Project description:Detecting chemical signals is important for identifying food sources and avoiding harmful agents. Like many animals, C. elegans use olfaction to chemotax towards their main food source, bacteria. However, little is known about the bacterial compounds governing C. elegans attraction to bacteria and the physiological importance of these compounds to bacteria. Here, we address these questions by investigating the function of a small RNA, P11, in the pathogen, Pseudomonas aeruginosa, that was previously shown to mediate learned pathogen avoidance. We discovered that this RNA also affects the attraction of untrained C. elegans to P. aeruginosa and does so by controlling production of ammonia, a volatile odorant produced during nitrogen assimilation. We describe the complex regulation of P. aeruginosa nitrogen assimilation, which is mediated by a partner-switching mechanism involving environmental nitrates, sensor proteins, and P11. In addition to mediating C. elegans attraction, we demonstrate that nitrogen assimilation mutants perturb bacterial fitness and pathogenesis during C. elegans infection by P. aeruginosa. These studies define ammonia as a major mediator of trans-kingdom signaling, implicate nitrogen assimilation as important for both bacteria and host organisms, and highlight how a bacterial metabolic pathway can either benefit or harm a host in different contexts.
Project description:Assimilation of nitrogen is an essential process in bacteria. The nitrogen regulation stress response is an adaptive mechanism used by nitrogen-starved Escherichia coli to scavenge for alternative nitrogen sources and requires the global transcriptional regulator NtrC.
Project description:Transcriptional profiling of Haloferax mediterranei in three culture media with different nitrogen sources: a) cells were grown stationary and exponentially on ammonium, b) cells were grown exponentially on nitrate, and c) cells were shifted to nitrogen starvation conditions. The main differences in the transcriptional profiles have been identified between the cultures with ammonium as nitrogen source and the cultures with nitrate or nitrogen starvation, supporting previous results which indicate the absence of ammonium as the factor causing the expression of genes involved in nitrate assimilation pathway.
Project description:Small RNAs have been studied in detail in Bacteria and Eukarya domain, but in the case of Archaea domain the knowledge is scarce and the physiological function of the majority is still uncertain. To extend the knowledge of sRNAs in Archaea domain and its possible role in the regulation of the nitrogen assimilation metabolism in haloarchaea, Haloferax mediterranei has been used as a model microorganism. Bioinformatic approach has allowed to predict 295 putative sRNAs genes in the genome of H. mediterranei, 88 of which have been verified by means of RNA-seq. The secondary structure of putative sRNAs and its possible targets have been identified. Curiously, some of them present as possible targets genes related to the nitrogen assimilation, as glutamate dehydrogenase or regulatory nitrogen protein PII. Analysis of RNA-seq data has also revealed differences in the expression pattern of 16 sRNAs according to the nitrogen source. Consequently, RNomic and the bioinformatic approaches used in this work have allowed the identification of new sRNAs in Hfx. mediterranei, some of which show different expression pattern depending on the nitrogen source. It suggests that these sRNAs could be involved in the regulation of nitrogen assimilation, being able to constitute important gene regulatory network.
Project description:Glutamine synthetase (GS), a key enzyme in biological nitrogen assimilation, is regulated in multiple ways in response to varying nitrogen sources and levels. Here we show a small regulatory RNA, NsiR4 (nitrogen stress induced RNA 4), which plays an important role in the regulation of GS in cyanobacteria. NsiR4 expression in the unicellular Synechocystis sp. PCC 6803 and in the filamentous, nitrogen-fixing Anabaena sp. PCC 7120 is stimulated through nitrogen-limitation via NtcA, the global transcriptional regulator of genes involved in nitrogen metabolism. NsiR4 is widely conserved throughout the cyanobacterial phylum, suggesting a conserved function. In silico target prediction, transcriptome profiling upon pulse overexpression and site-directed mutagenesis experiments using a heterologous reporter system showed that NsiR4 interacts with the 5’UTR of gifA mRNA, which encodes glutamine synthetase inactivating factor IF7. In Synechocystis, we observed an inverse relationship between the levels of NsiR4 and the accumulation of IF7 in vivo. This NsiR4-dependent modulation of gifA (IF7) mRNA accumulation influenced the glutamine pool and thus NH4+ assimilation via glutamine synthetase. As a second target, we identified ssr1528, a hitherto uncharacterized nitrogen-regulated gene. Competition experiments between wild type and an NsiR4 knock-out mutant showed that the lack of NsiR4 led to decreased acclimation capabilities of Synechocystis towards oscillating nitrogen levels. These results suggest a role for NsiR4 in the regulation of nitrogen metabolism in cyanobacteria, especially for the adaptation to rapid changes in available nitrogen sources and concentrations. NsiR4 is the first identified bacterial sRNA regulating the primary assimilation of a macronutrient.
Project description:In this approach, total transcript of the Mycobacterium smegmatis wild type SMR5 was compared under nitrogen surplus and starvation. This resulted in the change of transcript patterns of more than 500 genes due to nitrogen or general starvation response. Genes involved in uptake and assimilation of ammonium and alternative nitrogen sources showed enhanced transcript levels, genes involved in cell growth, protein biosynthesis etc showed reduced transcript levels. Two biological replicates were analyzed.
Project description:Transcriptional profiling of Haloferax mediterranei in three culture media with different nitrogen sources: a) cells were grown stationary and exponentially on ammonium, b) cells were grown exponentially on nitrate, and c) cells were shifted to nitrogen starvation conditions. The main differences in the transcriptional profiles have been identified between the cultures with ammonium as nitrogen source and the cultures with nitrate or nitrogen starvation, supporting previous results which indicate the absence of ammonium as the factor causing the expression of genes involved in nitrate assimilation pathway. Four-conditions experiment with four biological replicates, combined in a loop design.
Project description:Bacteria are major drivers of organic matter decomposition and play crucial roles in global nutrient cycling. Although the degradation of dead fungal biomass (necromass) is increasingly recognized as an important contributor to soil carbon (C) and nitrogen (N) cycling, the genes and metabolic pathways involved in necromass degradation are under characterized. In particular, how bacteria degrade necromass containing different quantities of melanin, which largely control rates of necromass decomposition in situ, is largely unknown. To address this gap, we conducted a multi-timepoint transcriptomic analysis using three Gram-negative, bacterial species grown on low or high melanin necromass of Hyaloscypha bicolor. The bacterial species, Cellvibrio japonicus, Chitinophaga pinensis, and Serratia marcescens, belong to genera known to degrade necromass in situ. We found that while bacterial growth was consistently higher on low than high melanin necromass, the CAZyme-encoding gene expression response of the three species was similar between the two necromass types. Interestingly, this trend was not shared for genes encoding nitrogen utilization, which varied in C. pinensis and S. marcescens during growth on high versus low melanin necromass. Additionally, this study tested the metabolic capabilities of these bacterial species to grow on a diversity of C and N sources and found that the three bacteria have substantially different abilities to utilize carbon and nitrogen compounds. Collectively, our data suggests that as necromass changes chemically over the course of degradation, certain bacterial species are favored based on their differential metabolic capacities.