IntroductionAdult-onset gastrointestinal malabsorption of the essential micronutrient, cobalamin (vitamin B 12 ), leads to potentially lethal manifestations as diverse as megaloblastic anemia and neutropenia, degeneration of spinal cord nerve tracts, and dementia. 1 In the very young, signs of cobalamin malabsorption may also include growth retardation or loss of developmental milestones or both. ImerslundGräsbeck syndrome (I-GS; also called megaloblastic anemia 1, OMIM no. 261100) is an autosomal recessive disorder characterized by selective malabsorption of cobalamin in the intestine and, most often, of specific low-molecular-weight proteins in renal proximal tubules. 2,3 Patients with I-GS typically present at 2 to 4 years of age with signs of cobalamin deficiency and proteinuria. More than 200 human cases and familial clusters have been reported in Finland, Norway, the Middle East, and Northern Africa. 1 Various null and missense mutations of the cubilin (CUBN) 4 and amnionless (AMN) genes 5 have been implicated in I-GS kindreds, but both loci have been excluded in some. 6 In addition, a number of patients suffering from gastric intrinsic factor (IF) deficiency have been misdiagnosed as having I-GS. 7 Cubilin is a 460-kDa multiligand receptor protein that mediates endocytosis of the IF-cobalamin complex from distal small intestinal chyme and of several proteins from glomerular filtrate in renal proximal tubules. 8 AMN is an approximately 48-kDa apical membrane protein also expressed in intestinal and proximal tubule epithelia but whose function is poorly defined. 5,9 Recent studies suggested that cubilin and AMN function as subunits of a receptor complex, now called "cubam." 10 However, no studies have directly assessed the effect of CUBN or AMN mutations on cubam expression in patients with I-GS because these proteins are expressed in inaccessible tissues, and the clinical disease is easily treated. Unfortunately, the AMN knockout mouse exhibits an embryonic lethal phenotype. 9 Canine I-GS was derived originally from purebred giant schnauzers (GSs) as a naturally occurring animal model of the human disorder [11][12][13] and has contributed significantly to understanding of its molecular biologic basis as well as aspects of cubilin function. 14-17 CUBN was excluded from the GS disease locus, 18 and the disorder was recently mapped to an approximately 4-Mb interval predicted to harbor AMN. 19 To define the molecular basis of the canine I-GS model, we sought AMN mutations segregating in the GS kindred and an unrelated kindred of Australian shepherd (AS) dogs and investigated in vivo AMN and cubilin expression in affected dogs. Additionally, comparison of these results to expression of mutant canine AMN in a heterologous mammalian-cell transfection system validates the cell-culture system for ex vivo Materials and methods AnimalsDogs were handled according to principles and protocols approved by the MSU All University Committee for Animal Use and Care. MSU University Laboratory Animal Resources housed dogs of...

Selective malabsorption of cobalamin (vitamin B(12)) accompanied by proteinuria, known as Imerslund-Gräsbeck syndrome or megaloblastic anemia 1 (I-GS, MGA1; OMIM 261100), is a rare autosomal recessive disorder. In Finnish kindreds, I-GS is caused by mutations in the cubilin gene ( CUBN), located on human Chromosome (Chr) 10. However, not all patients have CUBN mutations, and three distinct mutations in the amnionless gene, AMN, were very recently identified in patients from Norwegian and Israeli families. The present study demonstrates that in a large canine I-GS pedigree, the disease is genetically linked (peak multipoint LOD score 11.74) to a region on dog Chr 8 that exhibits conserved synteny with human Chr 14q. Multipoint analysis indicates that the canine disease gene lies in an interval between the echinoderm microtubule-associated, protein-like 1 ( EML1) gene and the telomere. A single critical recombinant further suggests that the disease gene is between markers in EML1 and the G protein-coupled receptor ( G2A) gene, defining an I-GS interval in the human genome that contains the AMN gene. Thus, these comparative-mapping data provide evidence that canine I-GS is a homologue of one form of the human disease and will provide a useful system for understanding the molecular mechanisms underlying the disease in humans.