ETO-2 Associates with SCL in Erythroid Cells and Megakaryocytes and Provides Repressor Functions in Erythropoiesis

Abstract

Lineage specification and cellular maturation require coordinated regulation of gene expression programs. In large part, this is dependent on the activator and repressor functions of protein complexes associated with tissue-specific transcriptional regulators. In this study, we have used a proteomic approach to characterize multiprotein complexes containing the key hematopoietic regulator SCL in erythroid and megakaryocytic cell lines. One of the novel SCL-interacting proteins identified in both cell types is the transcriptional corepressor ETO-2. Interaction between endogenous proteins was confirmed in primary cells. We then showed that SCL complexes are shared but also significantly differ in the two cell types. Importantly, SCL/ETO-2 interacts with another corepressor, Gfi-1b, in red cells but not megakaryocytes. The SCL/ETO-2/Gfi-1b association is lost during erythroid differentiation of primary fetal liver cells. Genetic studies of erythroid cells show that ETO-2 exerts a repressor effect on SCL target genes. We suggest that, through its association with SCL, ETO-2 represses gene expression in the early stages of erythroid differentiation and that alleviation/modulation of the repressive state is then required for expression of genes necessary for terminal erythroid maturation to proceed.

SUPPLEMENTAL MATERIAL

Supplemental material for this article may be found at http://mcb.asm.org/.

ACKNOWLEDGMENTS

We are very grateful to Anthony Jones and his team (Functional Genomics and Proteomics Laboratories, University of Birmingham) for mass spectrometry analyses; to Hedia Chagraoui, Mira Kassouf, and Douglas Vernimmen for invaluable advice; to Lauren Aronson for technical expertise; to Craig Waugh for FACS sorting; and to Ian Hickson and Roger Patient for critical reading of the manuscript.

A.H.S. is a recipient of a Leukemia Research Fund Clinical Scientist Award. T.E.'s laboratory is supported by a specialist program from the Leukemia Research Fund. P.V. is a Wellcome Trust Senior Clinical Fellow. C.P. is funded by the Medical Research Council.