Project description:In mitosis, most transcription factors detach from chromatin, but some are retained and bookmark genomic sites. Mitotic bookmarking has been implicated in lineage inheritance, pluripotency and reprogramming. However, the biological significance of this mechanism in vivo remains unclear. Here, we address mitotic retention of the hemogenic factors GATA2, GFI1B and FOS during haematopoietic specification. We show that GATA2 remains bound to chromatin throughout mitosis, in contrast to GFI1B and FOS, via C-terminal zinc finger-mediated DNA binding. GATA2 bookmarks a subset of its interphase targets that are co-enriched for RUNX1 and other regulators of definitive haematopoiesis. Remarkably, homozygous mice harbouring the cyclin B1 mitosis degradation domain upstream Gata2 partially phenocopy knockout mice. Degradation of GATA2 at mitotic exit abolishes definitive haematopoiesis at aorta-gonad-mesonephros, placenta and foetal liver, but does not impair yolk sac haematopoiesis. Our findings implicate GATA2-mediated mitotic bookmarking as critical for definitive haematopoiesis and highlight a dependency on bookmarkers for lineage commitment.
Project description:modENCODE_submission_3224 This submission comes from a modENCODE project of Michael Snyder. For full list of modENCODE projects, see http://www.genome.gov/26524648 Project Goal: We are identifying the DNA binding sites for 300 transcription factors in C. elegans. Each transcription factor gene is tagged with the same GFP fusion protein, permitting validation of the gene's correct spatio-temporal expression pattern in transgenic animals. Chromatin immunoprecipitation on each strain is peformed using an anti-GFP antibody, and any bound DNA is deep-sequenced using Solexa GA2 technology. For data usage terms and conditions, please refer to http://www.genome.gov/27528022 and http://www.genome.gov/Pages/Research/ENCODE/ENCODEDataReleasePolicyFinal2008.pdf EXPERIMENT TYPE: CHIP-seq. BIOLOGICAL SOURCE: Strain: OP304(official name : OP304 genotype : unc119(ed3);wgIs304(fos-1::TY1 EGFP FLAG;unc119) outcross : 3 mutagen : Bombard tags : GFP::3xFlag description : This strain's transgene was constructed by Mihail Sarov at the Max Planck Institute for Cell Biology in Tubiginen using Tony Hyman's recombineering pipeline. The resulting plasmid was used for biolistic transformation of an unc-119(ed3) strain. The FOS-1::EGFP fusion protein is expressed in the correct fos-1 spatio-temporal expression pattern. This strain was used for ChIP-seq experiments to map the in vivo binding sites for the FOS-1 transcription factor. made_by : R Waterston ); Developmental Stage: L2; Genotype: unc119(ed3);wgIs304(fos-1::TY1 EGFP FLAG;unc119); Sex: Hermaphrodite; EXPERIMENTAL FACTORS: Developmental Stage L2; Target gene fos-1; Strain OP304(official name : OP304 genotype : unc119(ed3);wgIs304(fos-1::TY1 EGFP FLAG;unc119) outcross : 3 mutagen : Bombard tags : GFP::3xFlag description : This strain's transgene was constructed by Mihail Sarov at the Max Planck Institute for Cell Biology in Tubiginen using Tony Hyman's recombineering pipeline. The resulting plasmid was used for biolistic transformation of an unc-119(ed3) strain. The FOS-1::EGFP fusion protein is expressed in the correct fos-1 spatio-temporal expression pattern. This strain was used for ChIP-seq experiments to map the in vivo binding sites for the FOS-1 transcription factor. made_by : R Waterston ); temp (temperature) 20 degree celsius
Project description:modENCODE_submission_3588 This submission comes from a modENCODE project of Michael Snyder. For full list of modENCODE projects, see http://www.genome.gov/26524648 Project Goal: We are identifying the DNA binding sites for 300 transcription factors in C. elegans. Each transcription factor gene is tagged with the same GFP fusion protein, permitting validation of the gene's correct spatio-temporal expression pattern in transgenic animals. Chromatin immunoprecipitation on each strain is peformed using an anti-GFP antibody, and any bound DNA is deep-sequenced using Solexa GA2 technology. For data usage terms and conditions, please refer to http://www.genome.gov/27528022 and http://www.genome.gov/Pages/Research/ENCODE/ENCODEDataReleasePolicyFinal2008.pdf EXPERIMENT TYPE: CHIP-seq. BIOLOGICAL SOURCE: Strain: OP304(official name : OP304 genotype : unc119(ed3);wgIs304(fos-1::TY1 EGFP FLAG;unc119) outcross : 3 mutagen : Bombard tags : GFP::3xFlag description : This strain's transgene was constructed by Mihail Sarov at the Max Planck Institute for Cell Biology in Tubiginen using Tony Hyman's recombineering pipeline. The resulting plasmid was used for biolistic transformation of an unc-119(ed3) strain. The FOS-1::EGFP fusion protein is expressed in the correct fos-1 spatio-temporal expression pattern. This strain was used for ChIP-seq experiments to map the in vivo binding sites for the FOS-1 transcription factor. made_by : R Waterston ); Developmental Stage: L4; Genotype: unc119(ed3);wgIs304(fos-1::TY1 EGFP FLAG;unc119); Sex: Hermaphrodite; EXPERIMENTAL FACTORS: Developmental Stage L4; Target gene fos-1; Strain OP304(official name : OP304 genotype : unc119(ed3);wgIs304(fos-1::TY1 EGFP FLAG;unc119) outcross : 3 mutagen : Bombard tags : GFP::3xFlag description : This strain's transgene was constructed by Mihail Sarov at the Max Planck Institute for Cell Biology in Tubiginen using Tony Hyman's recombineering pipeline. The resulting plasmid was used for biolistic transformation of an unc-119(ed3) strain. The FOS-1::EGFP fusion protein is expressed in the correct fos-1 spatio-temporal expression pattern. This strain was used for ChIP-seq experiments to map the in vivo binding sites for the FOS-1 transcription factor. made_by : R Waterston ); temp (temperature) 20 degree celsius
Project description:modENCODE_submission_3587 This submission comes from a modENCODE project of Michael Snyder. For full list of modENCODE projects, see http://www.genome.gov/26524648 Project Goal: We are identifying the DNA binding sites for 300 transcription factors in C. elegans. Each transcription factor gene is tagged with the same GFP fusion protein, permitting validation of the gene's correct spatio-temporal expression pattern in transgenic animals. Chromatin immunoprecipitation on each strain is peformed using an anti-GFP antibody, and any bound DNA is deep-sequenced using Solexa GA2 technology. For data usage terms and conditions, please refer to http://www.genome.gov/27528022 and http://www.genome.gov/Pages/Research/ENCODE/ENCODEDataReleasePolicyFinal2008.pdf EXPERIMENT TYPE: CHIP-seq. BIOLOGICAL SOURCE: Strain: OP304(official name : OP304 genotype : unc119(ed3);wgIs304(fos-1::TY1 EGFP FLAG;unc119) outcross : 3 mutagen : Bombard tags : GFP::3xFlag description : This strain's transgene was constructed by Mihail Sarov at the Max Planck Institute for Cell Biology in Tubiginen using Tony Hyman's recombineering pipeline. The resulting plasmid was used for biolistic transformation of an unc-119(ed3) strain. The FOS-1::EGFP fusion protein is expressed in the correct fos-1 spatio-temporal expression pattern. This strain was used for ChIP-seq experiments to map the in vivo binding sites for the FOS-1 transcription factor. made_by : R Waterston ); Developmental Stage: L3; Genotype: unc119(ed3);wgIs304(fos-1::TY1 EGFP FLAG;unc119); Sex: Hermaphrodite; EXPERIMENTAL FACTORS: Developmental Stage L3; Target gene fos-1; Strain OP304(official name : OP304 genotype : unc119(ed3);wgIs304(fos-1::TY1 EGFP FLAG;unc119) outcross : 3 mutagen : Bombard tags : GFP::3xFlag description : This strain's transgene was constructed by Mihail Sarov at the Max Planck Institute for Cell Biology in Tubiginen using Tony Hyman's recombineering pipeline. The resulting plasmid was used for biolistic transformation of an unc-119(ed3) strain. The FOS-1::EGFP fusion protein is expressed in the correct fos-1 spatio-temporal expression pattern. This strain was used for ChIP-seq experiments to map the in vivo binding sites for the FOS-1 transcription factor. made_by : R Waterston ); temp (temperature) 20 degree celsius
Project description:In mitosis, most transcription factors detach from chromatin, but some are retained and bookmark genomic sites. Mitotic bookmarking has been implicated in lineage inheritance, pluripotency and reprogramming. However, the biological significance of this mechanism in vivo remains unclear. Here, we address mitotic retention of the hemogenic factors GATA2, GFI1B and FOS during haematopoietic specification. We show that GATA2 remains bound to chromatin throughout mitosis, in contrast to GFI1B and FOS, via C-terminal zinc finger-mediated DNA binding. GATA2 bookmarks a subset of its interphase targets that are co-enriched for RUNX1 and other regulators of definitive haematopoiesis. Remarkably, homozygous mice harbouring the cyclin B1 mitosis degradation domain upstream Gata2 partially phenocopy knockout mice. Degradation of GATA2 at mitotic exit abolishes definitive haematopoiesis at aorta-gonad-mesonephros, placenta and foetal liver, but does not impair yolk sac haematopoiesis. Our findings implicate GATA2-mediated mitotic bookmarking as critical for definitive haematopoiesis and highlight a dependency on bookmarkers for lineage commitment.
Project description:More than half of the protein-coding genes in bacteria are organized in polycistronic operons composed of two or more genes. Whether the operon organization maintains the stoichiometric expression of the genes within an operon remain under debate. In this study, we performed a label-free data-independent acquisition hyper reaction monitoring mass-spectrometry (HRM-MS) experiment to quantify the Escherichia coli proteome in exponential phase and quantified 93.6% of the cytosolic proteins, covering 67.9% and 56.0% of the translating polycistronic operons in BW25113 and MG1655 strains, respectively. We found the shorter operons tend to be more tightly controlled for stoichiometry than longer operons, and those operons for metabolic pathways are less controlled for stoichiometry compared with operons for protein complexes, illustrating the multifaceted nature of the operon-wise regulation: the operon-wise unified transcriptional level and gene-specific translational level. This multi-level regulation benefits the host by optimizing the efficiency of the productivity of metabolic pathways and maintenance of different types of protein complexes.
Project description:modENCODE_submission_3387 This submission comes from a modENCODE project of Michael Snyder. For full list of modENCODE projects, see http://www.genome.gov/26524648 Project Goal: We are identifying the DNA binding sites for 300 transcription factors in C. elegans. Each transcription factor gene is tagged with the same GFP fusion protein, permitting validation of the gene's correct spatio-temporal expression pattern in transgenic animals. Chromatin immunoprecipitation on each strain is peformed using an anti-GFP antibody, and any bound DNA is deep-sequenced using Solexa GA2 technology. For data usage terms and conditions, please refer to http://www.genome.gov/27528022 and http://www.genome.gov/Pages/Research/ENCODE/ENCODEDataReleasePolicyFinal2008.pdf EXPERIMENT TYPE: CHIP-seq. BIOLOGICAL SOURCE: Strain: OP304(official name : OP304 genotype : unc119(ed3);wgIs304(fos-1::TY1 EGFP FLAG;unc119) outcross : 3 mutagen : Bombard tags : GFP::3xFlag description : This strain's transgene was constructed by Mihail Sarov at the Max Planck Institute for Cell Biology in Tubiginen using Tony Hyman's recombineering pipeline. The resulting plasmid was used for biolistic transformation of an unc-119(ed3) strain. The FOS-1::EGFP fusion protein is expressed in the correct fos-1 spatio-temporal expression pattern. This strain was used for ChIP-seq experiments to map the in vivo binding sites for the FOS-1 transcription factor. made_by : R Waterston ); Developmental Stage: fed L1; Genotype: unc119(ed3);wgIs304(fos-1::TY1 EGFP FLAG;unc119); Sex: Hermaphrodite; EXPERIMENTAL FACTORS: Developmental Stage fed L1; Target gene fos-1; Strain OP304(official name : OP304 genotype : unc119(ed3);wgIs304(fos-1::TY1 EGFP FLAG;unc119) outcross : 3 mutagen : Bombard tags : GFP::3xFlag description : This strain's transgene was constructed by Mihail Sarov at the Max Planck Institute for Cell Biology in Tubiginen using Tony Hyman's recombineering pipeline. The resulting plasmid was used for biolistic transformation of an unc-119(ed3) strain. The FOS-1::EGFP fusion protein is expressed in the correct fos-1 spatio-temporal expression pattern. This strain was used for ChIP-seq experiments to map the in vivo binding sites for the FOS-1 transcription factor. made_by : R Waterston ); temp (temperature) 20 degree celsius
Project description:The bifidogenic effect of the prebiotic inulin and its hydrolysed form (fructoligosaccharide, FOS) is well established since they promote the growth of specific beneficial (probiotic) gut bacteria such as bifidobacteria. Previous studies of the opportunistic nosocomial pathogen Pseudomonas aeruginosa PAO1 have shown that inulin and FOS reduce growth, biofilm formation, through decrease of its ability to motility and exotoxin secretion. However, the transcriptional basis for these phenotypic alterations remains unclear. To address this question we conducted RNA-sequence analysis. In most cases changes in the transcript level induced by inulin and FOS were similar. However, a set of transcripts were increased in expression response to inulin, but reduced in the presence of FOS. In the presence of inulin or FOS 260 and 217 transcript levels, respectively, were altered as compared to the control to which no polysaccharide was added. Importantly, changes in transcript levels of 57 and 83 genes were found to be specific for either inulin or FOS, respectively, indicating that both compounds trigger different changes. Gene pathway analysis of different exposure genes (DEG) revealed a specific FOS-mediated reduction in transcript levels of genes that participate in several canonical pathways involved in metabolism and growth, motility, biofilm formation, β-lactam resistance and in the modulation of type III and VI secretion system which were supported by real time quantitative PCR measurements. These results may provide solid information to suggest that FOS selectively modulates the P. aeruginosa PAO1 pathogenecity through distinct signaling pathways.