Transcobalamin II synthesized in the intestinal villi facilitates transfer of cobalamin to the portal blood (original) (raw)
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Journal of Biological Chemistry
The cellular uptake of cobalamin (Cbl, vitamin BIZ) is mediated by transcobalamin I1 (TCII), a plasma protein that binds Cbl and is secreted by human umbilical vein endothelial (HUVE) cells. These cells synthesize and secrete TCII and, therefore, served as the source of the complementary DNA (cDNA) library from which the TCII cDNA was isolated. This full-length cDNA consists of 1866 nucleotides that code for a leader peptide of 18 amino acids, a secreted protein of 409 amino acids, a 5"untranslated segment of 37 nucleotides, and a 3'-untranslated region of 548 nucleotides. A single 1.9-kilobase species of mRNA corresponding to the size of the cDNA was identified by Northern blot analysis of the RNA isolated from HUVE cells.
Rat transcobalamin: cloning and regulation of mRNA expression
The Journal of Physiology, 2004
Transcobalamin (TC) has been cloned and used for studying its gene expression in the rat. TC mRNA is distributed widely in adult rat tissues, but at different levels (kidney > liver > lung > yolk sac > intestine > heart > brain > spleen > muscle). TC mRNA levels were 4-fold higher in the jejunum and ileum compared to its levels in the duodenum. During postnatal development, TC mRNA levels in the ileum declined 4-fold from day 4 to day 12, but increased by 5-fold between days 12 and 24. In contrast, TC mRNA levels increased by 2.5-fold in the kidney from day 4 to day 12 and then declined by 2-fold by day 24. Adrenalectomy of adult rats resulted in a 4-fold decline in ileal levels of TC mRNA and a 50% decline in the ileal mucosal formation of the TC-[ 57 Co] cobalamin (Cbl) complex following oral administration of [ 57 Co]Cbl complexed to gastric intrinsic factor (IF). Cortisone treatment reversed these changes noted in the ileum. In contrast to ileum, kidney TC mRNA levels were not altered significantly in adrenalectomized rats before and after cortisone treatment. Taken together, this study has provided evidence for the regulation of TC gene expression in the rat kidney and intestine during their postnatal development, and cortisone selectively regulates ileal but not kidney TC mRNA levels.
Intrinsic Factor-mediated Absorption of Cobalamin by Guinea Pig Ileal Cells
Journal of Clinical Investigation, 1983
A B S T R A C T To investigate the fate of intrinsic factor and cobalamin during cobalamin absorption, we incubated enterocytes isolated from guinea pig ileum for periods of up to 30 min with 57Co-labeled cyanocobalamin bound either to human intrinsic factor or to rabbit intrinsic factor biosynthetically labeled with [35S]methionine. When the labeled complex was incubated for 30 min with isolated ileal cells under conditions that block cellular metabolism, virtually all cellular radioactivity could be removed by washing the cell surface with EDTA or acid. In contrast, washing removed only half the radioactivity from cells incubated at 370C in 02 When residual cellular radioactivity was extracted and analyzed by gel filtration, 80-94% of both the 35S and 57Co radioactivity eluted in the same fractions as the original complex. The remaining 6-20% eluted as free [57Co]cobalamin or [35S]methionine. To examine events occurring after 30 min, we instilled into tied-off ileal loops of intact guinea pigs radiolabeled intrinsic factor-cobalamin complex and extracted nondissociable radioactivity 2-4.5 h later. The proportion of extracted 57Co eluting as free cobalamin increased to 39-46%, that eluting as intrinsic factor-cobalamin complex declined to 22-45%, and 9-34% now eluted as a macromolecule that reacted with antitranscobalamin II antibody but not antiintrinsic factor antibody. Extracted 35S radioactivity eluted in several peaks in addition to the intrinsic factor peak. These findings suggest that (a) after reversible attachment of intrinsic factor-cobalamin complex to its ileal surface receptor, an energy-dependent process prevents removal of the complex from the cell Parts of this research were reported at the Annual Meetings of the American
Mapping the functional domains of human transcobalamin using monoclonal antibodies
FEBS Journal, 2005
Vitamin B 12 (cobalamin, Cbl) is absorbed in the distal ileum with the help of a specific binding protein intrinsic factor (IF) and appears in the circulation bound to another carrier transcobalamin (TC) . Tissue uptake of the TCAECbl complex (holo-TC) is mediated by specific receptors on the surface of the plasma membrane [2]. Holo-TC represents Cbl available for cellular uptake and a decrease in its level would indicate reduced absorption of the vitamin as well as systemic Cbl deficiency. Two new methods have recently been described for the measurement of holo-TC in plasma samples . Both methods employ TC-specific antibodies to capture the protein from plasma but lack the specificity needed for direct measurement of holo-TC in serum. The antigenic determinants and the functional domains of TC have not been identified.
Cobalamin-mediated regulation of transcobalamin receptor levels in rat organs
Archives of Biochemistry and Biophysics, 2007
Total gastrectomy (TG) causes cobalamin (Cbl) deficiency followed by increases in tumor necrosis factor (TNF)-a levels in the spinal cord (SC) of the rat. In order to understand how Cbl deficiency may influence cell Cbl transport, we have measured by immunoblotting protein levels of the receptor for the Cbl-transcobalamin (TC) complex (TC-R) in both animal and cell models. TC-R protein levels were elevated in the total membranes of duodenal mucosa, kidneys, liver, and SC of rats made Cbl-deficient (Cbl-D) by means of TG or feeding with a Cbl-D diet. Postoperative Cbl-replacement treatment normalized the TC-R protein levels in each of the tested organs, regardless of whether this treatment was given during the first two post-TG or during the third and fourth post-TG mo. In Caco-2 cells, progressively increasing TNF-a concentrations supplemented to culture medium induced an up-regulation of TC-R protein levels. We provide the first evidence of the regulation of a Cbl-specific receptor by the vitamin itself in some rat organs.
The Journal of cell biology, 1984
Absorption of cobalamin is facilitated by the binding of the intrinsic factor-cobalamin complex (IF-cbl) to specific receptors in the ileum. The physical and biochemical characteristics of this ligand-receptor binding reaction have been extensively studied, but little is known about the cellular mechanisms or receptor synthesis, intracellular transport, and expression on the microvillus surface membrane. We attempted to delineate these mechanisms by using ultrastructural immunocytochemistry to localize the IF-cbl receptor in the crypt, mid-villus, and villus tip regions of mucosal biopsies obtained from the ileum of anesthetized dogs. Prior to initiating the ileal localization studies, the antisera to purified canine IF-cbl receptor that was employed in our studies was shown to have specificity for site (e.g., ileal enterocytes vs. other cells within the gastrointestinal tract) and immunohistochemical specificity. Receptor synthesis in endoplasmic reticulum begins in crypt enterocyt...
Inherited Selective Intestinal Cobalamin Malabsorption and Cobalamin Deficiency in Dogs
Pediatric Research - PEDIAT RES, 1991
Inherited selective intestinal malabsorption of cobalamin (Cbl) was observed in a family of giant schnauzer dogs. Family studies and breeding experiments demonstrated simple autosomal recessive inheritance of this disease. Affected puppies exhibited chronic inappetence and failure to thrive beginning between 6 and 12 wk of age. Neutropenia with hypersegmentation, anemia with anisocytosis and poikilocytosis, and megaloblastic changes of the bone marrow were present. Serum Cbl concentrations were low, and methylmalonic aciduria and homocysteinemia were present. Parenteral, but not oral, cyanocobalamin administration rapidly eliminated all signs of Cbl deficiency except for low serum Cbl concentrations. Cbl malabsorption in affected dogs was documented by oral administration of [57Co]cyanocobalamin with or without simultaneous oral administration of intrinsic factor or normal dog gastric juice. Quantitation and function studies of intrinsic factor and transcobalamin-I1 from affected dogs revealed no abnormality. Other gastrointestinal functions and ileal morphology were normal, indicating a selective defect of Cbl absorption at the level of the ileal enterocyte. Immunoelectron microscopy of ileal biopsies showed that the receptor for intrinsic factor-Cbl complex was absent from the apical brush border microvillus pits of affected dogs. This canine disorder resembles inherited selective intestinal Cbl malabsorption (Imerslund-Grasbeck syndrome) in humans, and is a spontaneously occurring animal model of early onset Cbl deficiency. (Pediatr Res 29: 24-31, 1991) Abbreviations Cbl, cobalamin IF, intrinsic factor IF-Cbl, intrinsic factor-cobalamin complex TC-11, transcobalamin-I1 CN-Cbl, cyanocobalamin [57ColCN-Cbl, radioisotopically labeled cyanocobalamin UCBC, unsaturated cobalamin binding capacity MMA, methylmalonic acid THCys, total homocysteine