Document Detail


Modulation of function of sodium-dependent vitamin C transporter 1 (SVCT1) by Rab8a in intestinal epithelial cells: studies utilizing Caco-2 cells and Rab8a knockout mice.
MedLine Citation:
PMID:  23014846     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
BACKGROUND: Ascorbic acid (AA) is required for normal human health and development. Human intestine expresses two sodium-dependent vitamin C transporters (hSVCT-1 and -2) that mediate cellular AA transport, with hSVCT1 targeting to the apical membrane of polarized epithelia. Studies have shown a role for the Rab8a in the apical membrane targeting of transporters in intestinal cells.
AIMS: The purpose of this study was to determine whether Rab8a impacts the function and/or targeting of hSVCT1, and intestinal AA uptake.
METHODS: We used human intestinal cells and cells from a Rab8a knockout mouse. (14)C-AA uptake was performed to determine functionality. PCR and western blotting were performed to determine RNA and protein expression, respectively. Confocal imaging was performed to determine co-localization.
RESULTS: We show that hSVCT1 co-localized with Rab8a in intestinal cells. Knockdown of Rab8a lead to a significant inhibition in AA uptake and cell surface biotinylation studies revealed a lower cell surface expression of hSVCT1 in Rab8a siRNA-treated cells. Similarly, in the small intestine of a Rab8a knockout mouse, AA uptake was significantly inhibited. This effect again resulted from a decreased expression level of mSVCT1 protein, even though mRNA expression of SVCT1 was similar in intestinal cells from Rab8a knockout and wild-type litter-mates. The latter data are suggestive of enhanced lysosomal degradation of hSVCT1 protein in Rab8a-deficient cells; indeed, confocal imaging of Rab8a siRNA-treated intestinal cells revealed a strong overlap between hSVCT1-YFP and LAMP1-RFP.
CONCLUSIONS: These findings show a role for Rab8a in the physiological function of hSVCT1 in intestinal epithelia.
Authors:
Veedamali S Subramanian; Sandeep B Subramanya; Abhisek Ghosal; Jonathan S Marchant; Akihiro Harada; Hamid M Said
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Publication Detail:
Type:  Journal Article; Research Support, N.I.H., Extramural; Research Support, U.S. Gov't, Non-P.H.S.     Date:  2012-09-27
Journal Detail:
Title:  Digestive diseases and sciences     Volume:  58     ISSN:  1573-2568     ISO Abbreviation:  Dig. Dis. Sci.     Publication Date:  2013 Mar 
Date Detail:
Created Date:  2013-04-04     Completed Date:  2013-06-03     Revised Date:  2014-03-09    
Medline Journal Info:
Nlm Unique ID:  7902782     Medline TA:  Dig Dis Sci     Country:  United States    
Other Details:
Languages:  eng     Pagination:  641-9     Citation Subset:  AIM; IM    
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MeSH Terms
Descriptor/Qualifier:
Amino Acids / metabolism
Animals
Ascorbic Acid / metabolism*
Blotting, Western
Caco-2 Cells
Gene Knockdown Techniques
Gene Silencing
Humans
Intestines / metabolism*
Mice
Mice, Knockout
Protein Transport
RNA, Small Interfering
Real-Time Polymerase Chain Reaction
Sodium-Coupled Vitamin C Transporters / genetics,  metabolism*
rab GTP-Binding Proteins / genetics,  metabolism*
Grant Support
ID/Acronym/Agency:
DK 56061/DK/NIDDK NIH HHS; DK84094/DK/NIDDK NIH HHS; GM088790/GM/NIGMS NIH HHS; R01 DK058057/DK/NIDDK NIH HHS; R03 DK084094/DK/NIDDK NIH HHS; R37 DK056061/DK/NIDDK NIH HHS
Chemical
Reg. No./Substance:
0/Amino Acids; 0/RNA, Small Interfering; 0/Rab8a protein, mouse; 0/SLC23A1 protein, human; 0/Slc23a1 protein, mouse; 0/Sodium-Coupled Vitamin C Transporters; EC 3.6.1.-/rab GTP-Binding Proteins; EC 3.6.1.-./RAB8A protein, human; PQ6CK8PD0R/Ascorbic Acid
Comments/Corrections

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