Jérôme Franchel

Cloning and Characterization of a New Polyol Transporter (HbPLT2) in Hevea brasiliensis

Quebrachitol is a cyclic polyol and, along with sucrose, is one of the main sugars in Hevea latex. However, in contrast to sucrose, the mechanism and regulation of quebrachitol absorption is still unknown. Screening a latex-derived cDNA library using polyol transporter-specific probes, two full-length cDNAs were isolated, and named HbPLT1 and HbPLT2 (for Hevea brasiliensis polyol transporter 1 and 2, respectively). Their respective sequences exhibited close similarity with the previously cloned acyclic sugar polyol transporters, and shared the main features of the major facilitative superfamily. The functional activity of one of the cDNAs was determined by using an HbPLT2-complemented yeast strain. These strains displayed a marginal absorption of cyclic (inositol) and acyclic (mannitol and sorbitol) polyol but no absorption of sucrose, hexose and glycerol. Active absorption for xylitol was detected, and was competitively inhibited by quebrachitol. HbPLT1 and HbPLT2 expression patterns varied in response to different stimuli. Bark treatment with ethylene resulted in an early and significant up-regulation of HbPLT2 transcripts in laticifers as well as in inner bark cells, when compared with HbPLT1. Other treatments, especially mechanical wounding, strongly induced HbPLT2 transcripts. These data were consistent with the presence of ethylene and a wound-responsive regulatory cis-element on the sequence of the HbPLT2 promoter. All these findings together with those recently obtained for sucrose transporters and aquaporins are discussed in relation to the different roles for quebrachitol in Hevea brasiliensis.

Positional cloning of a candidate gene for resistance to the sunflower downy mildew, Plasmopara halstedii race 300

The resistance of sunflower to Plasmopara halstedii is conferred by major resistance genes denoted Pl. Previous genetic studies indicated that the majority of these genes are clustered on linkage groups 8 and 13. The Pl6 locus is one of the main clusters to have been identified, and confers resistance to several P. halstedii races. In this study, a map-based cloning strategy was implemented using a large segregating F2 population to establish a fine physical map of this cluster. A marker derived from a bacterial artificial chromosome (BAC) clone was found to be very tightly linked to the gene conferring resistance to race 300, and the corresponding BAC clone was sequenced and annotated. It contains several putative genes including three toll-interleukin receptor-nucleotide binding site-leucine rich repeats (TIR-NBS-LRR) genes. However, only one TIR-NBS-LRR appeared to be expressed, and thus constitutes a candidate gene for resistance to P. halstedii race 300.

Characterization of a cassiicolin-encoding gene from Corynespora cassiicola, pathogen of rubber tree (Hevea brasiliensis)

Corynespora Leaf Fall (CLF) is a major disease of rubber tree (Hevea brasiliensis) caused by the Ascomycota Corynespora cassiicola. Here we describe the cloning and characterization of a gene encoding cassiicolin (Cas), a glycosylated cystein-rich small secreted protein (SSP) identified as a potential CLF disease effector in rubber tree. Three isolates with contrasted levels of aggressiveness were analyzed comparatively. The cassiicolin gene was detected - and the toxin successfully purified - from the isolates with high and medium aggressiveness (CCP and CCAM3 respectively) but not from the isolate with the lowest aggressiveness (CCAM1), suggesting the existence of a different disease effector in the later. CCP and CCAM3 carried strictly identical cassiicolin genes and produced toxins of identical mass, as evidence by mass spectrometry analysis, thus suggesting conserved post-translational modifications in addition to sequence identity. The differences in aggressiveness between CCP and CCAM3 may be attributed to differences in cassiicolin transcript levels rather than qualitative variations in cassiicolin structure. Cassiicolin may play an important role in the early phase of infection since a peak of cassiicolin transcripts occurred in 1 or 2 days after inoculation (before the occurrence of the first symptoms), in both the tolerant and the susceptible cultivars.

Putative involvement of Thioredoxin h in early response to gravitropic stimulation of poplar stems.

Gravity perception and gravitropic response are essential for plant development. In herbaceous species, it is widely accepted that one of the primary events in gravity perception involves the displacement of amyloplasts within specialized cells. However, the early signaling events leading to stem reorientation are not fully known, especially in woody species in which primary and secondary growth occur. Thirty-six percent of the identified proteins that were differentially expressed after gravistimulation were established as potential Thioredoxin targets. In addition, Thioredoxin h expression was induced following gravistimulation. In situ immunolocalization indicated that Thioredoxin h protein co-localized with the amyloplasts located in the endodermal cells. These investigations suggest the involvement of Thioredoxin h in the first events of signal transduction in inclined poplar stems, leading to reaction wood formation.

Growth and molecular responses to long-distance stimuli in poplars: bending vs flame wounding

Inter-organ communication is essential for plants to coordinate development and acclimate to mechanical environmental fluctuations. The aim of this study was to investigate long-distance signaling in trees. We compared on young poplars the short-term effects of local flame wounding and of local stem bending for two distal responses: (1) stem primary growth and (2) the expression of mechanoresponsive genes in stem apices. We developed a non-contact measurement method based on the analysis of apex images in order to measure the primary growth of poplars. The results showed a phased stem elongation with alternating nocturnal circumnutation phases and diurnal growth arrest phases in Populus tremula × alba clone INRA 717-1B4. We applied real-time polymerase chain reaction (RT-PCR) amplifications in order to evaluate the PtaZFP2, PtaTCH2, PtaTCH4, PtaACS6 and PtaJAZ5 expressions. The flame wounding inhibited primary growth and triggered remote molecular responses. Flame wounding induced significant changes in stem elongation phases, coupled with inhibition of circumnutation. However, the circadian rhythm of phases remained unaltered and the treated plants were always phased with control plants during the days following the stress. For bent plants, the stimulated region of the stem showed an increased PtaJAZ5 expression, suggesting the jasmonates may be involved in local responses to bending. No significant remote responses to bending were observed.