Centromeres of most organisms are embedded within constitutive heterochromatin, the condensed regions of chromosomes that account for a large fraction of complex genomes. The functional significance of this centromere-heterochromatin relationship, if any, is unknown. One possibility is that heterochromatin provides a suitable environment for assembly of centromere components, such as special centromeric nucleosomes that contain distinctive histone H3-like proteins. We describe a Drosophila H3-like protein, Cid (for centromere identifier) that localizes exclusively to fly centromeres. When the cid upstream region drives expression of H3 and H2B histone-green fluorescent protein fusion genes in Drosophila cells, euchromatin-specific deposition results. Remarkably, when the cid upstream region drives expression of yeast, worm, and human centromeric histone-green fluorescent protein fusion proteins, localization is preferentially within Drosophila pericentric heterochromatin. Heterochromatin-specific localization also was seen for yeast and worm centromeric proteins constitutively expressed in human cells. Preferential localization to heterochromatin in heterologous systems is unexpected if centromere-specific or site-specific factors determine H3-like protein localization to centromeres. Rather, the heterochromatic state itself may help localize centromeric components.centromeres ͉ heterochromatin ͉ Drosophila

Heterochromatin protein 1 (HP1) is a non‐histone chromosomal protein in Drosophila with dosage‐dependent effects on heterochromatin‐mediated gene silencing. An evolutionarily conserved amino acid sequence in the N‐terminal half of HP1 (the ‘chromo domain’) shares > 60% sequence identity with a motif found in the Polycomb protein, a silencer of homeotic genes. We report here that point mutations in the HP1 chromo domain abolish the ability of HP1 to promote gene silencing. We show that the HP1 chromo domain, like the Polycomb chromo domain, has chromosome binding activity, but to distinct chromosomal sites. We constructed a chimeric HP1‐Polycomb protein, consisting of the chromo domain of Polycomb in the context of HP1, and show that it binds to both heterochromatin and Polycomb binding sites in polytene chromosomes. In flies expressing chimeric HP1‐Polycomb protein, endogenous HP1 is mislocalized to Polycomb binding sites, and endogenous polycomb is misdirected to the heterochromatic chromocenter, suggesting that both proteins are recruited to their distinct chromosomal binding sites through protein‐protein contacts. Chimeric HP1‐Polycomb protein expression in transgenic flies promotes heterochromatin‐mediated gene silencing, supporting the view that the chromo domain homology reflects a common mechanistic basis for homeotic and heterochromatic silencing.

We report the isolation and characterization of cDNA clones that encode a protein with the same DNA binding specificity as the immunoglobulin heavy chain enhancer binding protein E {ttEBP-E). We call the gene encoding this protein Ig/EBP-I. A fusion protein encoded by the cDNA binds specifically to ttEBP-E-binding sites (E sites) in both the IgH enhancer and the Vnl promoter. Sequence analysis reveals that Ig/EBP-I is a member of the "basic-zipper" family of DNA-binding proteins that are characterized by basic regions and heptad repeats of leucine residues. Among known family members, Ig/EBP-I demonstrates highest homology to C/EBP throughout the DNA-binding domain and leucine repeat region. Ig/EBP-I and C/EBP have highly overlapping binding specificities; both cloned proteins bind to the IgH enhancer and the VH1 promoter E sites, and Ig/EBP-1 binds to previously characterized C/EBP binding sites in the Rous sarcoma virus (RSV) LTR and the murine albumin promoter. Consistent with their homology in the leucine repeat region, Ig/EBP-I and C/EBP form heterodimers; Ig/EBP-I is the first member of this family that has been found to heterodimerize with the wellcharacterized C/EBP. Ig/EBP-I mRNA is present in all tissues and cell lines examined, although its levels vary almost 20-fold from different sources, with highest levels in early B cells. In tissues where Ig/EBP-1 and C/EBP are both present, heterodimers may be functionally important. The presence of Ig/EBP-I in fibroblasts and other tissues where C/EBP is not expressed suggests that Ig/EBP-I may be functionally important for the activity of the RSV enhancer in these cell types. Finally, elevated expression of Ig/EBP-I in early B cells may explain in part the enhancer-independent activity of Vx promoters early in B-cell development.

We examined the heterochromatic binding of GAGA factor and proliferation disrupter (Prod) proteins during the cell cycle in Drosophila melanogaster and sibling species. GAGA factor binding to the brownD ominant AG-rich satellite sequence insertion was seen at metaphase, however, no binding of GAGA factor to AG-rich sequences was observed at interphase in polytene or diploid nuclei. Comparable mitosis-specific binding was found for Prod protein to its target satellite in pericentric heterochromatin. At interphase, these proteins bind numerous dispersed sites in euchromatin, indicating that they move from euchromatin to heterochromatin and back every cell cycle. The presence of Prod in heterochromatin for a longer portion of the cell cycle than GAGA factor suggests that they cycle between euchromatin and heterochromatin independently. We propose that movement of GAGA factor and Prod from high affinity sites in euchromatin occurs upon condensation of metaphase chromosomes. Upon decondensation, GAGA factor and Prod shift from low affinity sites within satellite DNA back to euchromatic sites as a self-assembly process.