Expression and validation of PvPGIP genes for resistance to white mold (Sclerotinia sclerotiorum) in common beans (Phaseolus vulgaris L.).

The interaction between polygalacturonase-inhibiting proteins (PGIPs), produced by plants, and endopolygalacturonases (PGs), produced by fungi, limits the destructive potential of PGs and can trigger plant defense responses. This study aimed to i) investigate variation in the expression of different common bean (Phaseolus vulgaris L.) genotypes and its relationship with resistance to white mold (Sclerotinia sclerotiorum); ii) determine the expression levels of PvPGIP genes at different time points after inoculation with white mold; and iii) investigate differences in PvPGIP gene expression between two white mold isolates with different levels of aggressiveness. Four bean lines were analyzed, including two lines from a recurrent selection for white mold (50/5 and 84/6), one resistant line that was not adapted to Brazilian conditions (Cornell 605), and one susceptible line (Corujinha). Gene expression was investigated at 0, 1, 2, 3, and 5 days after inoculation. The isolate UFLA 03 caused no significant difference in the relative expression of any gene examined, and was inefficient in discriminating among the genotypes. For the isolate UFLA 116, all of the genes were differentially expressed, as they were associated with resistance to white mold, and the expressions increased until the third day after inoculation. The 50/5 line was not significantly different from the Corujinha line for all of the genes analyzed. However, this line had a resistance level that was similar to that of Cornell 605, according to the straw test. Therefore, the incorporation of PvPGIP genes can increase the resistance of lines derived from recurrent selection.

New Insights on Coffea miRNAs: Features and Evolutionary Conservation.

Small RNAs influence the gene expression at the post-transcriptional level by guiding messenger RNA (mRNA) cleavage, translational repression, and chromatin modifications. In addition to model plants, the microRNAs (miRNAs) have been identified in different crop species. In this work, we developed a specific pipeline to search for coffee miRNA homologs on expressed sequence tags (ESTs) and genome survey sequences (GSS) databases. As a result, 36 microRNAs were identified and a total of 616 and 362 potential targets for Coffea arabica and Coffea canephora, respectively. The evolutionary analyses of these molecules were performed by comparing the primary and secondary structures of precursors and mature miRNAs with their orthologs. Moreover, using a stem-loop RT-PCR assay, we evaluated the accumulation of mature miRNAs in genomes with different ploidy levels, detecting an increase in the miRNAs accumulation according to the ploidy raising. Finally, a 5' RACE (Rapid Amplification of cDNA Ends) assay was performed to verify the regulation of auxin responsive factor 8 (ARF8) by MIR167 in coffee plants. The great variety of target genes indicates the functional plasticity of these molecules and reinforces the importance of understanding the RNAi-dependent regulatory mechanisms. Our results expand the study of miRNAs and their target genes in this crop, providing new challenges to understand the biology of these species.

A proposed model for the flowering signaling pathway of sugarcane under photoperiodic control.

Molecular analysis of floral induction in Arabidopsis has identified several flowering time genes related to 4 response networks defined by the autonomous, gibberellin, photoperiod, and vernalization pathways. Although grass flowering processes include ancestral functions shared by both mono- and dicots, they have developed their own mechanisms to transmit floral induction signals. Despite its high production capacity and its important role in biofuel production, almost no information is available about the flowering process in sugarcane. We searched the Sugarcane Expressed Sequence Tags database to look for elements of the flowering signaling pathway under photoperiodic control. Sequences showing significant similarity to flowering time genes of other species were clustered, annotated, and analyzed for conserved domains. Multiple alignments comparing the sequences found in the sugarcane database and those from other species were performed and their phylogenetic relationship assessed using the MEGA 4.0 software. Electronic Northerns were run with Cluster and TreeView programs, allowing us to identify putative members of the photoperiod-controlled flowering pathway of sugarcane.

In silico characterization of putative members of the coffee (Coffea arabica) ethylene signaling pathway.

The plant hormone ethylene is involved in several developmental and physiological processes in plants, including senescence, fruit ripening and organ abscission, as well as in biotic and abiotic stress responses. Initiation of these processes involves complex regulation of both ethylene biosynthesis and the ability of cells to perceive the hormone and respond in an appropriate manner, a process which is regulated both spatially and temporally. Ethylene is a gaseous hormone whose sensitivity is a key factor to limiting its response in target cells. We made a search of the Coffee Expressed Sequence Tag (CAFEST) database for expressed sequence tags related to known elements of the ethylene signaling pathway. Sequences showing a reliable similarity were clusterized, annotated and analyzed for conserved domains. Multiple alignments comprising the sequences that we found and sequences of ethylene signaling elements from other species were made, and their phylogeny was assessed by phylogenetic trees constructed with the MEGA4 software. The expression profile was assessed by in silico Northern blot analysis performed using the Cluster and TreeView programs. The CAFEST database was found to have a large number of sequences related to previously described ethylene signaling pathway elements, allowing identification of putative members from almost every step of this pathway. The phylogenetic trees demonstrated high similarity between the sequences found in the CAFEST and those from other species, and the electronic Northern blot analysis detected their expression in various tissues, development stages and stress conditions.