Establishing the transcriptional programme for blood: the SCL stem cell enhancer is regulated by a multiprotein complex containing Ets and GATA factors.
Stem cells are a central feature of metazoan biology. Haematopoietic stem cells (HSCs) represent the best-characterized example of this phenomenon, but the molecular mechanisms responsible for their formation remain obscure. The stem cell leukaemia (SCL) gene encodes a basic helix-loop-helix (bHLH) transcription factor with an essential role in specifying HSCs. Here we have addressed the transcriptional hierarchy responsible for HSC formation by characterizing an SCL 3'' enhancer that targets expression to HSCs and endothelium and their bipotential precursors, the haemangioblast. We have identified three critical motifs, which are essential for enhancer function and bind GATA-2, Fli-1 and Elf-1 in vivo. Our results suggest that these transcription factors are key components of an enhanceosome responsible for activating SCL transcription and establishing the transcriptional programme required for HSC formation.
PubMed ID: 12065417
PMC ID: PMC126046
Article link: EMBO J.
Genes referenced: efna2 elf1 fli1 gata2 tal1
Article Images: [+] show captions
|Fig. 4. The +19 core enhancer is active in transgenic frogs and murine haematopoietic progenitor cells and critically depends on regions 1, 2 and 3. (A) An example of the variable ectopic expression patterns seen in a minority of transgenic stage 32 embryos with the SV/GFP construct (ectopic expression was never seen in the DLP region). (B) SV/GFP/H2.5 transgenic embryo showing reporter gene expression in the DLP region (arrow). (C–E) Stage 27 SV/GFP/H2.5 transgenic embryo stained for GFP (BCIP, turquoise) and endogenous SCL (BM Purple). d/e indicates the plane of the section shown in (D) and (E) which demonstrate that the purple staining (endogenous SCL) is contained within the area of transgene expression (turquoise) in the DLP (arrow). (F) Summary of transgenic frog experiments using wild-type and mutant versions of the SH0.6 core enhancer. DLP, embryos showing DLP staining with or without ectopic expression; ectopic, embryos showing ectopic expression only. The number in each bar represents the number of GFP-expressing embryos for each category. (G) The SV/luc/SH0.6 construct was active in 416B but not BW5147 cells in stable transfection assays. (H) Stable transfection assays in 416B cells demonstrate that the SH0.6 enhancer activity critically depends on the 227 bp 3′ fragment deleted in the SA0.4 construct, and that the three conserved regions mutated in the mut1, mut2 and mut3 constructs are each critical for enhancer function.|