Erythroid Krüppel-Like Factor Directly Activates the Basic Krüppel-Like Factor Gene in Erythroid Cells

Abstract

The Sp/Krüppel-like factor (Sp/Klf) family is comprised of around 25 zinc finger transcription factors that recognize CACCC boxes and GC-rich elements. We have investigated basic Krüppel-like factor (Bklf/Klf3) and show that in erythroid tissues its expression is highly dependent on another family member, erythroid Krüppel-like factor (Eklf/Klf1). We observe that Bklf mRNA is significantly reduced in erythroid tissues from Eklf-null murine embryos. We find that Bklf is driven primarily by two promoters, a ubiquitously active GC-rich upstream promoter, 1a, and an erythroid downstream promoter, 1b. Transcripts from the two promoters encode identical proteins. Interestingly, both the ubiquitous and the erythroid promoter are dependent on Eklf in erythroid cells. Eklf also activates both promoters in transient assays. Experiments utilizing an inducible form of Eklf demonstrate activation of the endogenous Bklf gene in the presence of an inhibitor of protein synthesis. The kinetics of activation are also consistent with Bklf being a direct Eklf target. Chromatin immunoprecipitation assays confirm that Eklf associates with both Bklf promoters. Eklf is typically an activator of transcription, whereas Bklf is noted as a repressor. Our results support the hypothesis that feedback cross-regulation occurs within the Sp/Klf family in vivo.

This work was supported by NIH grant NHLBI HL073443 and grants from the Australian ARC and NHMRC to M.C. A.F. is supported by an Australian Postgraduate Award.

We are grateful to Menie Merika and Stuart Orkin for providing pPac and pPac-Eklf and to James Bieker for supplying pSG5-Eklf. We also thank Hannah Nicholas, Richard Pearson, and Alexis Verger for their advice in preparing the manuscript. We express our gratitude to Dale Hancock, Denise Hodge, Richard Pearson, Nancy Sue, and Jane van Vliet for experimental support.