This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. [3H]luteolin binds nuclear type II [3H]estradiol binding sites covalently (Steroids 66707-719, 2001) and was used to identify the protein(s). Analyses of [3H]luteolin-labeled type II sites on SDS PAGE demonstrated specific binding to 11-kDa and 35-kDa proteins that were also detected in type II site preparations purified by C18-reversed phase HPLC. The 11-kDa protein was sequenced and identified as histone H4 and Western blotting confirmed the 11-kDa and 35-kDa proteins were acetylated forms of histone H4. Anti-histone H4 antibodies (but not H2A, H2B or H3 antibodies) immunoadsorbed type II sites from nuclear extracts and this was neutralized by specific competing peptides. Rabbit reticulocyte lysate translated histone H4, but not histones H2A, H2B or H3, demonstrated specific [3H]estradiol binding. Thus, histone H4 is the type II site and regulatory ligands such as methyl p-hydroxyphenyllactate (MeHPLA) and luteolin may control cell proliferation via modulation of hi stone H4 function.
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