat stress conditions (48 °C). The results showed that both Prxs, although in different organelles, influence the regeneration of ascorbate to a significant extent, but with different purposes. 2CysPrx affects abscisic acid (ABA) biosynthesis through ascorbate, while PrxIIB does it probably through the xanthophyll cycle. Moreover, 2CysPrx is key in H2O2 scavenging and in consequence in the regulation of ABA signaling downstream of reactive oxygen species and PrxIIB provides an important assistance for H2O2 peroxisome scavenges.

The resistance to a set of strains of Cucumber mosaic virus (CMV) in the melon accession PI 161375, cultivar 'Songwhan Charmi', is dependent on one recessive gene, cmv1, which confers total resistance, whereas a second set of strains is able to overcome it. We tested 11 strains of CMV subgroups I and II in the melon line SC12-1-99, which carries the gene cmv1, and showed that this gene confers resistance to strains of subgroup II only and that restriction is not related to either viral replication or cell-to-cell movement. This is the first time that a resistant trait has been correlated with CMV subgroups. Using infectious clones of the CMV strains LS (subgroup II) and FNY (subgroup I), we generated rearrangements and viral chimaeras between both strains and established that the determinant of virulence against the gene cmv1 resides in the first 209 amino acids of the movement protein, as this region from FNY is sufficient to confer virulence to the LS clone in the line SC12-1-99. A comparison of the sequences of the strains of both subgroups in this region shows that there are five main positions shared by all strains of subgroup II, which are different from those of subgroup I. Site-directed mutagenesis of the CMV-LS clone to substitute these residues for those of CMV-FNY revealed that a combination of four of these changes [the group 64-68 (SNNLL to HGRIA), and the point mutations R81C, G171T and A195I] was required for a complete gain of function of the LS MP in the resistant melon plant.

Cucumber mosaic virus (CMV) has the broadest host range among plant viruses, causing enormous losses in agriculture. In melon, strains of subgroup II are unable to establish a systemic infection in the near-isogenic line SC12-1-99, which carries the recessive resistance gene cmv1 from the accession PI 161375, cultivar ‘Songwhan Charmi’. Strains of subgroup I overcome cmv1 resistance in a manner dependent on the movement protein. We characterized the resistance conferred by cmv1 and established that CMV-LS (subgroup II) can move from cell to cell up to the veins in the inoculated leaf, but cannot enter the phloem. Immunogold labelling at transmission electron microscopy level showed that CMV-LS remains restricted to the bundle sheath (BS) cells in the resistant line, and does not invade vascular parenchyma or intermediary cells, whereas, in the susceptible line ‘Piel de Sapo’ (PS), the virus invades all vein cell types. These observations indicate that the resistant allele of cmv1 restricts systemic infection in a virus strain- and cell type-specific manner by acting as an important gatekeeper for virus progression from BS cells to phloem cells. Graft inoculation experiments showed that CMV-LS cannot move from the infected PS stock into the resistant cmv1 scion, thus suggesting an additional role for cmv1 related to CMV transport within or exit from the phloem. The characterization of this new form of recessive resistance, based on a restriction of virus systemic movement, opens up the possibility to design alternative approaches for breeding strategies in melon., This work was supported by grants AGL2009-12698-C02-01 and AGL2012-40130-C02-01 from the Spanish Ministry of Science and Innovation. CG-A was supported by grant BES-2010-030274 from the Spanish Ministry of Science and Innovation.

The complex resistance to cucumber mosaic virus (CMV) present in the exotic melon accession Sonwang Charmi PI161375 (SC) has been studied using two populations, a near-isogenic line (NIL) collection and a doubled haploid line (DHL) collection, both generated from a cross between SC and the cultivar Piel de Sapo as resistant and susceptible parents, respectively. The NIL collection had previously allowed us to describe a single recessive gene, cmv1, which conferred full resistance to CMV strains P9 and P104.82. Screening of the whole DHL population followed by quantitative trait locus (QTL) analysis revealed that resistance to the strains M6 and TL, both non-responsive to cmv1, was quantitative and governed by at least three QTLs. One of them, cmvqw12.1, co-located with cmv1 in linkage group (LG) XII. The QTL analysis mapped another two QTLs in LGIII (cmvqw3.1) and LGX (cmvqw10.1) and showed interaction between cmvqw12.1 and cmvqw3.1. Progeny of crosses between resistant DHLs carrying the three main QTLs showed complete resistance to the strain M6, validating the accuracy of the QTL analysis. However, in our screening, there were resistant DHLs carrying only two QTLs, suggesting that there are other regions involved in resistance to M6 and required when one of the main QTLs is missing. Therefore, resistance to CMV in melon SC is qualitative for some strains and quantitative for the rest. For this late resistance, cmv1 is necessary and explains most of the phenotypic variance, but it is not sufficient, and needs the interaction with other loci., The work was funded by grant AGL2009-12698-C02-01/AGR from the Spanish Ministerio de Ciencia e Innovación, and grant Consolider CDS2007-00036 from the Spanish Ministerio de Educación y Ciencia. CG-A was supported by Ministerio de Ciencia e Innovación. M.S. was supported by Junta de Ampliación de Estudios, from Consejo Superior de Investigaciones Científicas-doc. R.X.C. was supported by Brazilian Conselho Nacional de Desenvolvimento Científico e Tecnológico, CNPq.