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Catharanthus roseus produces a diverse range of specialized metabolites of the monoterpenoid indole alkaloid (MIA) class in a heavily branched pathway. Recent great progress in identification of MIA biosynthesis genes revealed that the different pathway branch genes are expressed in a highly cell type- and organ-specific and stress-dependent manner. This implies a complex control by specific transcription factors (TFs), only partly revealed today. We generated and mined a comprehensive compendium of publicly available C. roseus transcriptome data for MIA pathway branch-specific TFs. Functional analysis was performed through extensive comparative gene expression analysis and profiling of over 40 MIA metabolites in the C. roseus flower petal expression system. We identified additional members of the known BIS and ORCA regulators. Further detailed study of the ORCA TFs suggests subfunctionalization of ORCA paralogs in terms of target gene-specific regulation and synergistic activity with the central jasmonate response regulator MYC2. Moreover, we identified specific amino acid residues within the ORCA DNA-binding domains that contribute to the differential regulation of some MIA pathway branches. Our results advance our understanding of TF paralog specificity for which, despite the common occurrence of closely related paralogs in many species, comparative studies are scarce.

Abstract Background This study of the oropharyngeal microbiome complements the previously published AZIthromycin in Severe ASThma (AZISAST) clinical trial, where the use of azithromycin was assessed in subjects with exacerbation-prone severe asthma. Here, we determined the composition of the oropharyngeal microbial community by means of deep sequencing of the amplified 16S rRNA gene in oropharyngeal swabs from patients with exacerbation-prone severe asthma, at baseline and during and after 6 months treatment with azithromycin or placebo. Results A total of 1429 OTUs were observed, of which only 59 were represented by more than 0.02% of the reads. Firmicutes, Bacteroidetes, Fusobacteria, Proteobacteria and Actinobacteria were the most abundant phyla and Streptococcus and Prevotella were the most abundant genera in all the samples. Thirteen species only accounted for two thirds of the reads and two species only, i.e. Prevotella melaninogenica and Streptococcus mitis/pneumoniae, accounted for one fourth of the reads. We found that the overall composition of the oropharyngeal microbiome in patients with severe asthma is comparable to that of the healthy population, confirming the results of previous studies. Long term treatment (6 months) with azithromycin increased the species Streptococcus salivarius approximately 5-fold and decreased the species Leptotrichia wadei approximately 5-fold. This was confirmed by Boruta feature selection, which also indicated a significant decrease of L. buccalis/L. hofstadtii and of Fusobacterium nucleatum. Four of the 8 treated patients regained their initial microbial composition within one month after cessation of treatment. Conclusions Despite large diversity of the oropharyngeal microbiome, only a few species predominate. We confirm the absence of significant differences between the oropharyngeal microbiomes of people with and without severe asthma. Possibly, long term azithromycin treatment may have long term effects on the composition of the oropharygeal microbiome in half of the patients.

Sugarcane interacts with particular types of beneficial nitrogen-fixing bacteria that provide fixed-nitrogen and plant growth hormones to host plants, promoting an increase in plant biomass. Other benefits, as enhanced tolerance to abiotic stresses have been reported to some diazotrophs. Here we aim to study the effects of the association between the diazotroph Gluconacetobacter diazotrophicus PAL5 and sugarcane cv. SP70-1143 during water depletion by characterizing differential transcriptome profiles of sugarcane. RNA-seq libraries were generated from roots and shoots of sugarcane plants free of endophytes that were inoculated with G. diazotrophicus and subjected to water depletion for 3 days. A sugarcane reference transcriptome was constructed and used for the identification of differentially expressed transcripts. The differential profile of non-inoculated SP70-1143 suggests that it responds to water deficit stress by the activation of drought-responsive markers and hormone pathways, as ABA and Ethylene. qRT-PCR revealed that root samples had higher levels of G. diazotrophicus 3 days after water deficit, compared to roots of inoculated plants watered normally. With prolonged drought only inoculated plants survived, indicating that SP70-1143 plants colonized with G. diazotrophicus become more tolerant to drought stress than non-inoculated plants. Strengthening this hypothesis, several gene expression responses to drought were inactivated or regulated in an opposite manner, especially in roots, when plants were colonized by the bacteria. The data suggests that colonized roots would not be suffering from stress in the same way as non-inoculated plants. On the other hand, shoots specifically activate ABA-dependent signaling genes, which could act as key elements in the drought resistance conferred by G. diazotrophicus to SP70-1143. This work reports for the first time the involvement of G. diazotrophicus in the promotion of drought-tolerance to sugarcane cv. SP70-1143, and it describes the initial molecular events that may trigger the increased drought tolerance in the host plant.