A Krüppel-like factor 1 (KLF1) Mutation Associated with Severe Congenital Dyserythropoietic Anemia Alters Its DNA-Binding Specificity

Abstract

Krüppel-like factor 1 (KLF1/EKLF) is a transcription factor that globally activates genes involved in erythroid cell development. Various mutations are identified in the human KLF1 gene. The E325K mutation causes congenital dyserythropoietic anemia (CDA) type IV, characterized by severe anemia and non-erythroid-cell-related symptoms. The CDA mutation is in the second zinc finger of KLF1 at a position functionally involved in its interactions with DNA. The molecular parameters of how CDA-KLF1 exerts its biological effects have not been addressed. Here, using an in vitro selection strategy, we determined the preferred DNA-binding site for CDA-KLF1. Binding to the deduced consensus sequence is supported by in vitro gel shifts and by in vivo functional reporter gene studies. Two significant changes compared to wild-type (WT) binding are observed: G is selected as the middle nucleotide, and the 3′ portion of the consensus sequence is more degenerate. As a consequence, CDA-KLF1 did not bind the WT consensus sequence. However, activation of ectopic sites is promoted. Continuous activation of WT target genes occurs if they fortuitously contain the novel CDA site nearby. Our findings provide a molecular understanding of how a single mutation in the KLF1 zinc finger exerts effects on erythroid physiology in CDA type IV.

Transcriptional Control of Gene Expression and the Heterogeneous Cellular Identity of Erythroblastic Island Macrophages

“…One interesting approach would be to examine how EKLF expression in these subpopulations is initiated and regulated. Further, one must determine how EKLF function in macrophages differs from its erythroid counterpart; since EKLF has a conserved DNA binding sequence (5′CCMCRCCCN; ( Tallack et al, 2010 ; Gillinder et al, 2017 ; Kulczynska et al, 2020 )), what prevents EKLF binding to erythroid targets and driving the erythroid gene program in macrophages? One likelihood is that EKLF has separate molecular interactions with transcription cofactors leading to distinct chromatin dynamics at EKLF targets in erythroid cells and macrophages, perhaps determined by its cell- or stage-specific post-translational modification status (discussed in Yien and Bieker (2013) ).…”

Identification and Functional Analysis of Known and New Mutations in the Transcription Factor KLF1 Linked with β-Thalassemia-like Phenotypes

“…Mutations in the KLF1 gene have been reported to interfere with the erythropoiesis process, thus leading to severe hematological disorders, including congenital dyserythropoietic anemia (CDA) and congenital hemolytic anemia [ 18 , 19 , 20 ]. Additionally, KLF1 haploinsufficiency contributes to a number of benign hematological conditions, such as borderline-elevated HbA 2 levels, mild microcytosis, and/or hypochromia [ 21 , 22 ].…”

Congenital dyserythropoietic anemia type IV in the genetic era: A rare neonatal case report of rapid identification with a review of the literature

“… 2–11 Patients are found to have a mutation in the transcription factor Kruppel‐like factor 1 (KLF1), which has been implicated in various mild hematological disorders (see Table S1). 12 This is likely due to the KLF1 role at multiple levels of the erythropoietic process, and varying mutations may lead to different functional consequences on the transcription factor 13–15 . CDAIV is an autosomal dominant condition in which all patients harbor a de novo heterozygous mutation c.973G.A (p.E325K) (see Table 1).…”

“…CDAIV is an autosomal dominant condition in which all patients harbor a de novo heterozygous mutation c.973G.A (p.E325K) (see Table 1). 13–15 Most CDAIV patients present severe anemia, hemolysis, hepatosplenomegaly, hyperbilirubinemia, and persistence of fetal hemoglobin. They are often transiently transfusion‐dependent.…”