Neomorphic effects of the neonatal anemia (Nan-Eklf) mutation contribute to deficits throughout development

Abstract

Transcription factor control of cell-specific downstream targets can be significantly altered when the controlling factor is mutated. We show that the semi-dominant neonatal anemia (Nan) mutation in the EKLF/ KLF1 transcription factor leads to ectopic expression of proteins that are not normally expressed in the red blood cell, leading to systemic effects that exacerbate the intrinsic anemia in the adult and alter correct development in the early embryo. Even when expressed as a heterozygote, the Nan-EKLF protein accomplishes this by direct binding and aberrant activation of genes encoding secreted factors that exert a negative effect on erythropoiesis and iron use. Our data form the basis for a novel mechanism of physiological deficiency that is relevant to human dyserythropoietic anemia and likely other disease states.

KLF1 E325K-associated Congenital Dyserythropoietic Anemia Type IV: Insights Into the Variable Clinical Severity

“…Together, these alterations lead to numerous changes in erythroid gene expression, altering expression of direct KLF1 target genes and inducing ectopic expression of genes not typically expressed. 20 , 21 KLF1 mutant erythrocytes may have defects in proteins involved in growth and differentiation, maintenance of membrane integrity, hemoglobins and heme biosynthesis, iron homoeostasis, and regulation of metabolism. 22 …”

Mutant KLF1 in Adult Anemic Nan Mice Leads to Profound Transcriptome Changes and Disordered Erythropoiesis

“…ChIPseq confirmed ectopic Nan-KLF1 binding to an altered consensus sequence, CCM- NG C-CCN, with the result that >60% of Nan-KLF1 occupied sites do not overlap wild type (WT) KLF1 sites. Ectopic binding contributes to anemia in Nan through extrinsic mechanisms 15 . For example, hepcidin ( Hamp ) and interferon regulatory factor 7 ( Irf7 ), normally expressed in adult liver 16 and macrophages 17 , respectively, show dramatically increased expression in Nan fetal liver 13 and adult WT spleen and bone marrow 15 leading to increased serum hepcidin and interferon beta.…”

“…Ectopic binding contributes to anemia in Nan through extrinsic mechanisms 15 . For example, hepcidin ( Hamp ) and interferon regulatory factor 7 ( Irf7 ), normally expressed in adult liver 16 and macrophages 17 , respectively, show dramatically increased expression in Nan fetal liver 13 and adult WT spleen and bone marrow 15 leading to increased serum hepcidin and interferon beta. lncreased hepcidin with markedly decreased erythroferrone, which is not bound by Nan-KLF1 15 , limits iron availability 18 , 19 .…”

“…For example, a different substitution (E325K) at the position corresponding to Nan causes congenital dyserythropoietic anemia (CDA) type IV 22 . Prior transcriptome studies in Nan focused on fetal liver erythroid cells 1 , 13 , 15 . Here, we performed RNAseq in adult spleen erythroid precursors, comparing Nan to littermates made anemic by phlebotomy (WT-PHB).…”

Survey and evaluation of mutations in the human KLF1 transcription unit

“…The mutation alters the DNA binding specificity of EKLF such that it no longer binds promoters of a subset of its DNA targets 29 . In addition, recognition of a novel target DNA sequence by Nan-EKLF leads to ectopic expression of genes not normally expressed in the red cell, yielding a neomorphic phenotype with cellular and systemic consequences 32 , 33 .…”