Stage-Specific Repression by the EKLF Transcriptional Activator

Abstract

Dynamic changes in transcription factor function can be mediated by switching its interaction with coactivators and corepressors. Erythroid Krüppel-like factor (EKLF) is an erythroid cell-specific transcription factor that plays a critical role in ␤-globin gene activation via its interactions with CBP/p300 and SWI/SNF proteins. Unexpectedly, it also interacts with Sin3A and histone deacetylase 1 (HDAC1) corepressors via its zinc finger domain. We now find that selected point mutants can uncouple activation and repression and that an intact finger structure is not required for interactions with Sin3A/HDAC1 or for transrepression. Most intriguingly, EKLF repression exhibits stage specificity, with reversible EKLF-Sin3A interactions playing a key role in this process. Finally, we have located a key lysine residue that is both a substrate for CBP acetylation and required for Sin3A interaction. These data suggest a model whereby the stage of the erythroid cell alters the acetylation status of EKLF and plays a critical role in directing its coactivator-corepressor interactions and downstream transcriptional effects.A pivotal control point for the regulation of gene expression is at the initial generation of the transcript (8, 46). The eukaryotic cell has developed a wide range of control within which a gene can be activated from a dormant state or repressed from an activated state (59). These states can be temporally fixed by the additional controls imposed by chromatin structure and epigenetic marks upon both histones and DNA (60). Proteins that impose such determinants have been generally segregated into transcriptional activators and repressors (37). However, a recent realization that adds an additional layer of complexity is that such a demarcation of function is oversimplified, as some factors can do double duty as activators and repressors (3,20,21). These activities are controlled by a variety of external stimuli and provide a means by which the surprisingly low number of genes in the mammalian genome can exert multiple effects on genomic targets (33).Erythroid Krüppel-like factor (EKLF/KLF1) controls adult ␤-globin gene expression by the interaction of its three zinc fingers with the CAC element (5ЈCCACACCCT3Ј) located within the proximal promoter (4,40,48). Genetic ablation of EKLF leads to loss of the DNase-hypersensitive site at the ␤-promoter and absence of ␤-globin gene expression, resulting in embryonic lethality due to a profound ␤-thalassemia and toxic accumulation of ␣-globin chains (34,44,50,63). Murine yolk sac (primitive) erythroid cells express embryonic ␤-like globins and appear normal, but the lethality arises at the time of the switch to adult ␤-globin expression, which in the mouse occurs in the definitive erythroid cells of the fetal liver (58). EKLF's ability to interact with p300/CBP (67) and with the SWI/SNF complex (2) suggests a means by which EKLF integrates these components at the ␤-promoter and induces transcription initiation. These interactions are interrelated, as p300/CBP acetylat...

Functional Interactions between Erythroid Kruppel-like Factor (EKLF/KLF1) and Protein Phosphatase PPM1B/PP2Cβ

“…Although the existence of paradoxical roles in Ppm1b in erythroid development seems surprising, there is increasing evidence that EKLF itself plays various roles in erythroid populations at different stages of development (2,4,55). It is highly probable that these roles are due to different protein/protein interactions and post-translational modifications that take place over the course of development.…”

Non-random subcellular distribution of variant EKLF in erythroid cells

“…EKLF also activates protein-stabilizing, heme biosynthetic pathway, and red cell membrane protein genes in both primitive and definitive cells [17][18][19][20]. Although EKLF predominantly serves as a transcriptional activator, protein-protein interactions between EKLF and corepressors, such as mSin3A and HDAC1 [21], can result in the stage-specific repression of EKLF target genes [22]. Recent studies demonstrate that EKLF plays a novel role in negatively regulating megakaryocyte lineage commitment [23].…”

“…EKLF is also posttranslationally modified by phosphorylation [24], acetylation [16], ubiquitylation [25], and sumoylation [26]. Some of these alter protein/protein interactions and thus modify EKLF activity [22,26,27].…”

Structural and functional characterization of an atypical activation domain in erythroid Krüppel-like factor (EKLF)

“…EKLF specifically binds to CAACC elements (1,3) and either the absence of EKLF or mutations in the β-globin CAACC element leads to severe β-thalassemia (5,6,10). In addition, EKLF plays a role in generating an open chromatin structure on the adult β-globin gene through interactions with transcriptional regulatory factors (11)(12)(13)(14)(15)(16).…”