CD18 (␤ 2 leukocyte integrin) is a leukocyte-specific adhesion molecule that plays a crucial role in immune and inflammatory responses. A 79-nucleotide fragment of the CD18 promoter is sufficient to direct myeloid transcription. The CD18 promoter is bound by the B lymphocyte-and myeloid-restricted ets factor, PU.1, and disruption of the PU.1-binding sites significantly reduces promoter activity. However, PU.1 alone cannot fully account for the leukocyte-specific and myeloid-inducible transcription of CD18. We identified a ubiquitously expressed nuclear protein complex of extremely low electrophoretic mobility that also binds to this region of the CD18 promoter. This binding complex is a heterotetramer composed of GABP␣, an ets factor, and GABP␤, a subunit with homology to Drosophila Notch. GABP␣ competes with the lineage restricted factor, PU.1, for the same critical CD18 ets sites. The CD18 promoter is activated in myeloid cells by transfection with both GABP␣ and GABP␤ together, but not by either factor alone. Transfection of non-hematopoietic cells with the two GABP subunits together with PU

Normal cellular differentiation is linked to tightly regulated gene transcription. However, the DNA elements and trans-acting factors that regulate transcription in myeloid cells are poorly defined. CD18, the i3 chain of the leukocyte integrins, is transcriptionally regulated during myeloid differentiation. The CD18 promoter is active after transfection into myeloid cells. We demonstrate that a region of the CD18 promoter that contains two binding sites for the PU.1 transcription factor is required for activity in myeloid cells.These sites are bound by in vitro translated PU.i and by PU.A from myeloid nuclear extracts. Mutagenesis of these sites abrogates binding by PU.1 and substantially decreases promoter activity in myelold cells. Thus, the leukocyte-specific transcription factor PU.1 is required for myeloid activity of CD18.Monocytes and granulocytes, which are collectively referred to as myeloid cells, play critical roles in the immune and inflammatory responses. These mature hematopoietic cells differentiate from pluripotent bone marrow precursors under the influence of cytokines and other regulatory molecules. Cellular differentiation is linked to tightly regulated gene expression, and such regulation is frequently controlled at the level of transcription (1). Acute leukemia is a clinical manifestation of disordered hematopoietic differentiation; some types of acute leukemia are caused by abnormalities of transcription factors.Little is known of the sequence-specific DNA motifs and the trans-acting factors that regulate transcription in normal myeloid differentiation. PU.1, a member of the ets family of transcription factors, is expressed by B lymphocytes and monocyte/macrophages. It binds to DNA in a sequencespecific manner and transcriptionally regulates genes whose regulatory elements contain the PU

CD18, the ␤ chain of the leukocyte integrins, plays a crucial role in immune and inflammatory responses. CD18 is expressed exclusively by leukocytes, and it is transcriptionally regulated during the differentiation of myeloid cells. The ets factors, PU.1 and GABP, bind to three ets sites in the CD18 promoter, which are essential for high level myeloid expression of CD18. We now identify two binding sites for the transcription factor, Sp1, that flank these ets sites. Sp1 is the only factor from myeloid cells that binds to these sites in a sequencespecific manner. Mutagenesis of these sites abrogates Sp1 binding and significantly reduces the activity of the transfected CD18 promoter in myeloid cells. Transfection of Sp1 into Drosophila Schneider cells, which otherwise lack Sp1, activates the CD18 promoter dramatically. GABP also activates the CD18 promoter in Schneider cells. Co-transfection of Sp1 and GABP activates CD18 more than the sum of their individual effects, indicating that these factors cooperate to transcriptionally activate myeloid expression of CD18. These studies support a model of high level, lineagerestricted gene expression mediated by cooperative interactions between widely expressed transcription factors.Monocytes and granulocytes, which are collectively known as myeloid cells, play crucial roles in immune and inflammatory responses. As myeloid cells differentiate from immature bone marrow precursor cells, they express characteristic genes that are required in their roles as immune effector cells. Most genes that are expressed during myeloid differentiation are regulated at the level of transcription (1). Characterization of the DNA sequences and transcription factors that control myeloid gene expression has provided important insights into the molecular basis of normal myeloid differentiation.CD18 (␤ 2 leukocyte integrin) is a cell surface adhesion molecule that forms heterodimers with CD11a, CD11b, or CD11c to generate the antigens LFA-1, Mo-1 (Mac-1), and p150/95, respectively. These leukocyte-specific receptors mediate cell-cell and cell-matrix interactions and play important roles in immune and inflammatory responses (2). The clinical significance of CD18 is illustrated by leukocyte adhesion deficiency, in which insufficient CD18 causes recurrent bacterial and fungal infections and can lead to premature death (3).Expression of CD18 is restricted to myeloid cells and lymphocytes. CD18 expression increases significantly during myeloid differentiation due to increased transcription (4 -7). We (4) and others (8, 9) have cloned the gene that encodes CD18 and characterized its promoter. The transfected CD18 promoter exhibits the leukocyte-specific and myeloid-inducible activity of the endogenous CD18 gene (10). We have previously shown that two ets-related transcription factors, PU.1 and GABP, bind to the CD18 promoter and cooperate to activate leukocyte-specific expression of CD18 (10, 11).Sp1 is a DNA-binding nuclear protein that functions as a transcriptional activator (12)(13)(14). We now de...

1,25-Dihydroxyvitamin D3 inhibits the proliferation of the chronic myelogenous leukemia cell line RWLeu-4 but not the resistant variant, JMRD3. Although these cells exhibit no detectable differences in the vitamin D receptor, alterations in the interaction of nuclear extracts with the osteocalcin-1,25-dihydroxyvitamin D3-response element are noted. It is shown herein that the 1,25-dihydroxyvitamin D3 receptor binds to the osteocalcin-1,25-dihydroxyvitamin D3-response element along with activator protein-1 (AP-1) complexes and that the DNA binding activities of members of the Jun and Fos proto-oncogene families, which make up the AP-1 transcription factor, are differentially regulated by 1,25-dihydroxyvitamin D3. It is shown that JunD DNA binding activity is enhanced by 1,25-dihydroxyvitamin D3 during cell cycle arrest in the sensitive cells but is decreased in the resistant cells. These results suggest that the level of JunD DNA binding activity may be a critical factor in the regulation of proliferation.