Major erythrocyte membrane protein genes in EKLF-deficient mice.
OBJECTIVE
Mice deficient in the transcription factor erythroid Krüppel-like factor, KLF1 (EKLF) die approximately 14.5 days postcoitum of anemia, attributed to decreased expression of the beta-globin gene. The objectives of this study were to rescue EKLF-deficient embryos with mice expressing gamma-globin from beta-spectrin or ankyrin promoters and to characterize expression of the major erythrocyte membrane genes in EKLF-deficient cells.


METHODS
Transgenic beta-spectrin/gamma-globin or ankyrin/gamma-globin mice were bred onto EKLF-deficient and wild-type backgrounds. Animals were genotyped, gamma-globin mRNA levels measured, and hemoglobin electrophoresis performed. Steady-state mRNA levels and transcriptional rates of the major erythrocyte membrane protein genes were assayed.


RESULTS
beta-spectrin/gamma-globin or ankyrin/gamma-globin mice on EKLF-deficient and wild-type backgrounds had identical levels of gamma-globin mRNA, indicating EKLF-independence of these promoters. gamma-Globin expression improved globin chain imbalance, but hemolysis was not improved and no live-born EKLF-deficient/(A)gamma-globin mice were obtained. Circulating erythroid cells from EKLF-deficient/(A)gamma-globin embryos exhibited hemolysis reminiscent of that seen in patients with severe erythrocyte membrane defects. Levels of beta-spectrin, ankyrin, and band 3 mRNA, but not alpha-spectrin, were decreased in EKLF-deficient fetal liver RNA. In a run-on assay, levels of transcription of the ankyrin and band 3 genes were decreased in EKLF-deficient fetal liver nuclei.


CONCLUSIONS
These results indicate that the EKLF-responsive regions of the ankyrin and beta-spectrin genes are outside their promoters and that EKLF is necessary for full transcriptional activity of the ankyrin and band 3 genes; the results also provide additional evidence that defects in addition to beta-globin deficiency, including an abnormal erythrocyte membrane, contribute to the anemia and embryonic lethality in EKLF-deficient mice.
Nilson
Douglas G
DG
Hematopoiesis Section, Genetics and Molecular Biology Branch, National Human Genome Research Institute, National Institutes of Health, Bethesda, MD, USA.
Sabatino
Denise E
DE
Bodine
David M
DM
Gallagher
Patrick G
PG
D000818:Animals; D002467:Cell Nucleus / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D020964:Embryo Loss / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D004910:Erythrocyte Membrane / Q000235:genetics / Q000378:metabolism*; D004913:Erythrocytes, Abnormal / Q000378:metabolism* / Q000473:pathology; D005786:Gene Expression Regulation / Q000235:genetics*; D005914:Globins / Q000096:biosynthesis / Q000172:deficiency / Q000235:genetics; D006461:Hemolysis / Q000235:genetics; D006801:Humans; D051741:Kruppel-Like Transcription Factors / Q000172:deficiency / Q000378:metabolism*; D008099:Liver / Q000378:metabolism; D008565:Membrane Proteins / Q000096:biosynthesis* / Q000235:genetics; D051379:Mice; D018345:Mice, Knockout; D011401:Promoter Regions, Genetic / Q000235:genetics
D051741:Kruppel-Like Transcription Factors; D008565:Membrane Proteins; C080774:erythroid Kruppel-like factor; D005914:Globins