Project description:Expression of virulence genes in pathogenic E. coli is controlled in part by the transcription silencer H-NS and its paralogs (e.g., StpA), which sequester DNA in multi-kb nucleoprotein filaments to inhibit transcription initiation, elongation, or both. Some activators counter-silence initiation by displacing H-NS from promoters. How H-NS inhibition of elongation is overcome is not understood. In uropathogenic E. coli (UPEC), elongation regulator RfaH aids expression of some H-NS-silenced pathogenicity operons (e.g., hlyCABD encoding hemolysin). RfaH associates with elongation complexes (ECs) via direct contacts to a transiently exposed, nontemplate DNA-strand sequence called ops (operon polarity suppressor). RfaH–ops interactions establish long-lived RfaH–EC contacts that allow RfaH to recruit ribosomes to the nascent mRNA and to suppress transcriptional pausing and termination. Using ChIP-seq, we mapped the genome-scale distributions of RfaH, H-NS, StpA, RNA polymerase (RNAP), and σ70 in the UPEC strain CFT073. We identify 8 RfaH-activated operons, all of which were bound by H-NS and StpA. Four are new additions to the RfaH regulon. Deletion of RfaH caused premature termination whereas deletion of H-NS and StpA allowed elongation without RfaH. Thus, RfaH is an elongation counter-silencer of H-NS. Consistent with elongation counter-silencing, deletion of StpA alone decreased the effect of RfaH. StpA increases DNA bridging, which inhibits transcript elongation via topological constraints on RNAP. Residual RfaH effect when both H-NS and StpA were deleted was attributable to targeting of RfaH-regulated operons by a minor H-NS paralog, Hfp. These operons have evolved higher levels of H-NS–binding features, explaining minor-paralog targeting.
Project description:The global regulator H-NS represses transcription in gram negative bacteria. Sfh is a homologue of H-NS and is encoded by plasmid pSfR27. Sfh provides a 'stealth' function that allows pSfR27 to be transmitted to a new host without disrupting the competitive fitness of the new host We used ChIP-on-chip to profile Sfh (3xFLAG-tagged) and H-NS binding sites in Salmonella Typhimurium strain SL1344 and found that Sfh provides its 'stealth' function by targeting a sub-set of H-NS bound genes that display reduced levels of H-NS occupancy with the SL1344 chromosome upon acquisition of plasmid pSfR27
Project description:Viscum album is known for its special mode of cellular respiration. It lacks the mitochondrial NADH dehydrogenase complex (complex I of the respiratory chain) and has restricted capacities to generate mitochondrial adenosine triphosphate (ATP). We here present an investigation of the V. album energy metabolism taking place in mitochondria. Mitochondria were purified from young V. album leaves and membrane bound protein complexes characterized by Blue native polyacrylamide gel electrophoresis as well as by the complexome profiling approach. Proteins were systematically identified by label-free quantitative shotgun proteomics.
Project description:Viscum album is known for its special mode of cellular respiration. It lacks the mitochondrial NADH dehydrogenase complex (complex I of the respiratory chain) and has restricted capacities to generate mitochondrial adenosine triphosphate (ATP). We here present an investigation of the V. album energy metabolism taking place in the chloroplasts. Thylakoids were purified from young V. album leaves and membrane bound protein complexes characterized by Blue native polyacrylamide gel electrophoresis as well as by the complexome profiling approach. Proteins were systematically identified by label-free quantitative shotgun proteomics.
Project description:Transcriptional profiling of squamous cell carcinoma of oral tongue, comparing p53 NS+ and p53 NS- tumors. Goal was to determine differentially expressed genes between them based on global gene expression.