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TET enzymes are dioxygenases that promote DNA demethylation by oxidizing the methyl group of 5-methylcytosine to 5-hydroxymethylcytosine (5hmC). Here,we report a close correspondence between 5hmC-marked regions,chromatin accessibility and enhancer activity in B cells,and a strong enrichment for consensus binding motifs for basic region-leucine zipper (bZIP) transcription factors at TET-responsive genomic regions. Functionally,Tet2 and Tet3 regulate class switch recombination (CSR) in murine B cells by enhancing expression of Aicda,which encodes the activation-induced cytidine deaminase (AID) enzyme essential for CSR. TET enzymes deposit 5hmC,facilitate DNA demethylation,and maintain chromatin accessibility at two TET-responsive enhancer elements,TetE1 and TetE2,located within a superenhancer in the Aicda locus. Our data identify the bZIP transcription factor,ATF-like (BATF) as a key transcription factor involved in TET-dependent Aicda expression. 5hmC is not deposited at TetE1 in activated Batf-deficient B cells,indicating that BATF facilitates TET recruitment to this Aicda enhancer. Our study emphasizes the importance of TET enzymes for bolstering AID expression and highlights 5hmC as an epigenetic mark that captures enhancer dynamics during cell activation. View Publication

Extracellular cold-inducible RNA-binding protein (CIRP) exaggerates inflammation and tissue injury in sepsis. Neutrophil extracellular traps (NETs) are released by activated neutrophils during sepsis. NETs contribute to pathogen clearance,but excessive NET formation (NETosis) causes inflammation and tissue damage. Peptidylarginine deiminase 4 (PAD4) is associated with NETosis by increasing histone citrullination and chromatin decondensation. We hypothesized that CIRP induces NETosis in the lungs during sepsis via upregulating PAD4 expression. Sepsis was induced in C57BL/6 wild-type (WT) and CIRP-/- mice by cecal ligation and puncture (CLP). After 20 h of CLP induction,NETs in the lungs of WT and CIRP-/- mice were quantified by flow cytometry by staining the single cell suspensions with MPO and CitH3 Abs. PAD4 expression in the lungs of WT and CIRP-/- mice after sepsis was assessed by Western blotting. In vitro effects of recombinant mouse (rm) CIRP for NETosis and PAD4 expression in the bone marrow-derived neutrophils (BMDN) were assessed by flow cytometry and Western blotting,respectively. After 20 h of CLP,NETosis in the lungs was significantly decreased in CIRP-/- mice compared to WT mice,which also correlated with the decreased PAD4 expression. Intratracheal administration of rmCIRP into WT mice significantly increased NETosis and PAD4 expression in the lungs compared to vehicle-injected mice. In vitro culture of BMDN with rmCIRP significantly increased NETosis and PAD4 expression compared to PBS-treated control. Fluorescence microscopy revealed typical web-like structures consistent with NETs in rmCIRP-treated BMDN. Thus,CIRP serves as a novel inducer of NETosis via PAD4 during sepsis. View Publication