RESUMEN
The proton-coupled peptide transporter PEPT2 (SLC15A2) mediates the high-affinity low-capacity transport of small peptides as well as various oral peptide-like drugs in the kidney. In contrast to its well-characterized transport properties, there is less information available on its regulatory mechanism, although the interaction of PEPT2 to the PDZ (PSD-95, DglA, and ZO-1)-domain protein PDZK1 has been preliminarily reported. To examine whether PDZK1 is a physiological partner of PEPT2 in kidneys, we started from a yeast two-hybrid screen of a human kidney cDNA library with the C-terminus of PEPT2 (PEPT2 C-terminus (PEPT2-CT)) as bait. We could identify PDZK1 as one of the positive clones. This interaction requires the PDZ motif of PEPT2-CT detected by a yeast two-hybrid assay, in vitro binding assay and co-immunoprecipitation. The binding affinities of second and third PDZ domains of PDZK1 to PEPT2-CT were measured by surface plasmon resonance. Co-immunoprecipitation using human kidney membrane fraction and localization of PEPT2 in renal apical proximal tubules revealed the physiological meaning of this interaction in kidneys. Furthermore, we clarified the mechanism of enhanced glycylsarcosine (Gly-Sar) transport activity in PEPT2-expressing HEK293 cells after the PDZK1 coexpression. This augmentation was accompanied by a significant increase in the V(max) of Gly-Sar transport via PEPT2 and it was also associated with the increased surface expression level of PEPT2. These results indicate that the PEPT2-PDZK1 interaction thus plays a physiologically important role in both oligopeptide handling as well as peptide-like drug transport in the human kidney.
Asunto(s)
Proteínas Portadoras/metabolismo , Túbulos Renales Proximales/metabolismo , Simportadores/metabolismo , Transporte Biológico/fisiología , Proteínas Portadoras/química , Proteínas Portadoras/genética , Células Cultivadas , Biblioteca de Genes , Humanos , Inmunohistoquímica , Túbulos Renales Proximales/citología , Proteínas de la Membrana/química , Proteínas de la Membrana/genética , Proteínas de la Membrana/metabolismo , Unión Proteica/fisiología , Estructura Terciaria de Proteína , Resonancia por Plasmón de Superficie , Simportadores/genética , Técnicas del Sistema de Dos HíbridosRESUMEN
Elimination in urine and feces was compared between four perfluorinated fatty acids (PFCAs) with different carbon chain length. In male rats, perfluoroheptanoic acid (PFHA) was rapidly eliminated in urine with the proportion of 92% of the dose being eliminated within 120 h after an intraperitoneal injection. Perfluorooctanoic acid (PFOA), perfluorononanoic acid (PFNA) and perfluorodecanoic acid (PFDA) was eliminated in urine with the proportions of 55, 2.0 and 0.2% of the dose, respectively. By contrast, four PFCAs were eliminated in feces with the proportion of less than 5% of the dose within 120 h after an injection. In female rats, the proportions of PFOA and PFNA eliminated in urine within 120 h were 80% and 51% of the dose, respectively, which were significantly higher compared with those in male rats. There was the tendency that PFCA with longer carbon chain length is less eliminated in urine in both male and female rats. Fecal elimination of PFCAs was not different between PFCAs in female rats and comparable to those in male rats. The rates of biliary excretion of PFCAs in male rats were slower than those in female rats. Sex-related difference in urinary elimination of PFOA was abolished when male rats had been castrated. On the contrary, treatment with testosterone suppressed the elimination of PFOA in urine in both castrated male rats and female rats. The effect of testosterone was in a time- and dose-dependent manner. These results suggest that PFCAs are distinguished by their carbon chain length by a renal excretion system, which is regulated by testosterone.