Chromatin Boundaries Require the Functional Cooperation Between hSET1 and NURF Complexes

Abstract 646

Chromatin modification and remodeling activities play a central role in organizing nuclear functions in eukaryotic genome. Chromatin domains with characteristic epigenetic marks are organized by chromatin insulator. The chicken b-globin insulator, 5′HS4, is an excellent model system to study how insulator maintains gene function and prevents the encroachment of repressive heterochromatin. We showed previously that USF1/2 bound 5′HS4 insulator mediates chromatin barrier activity by recruiting and organizing active histone modifications in the chicken b-globin locus. However, it is not clear how insulator establishes such a barrier. To further understand the physical function of USF1, we purified the USF1-associated protein complexes. We found that USF1 forms a multiprotein complex with hSET1 and NURF exhibiting histone H3K4 methyltransferase and ATP-dependent chromatin remodeling activities, respectively. Both hSET1 and NURF complexes are specifically recruited to the 5′HS4 insulator by USF1 to retain the active chromatin structure. Knock-down of BPTF, a component of NURF complex, resulted in a rapid loss of barrier activity that protects a transgene from chromatin silencing. The loss of barrier by the BPTF knock-down is accompanied by an alternation of nucleosome positioning that expands the nucleosome to the nucleosome free linker region in the HS4 insulator site and increases repressive H3K9 dimethylation. Furthermore, the suppression of hSET1, a H3K4 methyltransferase, leads to not only a loss of barrier activity, but also a loss of H3K4 dimethylation and H3K9/K14 acetylation, as well as the recruitment of BPTF at chromatin insulator sites. Thus, our data reveal a molecular mechanism in which histone modifying enzyme hSET1 and chromatin remodeling complex NURF collaborate together to maintain the 5′HS4 chromatin barrier activity and to prevent the encroachment of adjacent heterochromatin.