Rodent intestinal folate transporters (SLC46A1): secondary structure, functional properties, and response to dietary folate restriction.
This laboratory recently identified a human gene that encodes a novel folate transporter [Homo sapiens proton-coupled folate transporter (HsPCFT); SLC46A1] required for intestinal folate absorption. This study focused on mouse (Mus musculus) PCFT (MmPCFT) and rat (Rattus norvegicus) PCFT (RnPCFT) and addresses their secondary structure, specificity, tissue expression, and regulation by dietary folates. Both rodent PCFT proteins traffic to the cell membrane with the NH(2)- and COOH-termini accessible to antibodies targeted to these domains only in permeabilized HeLa cells. This, together with computer-based topological analyses, is consistent with a model in which rodent PCFT proteins likely contain 12 transmembrane domains. Transport of [(3)H]folates was optimal at pH 5.5 and decreased with increasing pH due to an increase in K(m) and a decrease in V(max). At pH 7.0, folic acid and methotrexate influx was negligible, but there was residual (6S)5-methyltetrahydrofolate transport. Uptake of folates in PCFT-injected Xenopus oocytes was electrogenic and pH dependent. Folic acid influx K(m) values of MmPCFT and RnPCFT, assessed electrophysiologically, were 0.7 and 0.3 microM at pH 5.5 and 1.1 and 0.8 microM at pH 6.5, respectively. Rodent PCFTs were highly specific for monoglutamyl but not polyglutamyl methotrexate. MmPCFT mRNA was highly expressed in the duodenum, proximal jejunum, liver, and kidney with lesser expression in the brain and other tissues. MmPCFT protein was localized to the apical brush-border membrane of the duodenum and proximal jejunum. MmPCFT mRNA levels increased approximately 13-fold in the proximal small intestine in mice fed a folate-deficient vesus folate-replete diet, consistent with the critical role that PCFT plays in intestinal folate absorption.
Qiu
Andong
A
Department of Medicine, Albert Einstein College of Medicine, Bronx, New York 10461, USA.
Min
Sang Hee
SH
Jansen
Michaela
M
Malhotra
Usha
U
Tsai
Eugenia
E
Cabelof
Diane C
DC
Matherly
Larry H
LH
Zhao
Rongbao
R
Akabas
Myles H
MH
Goldman
I David
ID
D000595:Amino Acid Sequence; D000818:Animals; D027321:Anion Transport Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism*; D004195:Disease Models, Animal; D005492:Folic Acid / Q000378:metabolism*; D005493:Folic Acid Antagonists / Q000378:metabolism*; D005494:Folic Acid Deficiency / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007421:Intestine, Small / Q000378:metabolism*; D007700:Kinetics; D008297:Male; D008564:Membrane Potentials; D026901:Membrane Transport Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008727:Methotrexate / Q000031:analogs & derivatives / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D008871:Microvilli / Q000378:metabolism; D008969:Molecular Sequence Data; D009865:Oocytes; D011099:Polyglutamic Acid / Q000031:analogs & derivatives / Q000378:metabolism; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D058979:Proton-Coupled Folate Transporter; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D013763:Tetrahydrofolates / Q000378:metabolism; D014981:Xenopus
D027321:Anion Transport Proteins; D005493:Folic Acid Antagonists; D026901:Membrane Transport Proteins; D058979:Proton-Coupled Folate Transporter; D012333:RNA, Messenger; C516877:Slc46a1 protein, mouse; C525126:Slc46a1 protein, rat; D013763:Tetrahydrofolates; D011099:Polyglutamic Acid; C014085:methotrexate polyglutamate; D005492:Folic Acid; C005984:5-methyltetrahydrofolate; D008727:Methotrexate