Cleavage of disulfide bonds leads to inactivation and degradation of the type IIa, but not type IIb sodium phosphate cotransporter expressed in Xenopus laevis oocytes.
Tris(2-carboxyethyl)phosphine (TCEP) reduces (cleaves) disulfide bonds of the renal proximal tubule type IIa Na/Pi- cotransporter (rat NaPi IIa) and thereby inhibits its function. We tested the effect of TCEP on the murine type IIa Na/P(i)-cotransporter and the corresponding IIb intestinal isoform both expressed in Xenopus laevis oocytes. After incubation with TCEP the function of NaPi IIa was inhibited and protein amount was decreased. Injection of the lysosomal inhibitor leupeptin prevented degradation of the protein. Exposure of oocytes to TCEP at 0 degrees C led to a reduction in transport function without concomitant loss in Na/Pi IIa protein. In contrast to NaPi type IIa, the type IIb isoform was neither inhibited, nor degraded after incubation with TCEP. These results suggest that cleavage of disulfide bonds led to changes within the confirmation of the type IIa transporter that result in (i) inhibition of the transport activity and (ii) internalization and subsequent lysosomal degradation of transporter protein. Sequence comparisons suggest the involvement/presence of different disulfide bonds in type IIa and type IIb Na/P(i)-cotransporters.
Physiologisches Institut, Universität Zürich Irchel, Winterurerstr. 190, CH-8057 Zürich, Switzerland.
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Lambert
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Traebert
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Biber
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Murer
D000595:Amino Acid Sequence; D000818:Animals; D002352:Carrier Proteins / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D004220:Disulfides / Q000378:metabolism*; D015870:Gene Expression; D051379:Mice; D008969:Molecular Sequence Data; D009865:Oocytes / Q000187:drug effects / Q000378:metabolism; D010720:Phosphines / Q000494:pharmacology; D020033:Protein Isoforms / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D019163:Reducing Agents / Q000494:pharmacology; D017386:Sequence Homology, Amino Acid; D050608:Sodium-Phosphate Cotransporter Proteins; D050613:Sodium-Phosphate Cotransporter Proteins, Type IIa; D050614:Sodium-Phosphate Cotransporter Proteins, Type IIb; D027981:Symporters; D014982:Xenopus laevis
D002352:Carrier Proteins; D004220:Disulfides; D010720:Phosphines; D020033:Protein Isoforms; D019163:Reducing Agents; D050608:Sodium-Phosphate Cotransporter Proteins; D050613:Sodium-Phosphate Cotransporter Proteins, Type IIa; D050614:Sodium-Phosphate Cotransporter Proteins, Type IIb; D027981:Symporters; C493555:Slc34a1 protein, mouse; C493558:Slc34a2 protein, mouse; C080938:tris(2-carboxyethyl)phosphine