Characterization of sulphate transporters in isolated bovine articular chondrocytes.
Uptake of SO(4) (2-) by articular chondrocytes is an essential step in the pathway for sulphation of glycosaminoglycans (GAGs), with mutations in SO(4) (2-) transport proteins resulting in abnormalities of skeletal growth. In the present study, the transporters mediating SO(4) (2-) transport in bovine articular chondrocytes have been characterized. Expression of candidate transporters was determined using RT-PCR, while SO(4) (2-) transport was measured in radioisotope flux experiments. RT-PCR experiments showed that bovine articular chondrocytes express three transporters known to transport SO(4) (2-): AE2 (SLC4a2), DTDST (SLC26a2), and SLC26a11. Other transporters--NaS-1 (SLC13a1), SAT-1 (SLC26a1), DRA (SLC26a3), SLC26a6 (PAT1), SLC26a7, SLC26a8 (Tat-1), and SLC26a9--were, however, not detected. In functional experiments, SO(4) (2-) uptake was temperature-sensitive, inhibited by 60% by DIDS (50 microM) and exhibited saturation kinetics, with a K(m) value of 16 mM. Uptake was also inhibited at alkaline extracellular pH. In further experiments, a K(i) value for DIDS inhibition of SO(4) (2-) efflux of 5 microM was recorded. A DIDS-sensitive component of SO(4) (2-) efflux persisted in solutions lacking Cl(-) ions. These data are interpreted as evidence for the preferential operation of carrier-mediated exchange of SO(4) (2-) for Cl(-), while an alternative SO(4) (2-)-OH(-) exchange mode is also possible.
Meredith
David
D
Department of Physiology, Anatomy and Genetics, Sherrington Building, Parks Road, Oxford, OX1 3PT, UK.
Gehl
Katharina A
KA
Seymour
John
J
Ellory
J Clive
JC
Wilkins
Robert J
RJ
D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid / Q000494:pharmacology; D000818:Animals; D027321:Anion Transport Proteins / Q000235:genetics / Q000378:metabolism*; D017920:Antiporters / Q000235:genetics / Q000378:metabolism; D001692:Biological Transport; D002358:Cartilage, Articular / Q000378:metabolism*; D002417:Cattle; D019902:Chondrocytes / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D015870:Gene Expression; D012333:RNA, Messenger / Q000378:metabolism; D064606:SLC4A Proteins; D013431:Sulfates / Q000378:metabolism*
D027321:Anion Transport Proteins; D017920:Antiporters; D012333:RNA, Messenger; D064606:SLC4A Proteins; D013431:Sulfates; D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid