We previously showed that gamma interferon (IFN gamma) and its receptor subunit, IFNGR1, interacted with the promoter region of IFN gamma-activated genes along with transcription factor STAT1 alpha. Recent studies have suggested that activated Janus kinases pJAK2 and pJAK1 also played a role in gene activation by phosphorylation of histone H3 on tyrosine 41. This study addresses the question of the role of activated JAKs in specific gene activation by IFN gamma. We carried out chromatin immunoprecipitation (ChIP) followed by PCR in IFN gamma treated WISH cells and showed association of pJAK1, pJAK2, IFNGR1, and STAT1 on the same DNA sequence of the IRF-1 gene promoter. The beta-actin gene, which is not activated by IFN gamma, did not show this association. The movement of activated JAK to the nucleus and the IRF-1 promoter was confirmed by the combination of nuclear fractionation, confocal microscopy and DNA precipitation analysis using the biotinylated GAS promoter. Activated JAKs in the nucleus was associated with phosphorylated tyrosine 41 on histone H3 in the region of the GAS promoter. Unphosphorylated JAK2 was found to be constitutively present in the nucleus and was capable of undergoing activation in IFN gamma treated cells, most likely via nuclear IFNGR1. Association of pJAK2 and 1FNGR1 with histone H3 in IFN gamma treated cells was demonstrated by histone H3 immunoprecipitation. Unphosphorylated STAT1 protein was associated with histone H3 of untreated cells. IFN gamma treatment resulted in its disassociation and then re-association as pSTAT1. The results suggest a novel role for activated JAKs in epigenetic events for specific gene activation. (C) 2011 Elsevier Inc. All rights reserved.