Given the susceptibility of tomato plants to pests, the aim of the present study was to understand how hormones are involved in the formation of tomato natural defences against insect herbivory. Tomato hormone mutants, previously introgressed into the same genetic background of reference, were screened for alterations in trichome densities and allelochemical content. Ethylene, gibberellin, and auxin mutants indirectly showed alteration in trichome density, through effects on epidermal cell area. However, brassinosteroids (BRs) and jasmonates (JAs) directly affected trichome density and allelochemical content, and in an opposite fashion. The BR-deficient mutant dpy showed enhanced pubescence, zingiberene biosynthesis, and proteinase inhibitor expression; the opposite was observed for the JA-insensitive jai1-1 mutant. The dpy x jai1-1 double mutant showed that jai1-1 is epistatic to dpy, indicating that BR acts upstream of the JA signalling pathway. Herbivory tests with the poliphagous insect Spodoptera frugiperda and the tomato pest Tuta absoluta clearly confirmed the importance of the JA-BR interaction in defence against herbivory. The study underscores the importance of hormonal interactions on relevant agricultural traits and raises a novel biological mechanism in tomato that may differ from the BR and JA interaction already suggested for Arabidopsis.

ABSTRACT. Lack of continuous progress in Theobroma cacao (Malvaceae) breeding, especially associated with seed quality traits, requires more efficient selection methods based on genomic information. Reverse transcript quantitative PCR (RT-qPCR) has become the method of choice for gene expression analysis, but relative expression analysis requires various reference genes, which must be stable across various biological conditions. We sought suitable reference genes for various tissues of cacao, especially developing seeds. Ten potential reference genes were analyzed for stability at various stages of embryo development, leaves, stems, roots, flowers, and pod epicarp; seven of them were also evaluated in shoot tips treated either with hormones (salicylate; ethefon; methyl-jasmonate) or after inoculation with the fungus Moniliophthora perniciosa (Marasmiaceae sensu lato). For developing embryos, the three most stable genes were actin (ACT), polyubiquitin (PUB), and ribosomal protein L35 (Rpl35). In the analyses of various tissues, the most stable genes were malate dehydrogenase (MDH), glyceraldehyde 3-phosphate dehydrogenase (GAPDH), and acyl-carrier protein B (ACP B). GAPDH, MDH and tubulin (TUB) were the most appropriate for normalization when shoot apexes were treated with hormones, while ACT, TUB and Rpl35 were the most appropriate after inoculation with M. perniciosa. We conclude that for each plant system and biological or ontogenetical condition, there is a need to define suitable reference genes. This is the first report to define reference genes for expression studies in cacao.

BackgroundPhytophthora species secrete cytoplasmic effectors from a family named Crinkler (CRN), which are characterised by the presence of conserved specific domains in the N- and C-terminal regions. P. parasitica causes disease in a wide range of host plants, however the role of CRN effectors in these interactions remains unclear. Here, we aimed to: (i) identify candidate CRN encoding genes in P. parasitica genomes; (ii) evaluate the transcriptional expression of PpCRN (Phytophthora parasitica Crinkler candidate) during the P. parasitica interaction with Citrus sunki (high susceptible) and Poncirus trifoliata (resistant); and (iii) functionally characterize two PpCRNs in the model plant Nicotiana benthamiana.ResultsOur in silico analyses identified 80 putative PpCRN effectors in the genome of P. parasitica isolate ‘IAC 01/95.1’. Transcriptional analysis revealed differential gene expression of 20 PpCRN candidates during the interaction with the susceptible Citrus sunki and the resistant Poncirus trifoliata. We have also found that P. parasitica is able to recognize different citrus hosts and accordingly modulates PpCRNs expression. Additionally, two PpCRN effectors, namely PpCRN7 and PpCRN20, were further characterized via transient gene expression in N. benthamiana leaves. The elicitin INF-1-induced Hypersensitivity Response (HR) was increased by an additive effect driven by PpCRN7 expression, whereas PpCRN20 expression suppressed HR response in N. benthamiana leaves. Despite contrasting functions related to HR, both effectors increased the susceptibility of plants to P. parasitica.ConclusionsPpCRN7 and PpCRN20 have the ability to increase P. parasitica pathogenicity and may play important roles at different stages of infection. These PpCRN-associated mechanisms are now targets of biotechnological studies aiming to break pathogen’s virulence and to promote plant resistance.

Increasing evidence suggests that post-transcriptional regulation is a key player in the transition between mature pollen and the progamic phase (from pollination to fertilization). Nonetheless, the actors in this mRNA-based gene expression reprogramming are poorly understood. We demonstrate that the evolutionarily conserved RNA-binding protein (RBP) LARP6C is necessary for the transition from dry pollen to pollen tubes and the guided growth of pollen tubes towards the ovule in Arabidopsis thaliana. In dry pollen, LARP6C binds to transcripts encoding proteins that function in lipid synthesis and homeostasis, vesicular trafficking, and polarized cell growth. LARP6C also forms cytoplasmic granules that contain the poly(A) binding protein and possibly represent storage sites for translationally silent mRNAs. In pollen tubes, loss of LARP6C negatively affects the quantities and distribution of storage lipids, as well as vesicular trafficking. In Nicotiana benthamiana leaf cells and in planta, analysis of reporter mRNAs designed from the LARP6C target MGD2 provided evidence that LARP6C can shift from a repressor to an activator of translation when the pollen grain enters the progamic phase. We propose that LARP6C orchestrates the timely post-transcriptional regulation of a subset of mRNAs in pollen during the transition from the quiescent to active state and along the progamic phase to promote male fertilization in plants.