Studies of mouse models of anemia have long provided fundamental insights into red blood cell formation and function. Here we show that the semidominant mouse mutation Nan (“neonatal anemia”) carries a single amino acid change (E339D) within the second zinc finger of the erythroid Krüppel-like factor (EKLF), a critical erythroid regulatory transcription factor. The mutation alters the DNA-binding specificity of EKLF so that it no longer binds promoters of a subset of its DNA targets. Remarkably, even when mutant Nan and wild-type EKLF alleles are expressed at equivalent levels, the mutant form selectively interferes with expression of EKLF target genes whose promoter elements it no longer binds. This interference yields a distorted genetic output and selective protein deficiencies that differ from those seen in EKLF-heterozygous and EKLF-null red blood cells and presents a unique and unexpected mechanism of inherited disease.

Defects in a triad of organelles (melanosomes, platelet granules, and lysosomes) result in albinism, prolonged bleeding, and lysosome abnormalities in Hermansky-Pudlak syndrome (HPS). Defects in HPS1, a protein of unknown function, and in components of the AP-3 complex cause some, but not all, cases of HPS in humans. There have been 15 inherited models of HPS described in the mouse, underscoring its marked genetic heterogeneity. Here we characterize a new spontaneous mutation in the mouse, cappuccino (cno), that maps to mouse chromosome 5 in a region conserved with human 4p15-p16. Melanosomes ofcno/cno mice are immature and dramatically decreased in number in the eye and skin, resulting in severe oculocutaneous albinism. Platelet dense body contents (adenosine triphosphate, serotonin) are markedly deficient, leading to defective aggregation and prolonged bleeding. Lysosomal enzyme concentrations are significantly elevated in the kidney and liver. Genetic, immunofluorescence microscopy, and lysosomal protein trafficking studies indicate that the AP-3 complex is intact in cno/cno mice. It was concluded that the cappuccino gene encodes a product involved in an AP-3–independent mechanism critical to the biogenesis of lysosome-related organelles.

IntroductionHereditary spherocytosis (HS) is the most common inherited hemolytic anemia in people of Northern European descent, occurring with a frequency of approximately 1 in 2000. 1 HS is caused by defects in the red blood cell (RBC) membrane skeleton, a multiprotein structure located just beneath the lipid bilayer that imparts mechanical strength and elasticity to the RBC membrane. The major component of the membrane skeleton, spectrin, is present as tetramers of ␣-and ␤-subunits cross-linked into a 2-dimensional array by short actin filaments. [1][2][3] There are 2 major interactions between membrane skeleton components and integral membrane proteins that attach the spectrin array to the plasma membrane: (1) band 3-spectrin-ankyrin-protein 4.2 and (2) protein 4.1-p55-glycophorin C linkages. [4][5][6][7][8] The band 3-spectrin-ankyrin-protein 4.2 interactions are critical in the pathogenesis of HS. Defects in all of these proteins cause HS in humans and in mice. 1,9,10 Recent estimates are that approximately 50% of the mutations causing HS in European populations are in ankyrin, 20% in band 3, 20% in ␤-spectrin, 5% in protein 4.2, and less than 5% in ␣-spectrin. 1 Depending on the exact genetic defect, the hematologic phenotype varies from asymptomatic to life-threatening hemolysis requiring transfusion therapy and/or splenectomy. Secondary complications of HS include jaundice, gallstones, aplastic crises, and, more rarely, extramedullary hematopoietic masses and leg ulcers. 1 In the ankyrin-deficient nb mouse, progressive ataxia due to Purkinje cell degeneration occurs. 11 In other mouse models of HS, thrombosis and infarction are significant complicating factors. 12 Many specific molecular defects that cause HS in humans have been described. 1 Notably, as in other heritable syndromes, the clinical presentation varies significantly even among individuals with identical gene defects, illustrating the profound effects of genetic background on disease severity. 10 Identifying genetic modifiers by linkage analysis in humans is often difficult, if not impossible, due to environmental influences, genetic diversity, and small population (linkage group) size. Inbred mice, however, provide powerful tools for complex trait analysis. Moreover, as concordance between quantitative trait loci (QTL) in the mouse and human has been demonstrated for several diseases, complex trait analysis in the mouse has significant biomedical relevance. 13,14 Here, we describe a new spontaneous HS mutation in the mouse, wan, which arose on the C3H/HeJ (C3H) inbred strain, and identify QTL that modify the severity of the HS phenotype. Homozygous wan mice are severely anemic. A premature stop codon in the gene encoding erythroid band 3, Slc4a1 (solute carrier family 4 [anion exchanger], member 1; formerly Ae1, anion exchanger 1), results in complete deficiency of band 3 in homozygotes. In hybrid wan/wan newborns derived from F2 intercrosses between C3H wan/ϩ and Mus musculus castaneus (CAST/Ei), marked Reprints: Luanne L. Peters, The Jacks...

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Defects in a triad of organelles (melanosomes, platelet granules, and lysosomes) result in albinism, prolonged bleeding, and lysosome abnormalities in Hermansky-Pudlak syndrome (HPS). Defects in HPS1, a protein of unknown function, and in components of the AP-3 complex cause some, but not all, cases of HPS in humans. There have been 15 inherited models of HPS described in the mouse, underscoring its marked genetic heterogeneity. Here we characterize a new spontaneous mutation in the mouse, cappuccino (cno), that maps to mouse chromosome 5 in a region conserved with human 4p15-p16. Melanosomes ofcno/cno mice are immature and dramatically decreased in number in the eye and skin, resulting in severe oculocutaneous albinism. Platelet dense body contents (adenosine triphosphate, serotonin) are markedly deficient, leading to defective aggregation and prolonged bleeding. Lysosomal enzyme concentrations are significantly elevated in the kidney and liver. Genetic, immunofluorescence microscopy, and lysosomal protein trafficking studies indicate that the AP-3 complex is intact in cno/cno mice. It was concluded that the cappuccino gene encodes a product involved in an AP-3–independent mechanism critical to the biogenesis of lysosome-related organelles.