Project description:Pseudomonas syringae, the leading cause of crop diseases world-wide, uses type III secretion system (T3SS) to invade host plants. Our previous studies demonstrate that two-component system RhpRS enable P. syringae to coordinate T3SS gene expression, which is dependent on the phosphorylation state of RhpR in different environmental conditions. Homologs of RhpRS are distributed in a wide range of bacterial species, indicating a general regulatory mechanism. However, it remains elusive that how RhpRS relies on external signals and phosphorylation state to exercise its regulatory functions. To this end, we performed ChIP-seq assays to identify specific binding sites of RhpR and RhpRD70A in either King’s B (KB, T3SS-inhibiting medium) or minimal medium (MM, T3SS-inducing medium). Through data analysis, we identified 125 KB-dependent binding sites and 188 phosphorylation-dependent binding sites of RhpR. In KB, RhpR directly positively regulated cytochrome c550 production (via ccmA) and alcohol dehydrogenase activity (via adhB), but negatively regulated anthranilate synthase activity (via trpG) and protease activity (via hemB). Besides, phosphorylated RhpR (RhpR-P) directly negatively regulated T3SS (via hrpR and hopR1), swimming motility (via flhA), c-di-GMP level (via PSPPH_2590) and biofilm formation (via algD), while positively regulated twitching motility (via fimA) and lipopolysaccharide production (via PSPPH_2653). Our RNA-seq analyses newly identified 474 and 840 genes that regulated by RhpR in KB and MM respectively. Collectively, the present study revealed that rich nutrient condition allows RhpR to directly regulate the multiple metabolic pathways of P. syringae, while phosphorylation enables RhpR to specifically control virulence and cell envelope. RhpRS governs both virulence and multiple metabolic pathways by tuning its phosphorylation and sensing environmental signals in KB, respectively.
Project description:PSPTO_2222 and PSPTO_2222-2223 deletions mutants were grown in MM or KB, and their expression profiles were analyzed. 2222MM: PSPTO_2222 mutants cultured in MM. 3/2 MM: PSPTO_2222-2223 deletion mutants cultured in MM. 3-2 kb: PSPTO_2222-2223 deletion mutants cultured in KB. P2KB: PSPTO_2222 mutants cultured in KB.
Project description:Estrogen Receptor alpha (ERα) is a key driver of most breast cancers, and it is the target of endocrine therapies used in the clinic to treat women with ERα positive (ER+) breast cancer. The two methods ChIP-seq (chromatin immunoprecipitation coupled with deep sequencing) and RIME (Rapid Immunoprecipitation of Endogenous Proteins) have greatly improved our understanding of ERα function during breast cancer progression and in response to anti-estrogens. A critical component of both ChIP-seq and RIME protocols is the antibody that is used to pull down the bait protein. To date, most of the ChIP-seq and RIME experiments for the study of ERα have been performed using the sc-543 antibody from Santa Cruz Biotechnology. However, this antibody has been discontinued, thereby severely impacting the study of ERα in normal physiology as well as diseases such as breast cancer and ovarian cancer. Here, we compare the sc-543 antibody with other commercially available antibodies, and we show that 06-935 (EMD Millipore) and ab3575 (Abcam) antibodies can successfully replace the sc-543 antibody for ChIP-seq and RIME experiments.
Project description:We used ChIP-seq to map the binding of of C-terminally FLAG3-tagged PhoB to the Escherichia coli K-12 genome during growth in MOPS minimal media with low phosphate or high phosphate (0.2 mM or 1.32 mM, respectively). As a control, we performed ChIP-seq in an untagged strain. We also used ChIP-seq to map Sigma 70 (associates with initiating RNA polymerase) binding across the E. coli K-12 genome in wild-type, ΔphoB, Δhns, and Δhns ΔphoB strains grown in low phosphate medium to determine whether PhoB modulates recruitment of RNA polymerase to promoters and whether this is modulated by H-NS.
Project description:Most patients with multiple myeloma (MM) will eventually relapse and current treatments have limited effect. Herein, we demonstrate that succinate dehydrogenase subunitA (SDHA) was low expressed in MM patients, and patients with SDHA relatively high expression had long overall survival and progression-free survival. Furthermore, SDHA high expression inhibited proliferation and invasion in multiple myeloma (MM) cell lines and enhanced the anti-tumor and synergistic effect of chemotherapeutics. To investgate regulation of SDHA on epigenetic level, we conducted ChIP-seq experiments to compare the H3K27ac signal between H929 cells treated with DMSO and H929 cells treated with chidamide.
Project description:Proinflammatory stimuli rapidly and globally remodel chromatin landscape, thereby enabling transcriptional responses. Yet, the mechanisms coupling chromatin regulators to the master regulatory inflammatory transcription factor NF-kB remain poorly understood. We report in human endothelial cells (ECs) that activated NF-kB binds to enhancers, provoking a rapid, global redistribution of BRD4 preferentially at super-enhancers, large enhancer domains highly bound by chromatin regulators. Newly established NF-kB super-enhancers drive nearby canonical inflammatory response genes. In both ECs and macrophages BET bromodomain inhibition prevents super-enhancer formation downstream of NF-kB activation, abrogating proinflammatory transcription. In TNFa-activated endothelium this culminates in functional suppression of leukocyte rolling, adhesion and transmigration. Sustained BET bromodomain inhibitor treatment of LDLr -/- animals suppresses atherogenesis, a disease process rooted in pathological vascular inflammation involving endothelium and macrophages. These data establish BET-bromodomains as key effectors of inflammatory response through their role in the dynamic, global reorganization of super-enhancers during NF-kB activation. ChIP-Seq for various transcription factors, RNA Polymerase II, and histone modifications in human endothelial cells
Project description:Transcriptionnal profiling of C. perfringens 13 strain comparing growth in minimal medium with 1 mM homocysteine with growth in minimal medium with 0.5 mM cystine.