HISTONES IN TRANSIT - CYTOSOLIC HISTONE COMPLEXES AND DIACETYLATION OF H4 DURING NUCLEOSOME ASSEMBLY IN HUMAN-CELLS

The organization and acetylation of nascent histones prior to their st able incorporation into chromatin were examined. Through sedimentation and immunoprecipitation analyses of HeLa cytosolic extracts, two soma tic non-nucleosomal histone complexes were detected: one containing na scent H3 and H4, and a second containing H2A (and probably H2B) in ass ociation with the nonhistone protein NAP-1. The H3/H4 complex has a se dimentation coefficient of 5-6S, consistent with the presence of one o r more escort proteins. H4 in the cytosolic H3/H4 complex is diacetyla ted, fully in accord with the acetylation state of newly synthesized H 4 in chromatin. The diacetylation of nascent human H4 is therefore com pleted prior to nucleosome assembly. As part of our studies of the nas cent H3/H4 complex, the cytoplasmic histone acetyltransferase most lik ely responsible for acetylating newly synthesized H4 was also investig ated. HeLa histone acetyltransferase B (HAT B) acetylates H4 but not H 3 in vitro, and maximally diacetylates H4 even in the presence of sodi um butyrate. Human HAT B acetylates H4 exclusively on the lysine resid ues at positions 5 and 12, in complete agreement with the highly conse rved acetylation pattern of nascent nucleosomal H4 (Sobel et al., 1995 ), and has a native molecular weight of similar to 100 kDa. Based on o ur findings a model is presented for the involvement of histone acetyl ation and NAP-1 in H2A/H2B deposition and exchange, during nucleosome assembly and chromatin remodeling in vivo.