Project description:Nitrogen fixation is a highly energy-demanding process and highly regulated at multiple levels. The two major signals that regulate nitrogen fixation in most diazotrophs are oxygen and ammonia. In order to study the complex regulated mechanism and to highlight the complete nitrogen fixing system in genome level, here we present the transcriptional profiles of the nitrogen fixation genes of P.stutzeri A1501 in different growth conditions with a genome-wide DNA microarray. In this study, the three samples of "P.stutzeri A1501 treated with 0.1mM ammonia and 0.5% Oxygen tension","P.stutzeri A1501 treated with 0.1mM ammonia and 0.5% Oxygen tension-2" and "P.stutzeri A1501 treated with 0.1mM ammonia and 0.5% Oxygen tension-3" were three repeat experiments, while, the other three samples of "P.stutzeri A1501 treated with 20mM ammonia and 0.5% Oxygen tension-1", "P.stutzeri A1501 treated with 20mM ammonia and 0.5% Oxygen tension-2" and "P.stutzeri A1501 treated with 20mM ammonia and 0.5% Oxygen tension-3" were three repeat experiments. The gene expressions under these two growth phases were compared to investigate which genes' expression were effected by different ammonia concentrations. Keywords: nitrogen fixation, nitrogen repression
Project description:A whole genome DNA microarray was used to undertake a global transcriptional analysis of nitrogen fixation and ammonium repression in Pseudomonas stutzeri A1501. The aim of this study was to identify the genes that are up-regulated under nitrogen fixation conditions and rapidly down-regulated as soon as 10 min after ammonia shock. The expression changed genes may be the candidate genes for the ammonia signal transmission or be involved in the nitrogen regulatory mechanism.
Project description:NtrC and Nac have been known to be responsible for responding to nitrogen limitting conditions. In order to elucidate NtrC and Nac regulons in a genome-wide manner, RNA-seq and ChIP-exo experiments were performed for those two transcription factors under 4 different nitrogen sources, ammonia, glutamine, cytidine and cytosine.
Project description:Nitrogenase is the key enzyme involved in nitrogen fixation and uses low potential electrons delivered by ferredoxin or flavodoxin to reduce dinitrogen gas (N2) to produce ammonia and hydrogen. Although the phototrophic alphaproteobacterium Rhodopseudomonas palustris encodes many proteins that can reduce ferredoxin, the electron bifurcating FixABCX complex is the only one shown to support nitrogen fixation. To gain insight into why R. palustris is unable to use these other enzymes to reduce ferredoxin in the absence of FixABCX, we isolated a suppressor of R. palustris DfixC that allowed this strain to grow under nitrogen-fixing conditions. We found two mutations were necessary and sufficient to restore growth under nitrogen-fixing conditions in the absence of a functional FixABCX. One mutation was in the primary ferredoxin involved in nitrogen fixation, fer1, and the other mutation was in rpa0678, a homolog of NAD+-dependent ferredoxin:NADPH oxidoreductase, which carries out flavin-based electron bifurcation to generate reduced Fd. We present evidence that Rpa0678 plays a role in electron transfer to benzoyl-CoA reductase, the key enzyme involved in anaerobic aromatic compound degradation. Together these findings indicate that the electron transfer pathway for anaerobic aromatic compound degradation was re-purposed to support nitrogen fixation in the suppressor strain.
Project description:Here, we report the transcriptome of Anabaena sp. strain 7120, a cyanobacterium that forms specialized nitrogen-fixing cells called heterocysts. Our data suggests that cyanobacteria frequently have more complex transcripts than thought, with large 5' UTRs, numerous antisense transcripts, and multiple transcriptional start sites or processing sites. Four samples of total filament RNA were sequenced with Illumina 40bp reads using directional RNA sequencing (see the Illumina small RNA prep protocol). The samples are 0hr (vegetative cells grown in the presence of ammonia) and 6hr, 12hr, and 21hr cells (after nitrogen step down).
Project description:NtrC and Nac have been known to be responsible for responding to nitrogen limitting conditions. In order to elucidate NtrC and Nac regulons in a genome-wide manner, RNA-seq and ChIP-exo experiments were performed for those two transcription factors under 4 different nitrogen sources, ammonia, glutamine, cytidine and cytosine.
Project description:Giant Axonal Neuropathy (GAN) is a pediatric neurodegenerative disease caused by loss-of-function mutations in the E3 ubiquitin ligase adaptor gigaxonin, which is encoded by the GAN (KLHL16) gene. Gigaxonin regulates the degradation of multiple intermediate filament (IF) proteins, including neurofilaments, peripherin, GFAP, and vimentin, which aggregate in GAN patient cells. Understanding how IFs and their aggregates are processed under stress can reveal new GAN disease mechanisms and potential targets for therapy. Here we tested the hypothesis that hypotonic stress-induced vimentin proteolysis is impaired in GAN. In this mass spectrometry-based proteomics analysis, phosphorylation at Ser-412, located at the junction between the central “rod” domain and C-terminal “tail” domain on vimentin, was identified to be involved in this stress response. Over-expression studies using either phospho-deficient (S412A) or phospho-mimic (S412D) mutants revealed that Ser-412 is important for filament organization, solubility dynamics, and cleavage of vimentin upon hypotonic stress exposure. Collectively, our work reveals that osmotic stress induces calpain- and proteasome-mediated vimentin degradation and IF network breakdown.
Project description:A whole genome DNA microarray was used to undertake a global transcriptional analysis of nitrogen fixation and ammonium repression in Pseudomonas stutzeri A1501. The aim of this study was to identify the genes that are up-regulated under nitrogen fixation conditions and rapidly down-regulated as soon as 10 min after ammonia shock. The expression changed genes may be the candidate genes for the ammonia signal transmission or be involved in the nitrogen regulatory mechanism. First, P. stutzeri A1501 was treated with 0.1 mM ammonia and 0.5% Oxygen tension until the nitrogenase activity was detectable. Then the cells were sudden shifted from the nitrogen fixation conditions to the ammonia repression conditions by addition of 20 mM ammonia for 10min. Subsequently, the bacterium was collected and began the RNA extraction process. Thus, we compare the expression profilings in these two conditions in order to identify the candidate genes.