Extrinsic and intrinsic control by EKLF (KLF1) within a specialized erythroid niche.

The erythroblastic island provides an important nutritional and survival support niche for efficient erythropoietic differentiation. Island integrity is reliant on adhesive interactions between erythroid and macrophage cells. We show that erythroblastic islands can be formed from single progenitor cells present in differentiating embryoid bodies, and that these correspond to erythro-myeloid progenitors (EMPs) that first appear in the yolk sac of the early developing embryo. Erythroid Krüppel-like factor (EKLF; KLF1), a crucial zinc finger transcription factor, is expressed in the EMPs, and plays an extrinsic role in erythroid maturation by being expressed in the supportive macrophage of the erythroblastic island and regulating relevant genes important for island integrity within these cells. Together with its well-established intrinsic contributions to erythropoiesis, EKLF thus plays a coordinating role between two different cell types whose interaction provides the optimal environment to generate a mature red blood cell.

Novel role for EKLF in megakaryocyte lineage commitment.

Megakaryocytes and erythroid cells are thought to derive from a common progenitor during hematopoietic differentiation. Although a number of transcriptional regulators are important for this process, they do not explain the bipotential result. We now show by gain- and loss-of-function studies that erythroid Krüppel-like factor (EKLF), a transcription factor whose role in erythroid gene regulation is well established, plays an unexpected directive role in the megakaryocyte lineage. EKLF inhibits the formation of megakaryocytes while at the same time stimulating erythroid differentiation. Quantitative examination of expression during hematopoiesis shows that, unlike genes whose presence is required for establishment of both lineages, EKLF is uniquely down-regulated in megakaryocytes after formation of the megakaryocyte-erythroid progenitor. Expression profiling and molecular analyses support these observations and suggest that megakaryocytic inhibition is achieved, at least in part, by EKLF repression of Fli-1 message levels.

Altered regulation of beta-like globin genes by a redesigned erythroid transcription factor.

OBJECTIVE: Targeted regulation of beta-like globin genes was studied using designer zinc finger transcription factors containing the DNA binding domain of the red cell specific transcription factor erythroid Kruppel-like factor (EKLF) fused to repression domains. METHODS: Globin gene expression was analyzed after introduction of the modified transcription factors into cell lines, embryonic stem cells and transgenic mice. RESULTS: As would be predicted, when introduced transiently into cells these transcription factors were effective in repressing the adult beta-globin promoter CACCC element, which is the natural target for EKLF. In murine erythroleukemia cells repression of the adult beta-globin gene was accompanied by a reactivation of the endogenous embryonic betaH1-globin gene. Studies in differentiated embryonic stem cells and transgenic mice confirmed the reactivation of embryonic gene expression during development. CONCLUSION: Our studies support a competition model for beta-globin gene expression and underscore the importance of EKLF in the embryonic/fetal-to-adult globin switch. They also demonstrate the feasibility of designer zinc finger transcription factors in the study of transcriptional control mechanisms at the beta-globin locus and as potential gene therapy agents for sickle cell disease and related hemoglobinopathies.