An Ikaros-Containing Chromatin-Remodeling Complex in Adult-Type Erythroid Cells

Abstract

We have previously described a SWI/SNF-related protein complex (PYR complex) that is restricted to definitive (adult-type) hematopoietic cells and that specifically binds DNA sequences containing long stretches of pyrimidines. Deletion of an intergenic DNA-binding site for this complex from a human ␤-globin locus construct results in delayed human ␥-to ␤-globin switching in transgenic mice, suggesting that the PYR complex acts to facilitate the switch. We now show that PYR complex DNA-binding activity also copurifies with subunits of a second type of chromatin-remodeling complex, nucleosome-remodeling deacetylase (NuRD), that has been shown to have both nucleosome-remodeling and histone deacetylase activities. Gel supershift assays using antibodies to the ATPase-helicase subunit of the NuRD complex, Mi-2 (CHD4), confirm that Mi-2 is a component of the PYR complex. In addition, we show that the hematopoietic cell-restricted zinc finger protein Ikaros copurifies with PYR complex DNA-binding activity and that antibodies to Ikaros also supershift the complex. We also show that NuRD and SWI/SNF components coimmunopurify with each other as well as with Ikaros. Competition gel shift experiments using partially purified PYR complex and recombinant Ikaros protein indicate that Ikaros functions as a DNA-binding subunit of the PYR complex. Our results suggest that Ikaros targets two types of chromatin-remodeling factors-activators (SWI/SNF) and repressors (NuRD)-in a single complex (PYR complex) to the ␤-globin locus in adult erythroid cells. At the time of the switch from fetal to adult globin production, the PYR complex is assembled and may function to repress ␥-globin gene expression and facilitate ␥-to ␤-globin switching.

Heterogeneous nuclear ribonucleoprotein C1/C2, MeCP1, and SWI/SNF form a chromatin remodeling complex at the β-globin locus control region

“…Reports on the recruitment of chromatin remodeling complexes on the ␤-globin locus are restricted to the ␤-globin promoter and a pyrimidine-rich DNA sequence upstream of the ␦-globin promoter (30)(31)(32). GATA-1 and MafK have been described as associated with chromatin remodeling complexes (33,34,44).…”

The role of Ikaros in human erythroid differentiation

“…In the second step (days 10-13), cells were diluted at 5 ϫ 10 4 or 10 5 cells/mL (for CB or mPB, respectively) and cocultured on an adherent MS5 stromal layer in basal medium supplemented with erythropoietin. In the third step (days [13][14][15][16][17][18][19][20], cells were cultured on MS5 cells in basal medium without cytokine.…”

“…In MEL (murine erythroleukemia) cells, Ikaros is associated with the chromatin remodelling PYR complex which binds to an intergenic domain between the ␥-globin and ␤-globin genes, and facilitates the switch between these 2 globins. 14 Moreover, Ikaros null mice show a delay in murine embryonic to adult ␤-globin switching, due to the lack of the chromatin remodeling PYR complex. Delayed switching between human ␥-and ␤-globins is also observed in mice with a human ␤-globin minilocus and lacking the Ikaros gene.…”

When the SWI/SNF complex remodels … the cell cycle

“…Within this complex, Brg1, Baf170, Baf155 and SNF5/INI1 are associated with the histone deacetylase HDAC3 and the corepressor Kap1/TIF1b (Underhill et al, 2000). In lymphoid cells, SWI/SNF subunits are also associated with the transcriptional regulator Ikaros required for normal B and T cell di erentiation (Kim et al, 1999;O'Neill et al, 2000). Similarly, erythroid cells contain a SWI/SNF-like complex associated with EKLF, a key regulator of b-globin gene expression (Armstrong et al, 1998;Kadam et al, 2000).…”