Molecular mechanism of the intestinal biotin transport process.
Previous studies have characterized different aspects of the cellular/membrane mechanism and regulation of the intestinal uptake process of the water-soluble vitamin biotin. Little, however, is known about the molecular mechanisms of the uptake process. In this study, we have identified a cDNA from rat small intestine that appears to be involved in biotin transport. The open reading frame of this cloned cDNA consisted of 1,905 bases and was identical to that identified for the vitamin transporter in placental tissue. Significant heterogeneity, however, was found in the 5' untranslated region of this clone, with three distinct variants (II, III, IV) being identified in the small intestine; the placental variant (variant I), however, was not present in the small gut. Variant II was found to be the predominant form expressed in the rat small and large intestines. Functional identity of the cloned intestinal cDNA was confirmed by stable expression in COS-7 cells, which showed a four- to fivefold increase in biotin uptake in transfected COS-7 cells compared with controls. The induced biotin uptake in transfected COS-7 cells was found to be 1) Na(+) dependent, 2) saturable as a function of concentration with an apparent K(m) of 8. 77 microM and a V(max) of 779.7 pmol. mg protein(-1). 3 min(-1), and 3) inhibited by unlabeled biotin and pantothenic acid and their structural analogs. The distribution of complementary mRNA transcripts of the cloned cDNA along the vertical and longitudinal axes of the intestinal tract was also determined. Results of this study describe the molecular characteristics of the intestinal biotin absorption process and report the identification of a cDNA that encodes a Na(+)-dependent biotin uptake carrier that appears to exist in the form of multiple variants.
Chatterjee
N S
NS
Medical Research Service, Veterans Affairs Medical Center, Long Beach 90822, USA.
Kumar
C K
CK
Ortiz
A
A
Rubin
S A
SA
Said
H M
HM
D020121:5' Untranslated Regions / Q000235:genetics; D000818:Animals; D001483:Base Sequence; D019556:COS Cells; D002352:Carrier Proteins / Q000235:genetics*; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics / Q000378:metabolism; D014644:Genetic Variation; D007413:Intestinal Mucosa / Q000378:metabolism*; D008565:Membrane Proteins / Q000235:genetics*; D008969:Molecular Sequence Data; D016366:Open Reading Frames; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D020133:Reverse Transcriptase Polymerase Chain Reaction; D027981:Symporters; D014018:Tissue Distribution; D014162:Transfection
D020121:5' Untranslated Regions; D002352:Carrier Proteins; D018076:DNA, Complementary; D008565:Membrane Proteins; D012333:RNA, Messenger; D027981:Symporters; C119981:biotin transporter