James J Bieker, PhD

 
     The molecular events that confer the ability to express lineage-specific genes upon an initially uncommitted, pluripotent hematopoietic stem cell remain a major question in cell differentiation. Use of an immortalized erythroid cell line as a means to isolate genes that may be important for erythroid function allowed us to identify a novel, erythroid-specific gene, which was named EKLF (erythroid Kruppel-like factor).

     EKLF binds to and activates transcription from the CACCC element, one of a trio of localized promoter and enhancer DNA binding sites known to be crucial for transcription of globin and othererythroid cell-specific genes. Biological analyses reveal that murine EKLF is expressed in primitive erythroid cells by embryonic day 7.5, and in definitive erythroid cells within the hepatic primordia by embryonic day 9.5. However, its ability to preferentially activate an adult ß-globin promoter over a linked fetal gamma-globin promoter led us to propose that EKLF may be an important factor for gamma- to ß-globin gene switching. This was verified by studies showing that EKLF is absolutely required for normal red cell development, since its genetic disruption leads to death by embryonic day 14-16 (precisely the time of the switch in mice) due to a deficiency of mature, definitive red cells. EKLF-deficient mice exhibit drastically low-globin expression at the transcriptional level, i.e., a severe ß-thalassemia phenotype, and contain an altered chromatin structure at the ß-globin locus. These molecular and biological studies have established that EKLF is an essential component required for globin switching and completion of the definitive erythroid program. Disorders of hemoglobin expression can lead to a variety of hemoglobinopathies, including sickle cell anemia and -thalassemia (Cooley's anemia). As a result, our examination of EKLF's mechanism of action has illuminated how it regulates the globin locus, and has provided us with a way to reconstruct EKLF so that it can potentially rectify one type of hemoglobin disorder. 

     Our discovery of EKLF has stimulated other investigators to search for analogous genes that can work in a similar fashion to regulate unique targets in other tissues.  EKLF is now the founding member (KLF1) of a family of seventeen proteins, some of which have been directly implicated in suppression of a specific subset of cancers.  We are vigorously continuing its study using a number of approaches, including biochemical and structure/function analyses of the EKLF protein, identification of its protein partners, and monitoring how EKLF expression itself is so precisely regulated during development. In addition, differentiating embryonic stem cells in culture are being used as one powerful approach to address these issues in diverse ways: to identify extracellular molecules and illuminate the intracellular pathway they use to play a directive role in erythroid gene expression; to functionally test the cis-acting sequences that control one of these downstream targets, EKLF; to establish gain-of-function studies that address lineage determination mechanisms during developing and differentiation; and to gain further insight into globin switching mechanisms and identify ways to alter the normal pattern of expression.

     Our recent studies show that EKLF becomes acetylated by virtue of its association with a subset of coactivators, leading to enhanced interaction with chromatin remodelers that leads to activated transcription. Surprisingly, EKLF can also associate with corepressors and decrease transcription at selected promoters, suggesting other activities beyond activation of the adult ß-globin gene.  Finally, the BMP4/Smad pathway plays a critical role in transcriptional activation of EKLF in the erythroid cell, likely via a relatively small promoter region proximal to its initiation site.

     For more information, please visit the Bieker Laboratory website.

     For a complete list of published work: http://www.ncbi.nlm.nih.gov/sites/myncbi/james.bieker.1/bibliography/40735946/public/?sort=date&direction=descending