Identification of a putative lysosomal cobalamin exporter altered in the cblF defect of vitamin B12 metabolism

Abstract

Vitamin B(12) (cobalamin) is essential in animals for metabolism of branched chain amino acids and odd chain fatty acids, and for remethylation of homocysteine to methionine. In the cblF inborn error of vitamin B(12) metabolism, free vitamin accumulates in lysosomes, thus hindering its conversion to cofactors. Using homozygosity mapping in 12 unrelated cblF individuals and microcell-mediated chromosome transfer, we identified a candidate gene on chromosome 6q13, LMBRD1, encoding LMBD1, a lysosomal membrane protein with homology to lipocalin membrane receptor LIMR. We identified five different frameshift mutations in LMBRD1 resulting in loss of LMBD1 function, with 18 of the 24 disease chromosomes carrying the same mutation embedded in a common 1.34-Mb haplotype. Transfection of fibroblasts of individuals with cblF with wild-type LMBD1 rescued cobalamin coenzyme synthesis and function. This work identifies LMBRD1 as the gene underlying the cblF defect of cobalamin metabolism and suggests that LMBD1 is a lysosomal membrane exporter for cobalamin.

A Patient With an Inborn Error of Vitamin B12 Metabolism (cblF) Detected by Newborn Screening

“…To date there are 15 reported cblF patients. [3][4][5] Several have had decreased serum vitamin B 12 levels, apparently reflecting a role for the lysosome in intestinal uptake of ingested cobalamin; the Schilling test for cobalamin absorption has been reported to be abnormal in cblF patients. 6 A second disorder with an identical biochemical and cellular phenotype, cblJ, caused by mutations in the gene which codes for a different lysosomal membrane protein, ABCD4, has recently been described in 3 patients.…”

“…3 Endocytosis of transcobalamin-bound circulating cobalamin is mediated by the transcobalamin receptor, with dissociation of the transcobalamin-cobalamin complex and proteolysis of transcobalamin occurring in the lysosome. LMBD1 is thought to play a role in transporting cobalamin from the lysosome into the cytoplasm after its release from transcobalamin.…”

Diagnostic Exome Sequencing and Tailored Bioinformatics of the Parents of a Deceased Child with Cobalamin Deficiency Suggests Digenic Inheritance of the MTR and LMBRD1 Genes

“…c.3518C>T (p.P1173L) is the most common alteration observed in the MTR gene, with a frequency of about 40% (16/38 chromosomes) of patients with cblG deficiency (Watkins et al 2002). LMBRD1 c.1056delG (p.L352Lfs*18) is the most frequently reported alteration, found to be present in 75% (18/24 chromosomes) of one cohort of 12 patients with cblF deficiency (Rutsch et al 2009(Rutsch et al , 2011.…”

“…Other clinical findings in patients with cblG include decreased S-adenosylmethionine, lethargy, feeding difficulties, vomiting, abnormal tonus, mental retardation, failure to thrive, blindness, ataxia, delayed myelination, and megaloblastic anemia (Sarafoglou and Hoffman 2009). LMBRD1 is a lysosomal membrane protein thought to be involved in lysosomal export of cobalamin in which alteration leads to the cblF complementation class of the cobalamin metabolism disorders (Rutsch et al 2009(Rutsch et al , 2011Gailus et al 2010). Cells from cblF patients accumulate large amounts of free cobalamin within the lysosomes, but there is a deficiency of both cobalamin coenzyme derivatives and decreased activity of MMA mutase and methionine sythetase (reviewed in Watkins and Rosenblatt, 2011).…”

“…In fact, a recent review on digenic inheritance cautions medical geneticists to consider digenic inheritance as a possible mechanism for patients with novel phenotypes (Schaffer 2013). Further, the clinical features observed among patients with LMBRD1 are widely variable (Froese and Gravel 2010;Watkins and Rosenblatt 2011), and specifically patients with the c.1056delG alteration in the compound heterozygous or homozygous state show considerable phenotypic variability ranging from death in infancy to asymptomatic long-term survival (Rutsch et al 2009). The unaffected mother, unaffected father, and unaffected sister do not carry both the MTR and LMBRD1 alterations which would have ruled out the possibility for a digenic affect.…”