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Microwave oven-assisted derivatization of histone proteoforms using trimethylacetic anhydride

ABSTRACT: We introduce a new reagent – Trimethylacetic anhydride (TMA) for highly efficient histone derivatization, which is facilitated by microwave irradiation to reduce sample preparation duration. The derivatization of unmodified NH2- groups at histone lysine residues is necessary prior LC-MS analysis using bottom-up approach. Our methodology shows high labelling efficiency and demonstrates its utilization for histone posttranslational modification abundance monitoring. Therefore, to be an alternative to conventionally used methods.

INSTRUMENT(S): Orbitrap Fusion Lumos

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): B Cell, Cell Culture

SUBMITTER: Hana Kucharikova

LAB HEAD: Zbynek Zdrahal

PROVIDER: PXD019502 | Pride | 2021-06-14

REPOSITORIES: Pride

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Project description:Dramatic changes of gene expressions are known to occur in human endometrial stromal cells (ESC) during decidualization. The changes in gene expression are associated with changes of chromatin structure, which are regulated by epigenetic mechanisms such as histone modifications. Here, we investigated genome-wide changes in histone modifications and mRNA expressions associated with decidualization in human ESC using chromatin immunoprecipitation (ChIP) combined with next-generation sequencing. ESC were incubated with estradiol and medroxyprogesterone acetate for 14 days to induce decidualization. The ChIP-sequence data showed that induction of decidualization increased H3K27ac and H3K4me3 signals in many genomic regions but decreased in only a few regions. Most (80%) of the H3K27ac-increased regions and half of the H3K4me3-increased regions were located in the distal promoter regions (more than 3 kb upstream or downstream of the transcription start site). RNA-sequence showed that induction of decidualization up-regulated 881 genes, 223 of which had H3K27ac- or H3K4me3-increased regions in the proximal and distal promoter regions. Induction of decidualization increased the mRNA levels of these genes more than it increased the mRNA levels of genes without H3K27ac- or H3K4me3-increased regions. Pathway analysis revealed that up-regulated genes with the H3K27ac- or H3K4me3-increased regions were associated with insulin signaling. These results show that histone modification statuses genome-widely change in human ESC by induction of decidualization. The main changes of histone modifications are increases of H3K27ac and H3K4me3 in both the proximal and distal promoter regions, which are involved in the up-regulation of gene expression that occurs during decidualization. Examination of 4 different histone modifications in human endometrial stromal cells with or without EP inductions for 2 individuals. (EP induction: induction with estradiol (10-8 M) and medroxyprogesterone acetate (10-6 M))

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Microwave oven-assisted derivatization of histone proteoforms using trimethylacetic anhydride

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