A dileucine motif targets the sulfate anion transporter sat-1 to the basolateral membrane in renal cell lines.
The sat-1 transporter mediates sulfate/bicarbonate/oxalate anion exchange in vivo at the basolateral membrane of the kidney proximal tubule. In the present study, we show two renal cell lines [Madin-Darby canine kidney (MDCK) and porcine proximal tubular kidney (LLC-PK1) cells] that similarly target sat-1 exclusively to the basolateral membrane. To identify possible sorting determinants, we generated truncations of the sat-1 cytoplasmic COOH terminus, fused to enhanced green fluorescence protein (EGFP) or the human IL-2 receptor alpha-chain (Tac) protein, and both fusion constructs were transiently transfected into MDCK cells. Confocal microscopy revealed that removal of the last three residues on the sat-1 COOH terminus, a putative PDZ domain, had no effect on basolateral sorting in MDCK cells or on sulfate transport in Xenopus oocytes. Removal of the last 30 residues led to an intracellular expression for the GFP fusion protein and an apical expression for the Tac fusion protein, suggesting that a possible sorting motif lies between the last 3 and 30 residues of the sat-1 COOH terminus. Elimination of a dileucine motif at position 677/678 resulted in the loss of basolateral sorting, suggesting that this motif is required for sat-1 targeting to the basolateral membrane. This posttranslational mechanism may be important for the regulation of sulfate reabsorption and oxalate secretion by sat-1 in the kidney proximal tubule.
Regeer
Ralf R
RR
Department of Physiology and Pharmacology, School of Biomedical Sciences, University of Queensland, St. Lucia, QLD 4072, Australia. d.markovich@uq.edu.au
Markovich
Daniel
D
D000818:Animals; D027321:Anion Transport Proteins; D017920:Antiporters / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism; D016764:Cell Polarity / Q000502:physiology; D015870:Gene Expression; D049452:Green Fluorescent Proteins; D007202:Indicators and Reagents / Q000378:metabolism; D007668:Kidney / Q000166:cytology*; D018374:LLC-PK1 Cells; D007930:Leucine / Q000737:chemistry / Q000378:metabolism*; D008164:Luminescent Proteins / Q000235:genetics; D009865:Oocytes / Q000378:metabolism; D017434:Protein Structure, Tertiary; D021381:Protein Transport / Q000502:physiology; D051381:Rats; D000076988:Sulfate Transporters; D013431:Sulfates / Q000378:metabolism*; D013552:Swine; D014982:Xenopus laevis
D027321:Anion Transport Proteins; D017920:Antiporters; D007202:Indicators and Reagents; D008164:Luminescent Proteins; C056459:Slc26a1 protein, rat; D000076988:Sulfate Transporters; D013431:Sulfates; D049452:Green Fluorescent Proteins; D007930:Leucine