The SCL complex regulates c-kit expression in hematopoietic cells through functional interaction with Sp1.
The combinatorial interaction among transcription factors is believed to determine hematopoietic cell fate. Stem cell leukemia (SCL, also known as TAL1 [T-cell acute lymphoblastic leukemia 1]) is a tissue-specific basic helix-loop-helix (bHLH) factor that plays a central function in hematopoietic development; however, its target genes and molecular mode of action remain to be elucidated. Here we show that SCL and the c-Kit receptor are coexpressed in hematopoietic progenitors at the single-cell level and that SCL induces c-kit in chromatin, as ectopic SCL expression in transgenic mice sustains c-kit transcription in developing B lymphocytes, in which both genes are normally down-regulated. Through transient transfection assays and coimmunoprecipitation of endogenous proteins, we define the role of SCL as a nucleation factor for a multifactorial complex (SCL complex) that specifically enhances c-kit promoter activity without affecting the activity of myelomonocytic promoters. This complex, containing hematopoietic-specific (SCL, Lim-only 2 (LMO2), GATA-1/GATA-2) and ubiquitous (E2A, LIM- domain binding protein 1 [Ldb-1]) factors, is tethered to DNA via a specificity protein 1 (Sp1) motif, through direct interactions between elements of the SCL complex and the Sp1 zinc finger protein. Furthermore, we demonstrate by chromatin immunoprecipitation that SCL, E2A, and Sp1 specifically co-occupy the c-kit promoter in vivo. We therefore conclude that c-kit is a direct target of the SCL complex. Proper activation of the c-kit promoter depends on the combinatorial interaction of all members of the complex. Since SCL is down-regulated in maturing cells while its partners remain expressed, our observations suggest that loss of SCL inactivates the SCL complex, which may be an important event in the differentiation of pluripotent hematopoietic cells.
Lécuyer
Eric
E
Clinical Research Institute of Montreal and from the Departments of Pharmacology, Biochemistry, and Molecular Biology, Université de Montréal, Quebec, Canada.
Herblot
Sabine
S
Saint-Denis
Marianne
M
Martin
Richard
R
Begley
C Glenn
CG
Porcher
Catherine
C
Orkin
Stuart H
SH
Hoang
Trang
T
D016475:3T3 Cells; D000818:Animals; D001483:Base Sequence; D051792:Basic Helix-Loop-Helix Proteins; D001854:Bone Marrow Cells / Q000166:cytology / Q000502:physiology; D002460:Cell Line; D017931:DNA Primers; D004268:DNA-Binding Proteins / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D005786:Gene Expression Regulation; D018257:Helix-Loop-Helix Motifs; D006412:Hematopoietic Stem Cells / Q000502:physiology*; D051379:Mice; D018345:Mice, Knockout; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D011518:Proto-Oncogene Proteins / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D019009:Proto-Oncogene Proteins c-kit / Q000235:genetics*; D011519:Proto-Oncogenes; D011994:Recombinant Proteins / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016329:Sp1 Transcription Factor / Q000378:metabolism*; D000073942:T-Cell Acute Lymphocytic Leukemia Protein 1; D014157:Transcription Factors / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D014162:Transfection
D051792:Basic Helix-Loop-Helix Proteins; D017931:DNA Primers; D004268:DNA-Binding Proteins; D011518:Proto-Oncogene Proteins; D019009:Proto-Oncogene Proteins c-kit; D011994:Recombinant Proteins; D016329:Sp1 Transcription Factor; D000073942:T-Cell Acute Lymphocytic Leukemia Protein 1; D014157:Transcription Factors; C489366:Tal1 protein, mouse