Characterization of mouse organic anion transporter 5 as a renal steroid sulfate transporter

A family of organic anion transporters (OAT) recently identified has important roles for the excretion or reabsorption of endogenous and exogenous compounds, and several new isoforms have been reported in this decade. Although the transepithelial transport properties of organic anions are gradually being understood, many portions of their functional characteristics in functions remain to be elucidated. A recently reported new cDNA encoding a mouse OAT5 (mOAT5) was constructed, using 3'-RACE PCR, with the total RNA isolated from a mouse kidney. When mOAT5 was expressed in Xenopus oocytes, mOAT5 transported estrone sulfate, dehydroepiandrosterone sulfate and ochratoxin A. Estrone sulfate uptake by mOAT5 displayed a time-dependent and sodium-independent manner. The Km values of estrone sulfate and dehydroepiandrosterone sulfate were 2.2 and 3.8 microM, respectively. mOAT5 interacted with chemically heterogeneous steroid or organic sulfates, such as nitrophenyl sulfate, methylumbelliferyl sulfate and estradiol sulfates. In contrast to the sulfate conjugates, mOAT5-mediated estrone sulfate uptake was not inhibited by the steroid or organic glucuronides. The mOAT5 protein having about 85 kDa molecular weight was shown to be mainly localized in the apical membrane of the proximal tubules of the outer medulla. These results suggest an important role of mOAT5 for the excretion or reabsorption of steroid sulfates in the kidney.     

Identification of a novel voltage-driven organic anion transporter present at apical membrane of renal proximal tubule

A novel transport protein with the properties of voltage-driven organic anion transport was isolated from pig kidney cortex by expression cloning in Xenopus laevis oocytes. A cDNA library was constructed from size-fractionated poly(A)+ RNA and screened for p-aminohippurate (PAH) transport in high potassium medium. A 1856-base pair cDNA encoding a 467-amino acid peptide designated as OATV1 (voltage-driven organic anion transporter 1) was isolated. The predicted amino acid sequence of OATV1 exhibited 60-65% identity to those of human, rat, rabbit, and mouse sodium-dependent phosphate cotransporter type 1 (NPT1), although OATV1 did not transport phosphate. The homology of this transporter to known members of the organic anion transporter family (OAT family) was about 25-30%. OATV1-mediated PAH transport was affected by the changes in membrane potential. The transport was Na+-independent and enhanced at high concentrations of extracellular potassium and low concentrations of extracellular chloride. Under the voltage clamp condition, extracellularly applied PAH induced outward currents in oocytes expressing OATV1. The current showed steep voltage dependence, consistent with the voltage-driven transport of PAH by OATV1. The PAH transport was inhibited by various organic anions but not by organic cations, indicating the multispecific nature of OATV1 for anionic compounds. This transport protein is localized at the apical membrane of renal proximal tubule, consistent with the proposed localization of a voltage-driven organic anion transporter. Therefore, it is proposed that OATV1 plays an important role to excrete drugs, xenobiotics, and their metabolites driven by membrane voltage through the apical membrane of the tubular epithelial cells into the urine.     

Functional expression of pig renal organic anion transporter 3 (pOAT3)

With the cloning of pig renal organic anion transporter 1 (pOAT1) (Biochimie 84 (2002) 1219) we set up a model system for comparative studies of cloned and natively isolated membrane located transport proteins. Meanwhile, another transport protein involved in p-aminohippurate (PAH) uptake on the basolateral side of the proximal tubule cells was identified, designated organic anion transporter 3 (OAT3). To explore the contribution of pOAT1 to the PAH clearance in comparison to OAT3, it was the aim of this study to extend our model by cloning of the pig ortholog of OAT3. Sequence comparisons of human organic anion transporter 3 (hOAT3) with the expressed sequence tag (EST) database revealed a clone and partial sequence of the pig renal organic anion transporter 3 (pOAT3) ortholog. Sequencing of the entire open reading frame resulted in a protein of 543 amino acid residues encoded by 1632 base pairs (EMBL Acc. No. AJ587003). It showed high homologies of 81%, 80%, 76%, and 77% to the human, rabbit, rat, and mouse OAT3, respectively. A functional characterization of pOAT3 in Xenopus laevis oocytes yielded an apparent Km (Kt) for [3H]estrone sulfate of 7.8 +/- 1.3 microM. Moreover, pOAT3 mediated [3H]estrone sulfate uptake was almost abolished by 0.5 mM of glutarate, dehydroepiandosterone sulfate, or probenecid consistent with the hallmarks of OAT3 function.     

Cloning of the pig renal organic anion transporter 1 (pOAT1)

A pig kidney cDNA library was screened for the porcine ortholog of the multispecific organic anion transporter 1 (pOAT1). Several positive clones were isolated resulting in two alternatively spliced cDNA clones of pOAT1 (pOAT1 and pOAT1A). pOAT1-cDNAs consist of 2126 or 1895 base pairs (EMBL Acc. No. AJ308234 and AJ308235) encoding 547 or 533 amino acid residue proteins with 89, 87, 83 and 81% homology to the human, rabbit, rat, and mouse OAT1, respectively. Heterologous expression of pOAT1 in Xenopus laevis oocytes revealed an apparent K(m) for [3H]PAH of 3.75 +/- 1.6 microM. [3H]PAH uptake mediated by pOAT1 was abolished by 0.5 mM glutarate or 1 mM probenecid. Functional characterization of pOAT1A did not show any affinity for [3H]PAH. In summary, we cloned two alternative splice variants of the pig ortholog of organic anion transporter 1. One splice form (pOAT1) showed typical functional characteristics of organic anion transporter 1, whereas the second form appears not to transport PAH.     

Functional differences in steroid sulfate uptake of organic anion transporter 4 (OAT4) and organic anion transporting polypeptide 2B1 (OATP2B1) in human placenta

Human trophoblasts depend on the supply of external precursors such as dehydroepiandrosterone-3-sulfate (DHEA-S) and 16alpha-OH-DHEA-S for synthesis of estrogens. Recently, we have characterized the uptake of DHEA-S by isolated mononucleated trophoblasts and identified different transporter polypeptides involved in this process. Immunohistochemistry of 1st and 3rd trimester placenta detected organic anion transporter 4 (OAT4) and organic anion transporting polypeptide 2B1 (OATP2B1, former name OATP-B) in cytotrophoblast membranes and at the basal surface of the syncytiotrophoblast, indicating that both transporter polypeptides are involved in placental uptake of foetal derived steroid sulfates. In the present study we have characterized and compared the kinetics of DHEA-S and estrone sulfate (E(1)S) uptake by these transporters stably expressed in FlpIn -HEK293 cells using the Flp recombinase-mediated site-specific recombination. Uptake of E(1)S by OAT4- and OATP2B1-transfected cells was highly increased compared to the non-transfected cells. In contrast, DHEA-S uptake was only highly increased in OAT4 (40 times), but only weakly enhanced in OATP2B1 cells. The uptake of DHEA-S and E(1)S by OAT4 was partly Na(+)-dependent (about 50%), whereas uptake of DHEA-S by OATP2B1 was Na(+)-independent. Kinetic analysis of the initial uptake rates of E(1)S by OAT4 and OATP2B1 gave very similar values for K(m) (about 20microM) and V(max) (about 600pmol/(minxmg protein)). In contrast, the affinity of DHEA-S towards OATP2B1 was about 10 times lower (K(m)>200microM) then for OAT4 (K(m)=29microM). Our results suggest different physiological roles of the two transporter polypeptides in placental uptake of foetal derived steroid sulfates. OATP2B1 seems not to be involved in de novo synthesis of placental estrogens but may contribute to the clearance of estrogen sulfates from foetal circulation.     

Regulation by protein kinase C of organic anion transport driven by rat organic anion transporter 3 (rOAT3)

The organic anion transporter 3 (rOAT3) is a multispecific OAT localized at the basolateral membrane of the proximal tubule. The purpose of this study was to elucidate the role of protein kinase C (PKC) in the regulation of organic anion transport driven by rOAT3 and its mechanism of action. For this purpose, we established and utilized cells derived from the second segment of proximal tubule from mice stably expressing rOAT3 (S2 rOAT3). Phorbol 12-myristate 13-acetate (PMA), a PKC stimulator, attenuated the cellular uptake of estrone sulfate (ES), a prototype organic anion for rOAT3, in a dose- and time-dependent manner. PMA treatment resulted in a decrease in the Vmax, but not the Km of uptake of ES in S2 rOAT3. Treatment of S2 rOAT3 with other PKC stimulators or diacylglycerols also inhibited the uptake of ES, whereas that with an inactive phorbol ester did not. Chelerythrine chloride, a PKC inhibitor, reversed the PMA-induced decrease in uptake of ES in S2 rOAT3. These results suggest that PKC activation downregulates rOAT3-mediated organic anion transport. This down-regulation may be due to the inhibition of translocation or internalization of the rOAT3 protein, resulting in the decrease in the Vmax of rOAT3-mediated organic anion transport.     

Evidence for rat organic anion transporter 3 association with caveolin-1 in rat kidney

The rat organic anion transporter 3 (rOAT3) has recently been identified as the third isoform of the OAT family. The mechanisms that regulate rOAT3's functions remain to be elucidated. rOAT3 contributes for moving a number of negatively charged organic compounds between cells and their extracellular milieu. Caveolin (Cav) also plays a role as a membrane transporter. To address the relationship of these two proteins, we investigated the protein-protein interaction between rOAT3 and Cav-1. The rOAT3 mRNA and protein expression were observed in the rat kidney, and the expressions of Cav-1 mRNA and protein were also detected in the kidney. Confocal microscopy of the immuno-cytochemistry experiments using primary cultured renal proximal tubular cells showed that rOAT3 and Cav-1 were co-localized at the plasma membrane. This finding was confirmed by Western blot analysis using isolated caveolae-enriched membrane fractions from the rat kidney and immuno-precipitation experimentation. When rOAT3's synthesized cRNA of rOAT3 along with the antisense oligo deoxynucleotide ofXenopusCav-1 were co-injected intoXenopusoocytes, the [(3)H] estrone sulfate uptake was significantly decreased. These findings suggest that rOAT3 and caveolin-1 share a cellular expression in the plasma membrane and Cav-1 up-regulates the organic anionic compound uptake via rOAT3 under normal physiological conditions.     

Characterization of an organic anion transport system in a placental cell line

Transporters within the placenta play a crucial role in the distribution of nutrients and xenobiotics across the maternal-fetal interface. An organic anion transport system was identified on the apical membrane of the rat placenta cell line HRP-1, a model for the placenta barrier. The apical uptake of 3H-labeled organic anion estrone sulfate in HRP-1 cells was saturable (Km = 4.67 microM), temperature and Na+ dependent, Li+ tolerant, and pH sensitive. The substrate specificity of the transport system includes various steroid sulfates, such as beta-estradiol 3,17-disulfate, 17 beta-estradiol 3-sulfate, and dehydroepiandrosterone 3-sulfate (DHEAS) but does not include taurocholate, p-aminohippuric acid (PAH), and tetraethylammonium. Preincubation of HRP-1 cells with 8-bromo-cAMP (a cAMP analog) and forskolin (an adenylyl cyclase activator) acutely stimulated the apical transport activity. This stimulation was further enhanced in the presence of IBMX (a phosphodiesterase inhibitor). Together these data show that the apical membrane of HRP-1 cells expresses an organic anion transport system that is regulated by cellular cAMP levels. This transport system appears to be different from the known taurocholate-transporting organic anion-transporting polypeptides and PAH-transporting organic anion transporters, both of which also mediate the transport of estrone sulfate and DHEAS.     

The SUMO-Specific Protease Senp2 Regulates SUMOylation,Expression and Function of Human Organic Anion Transporter 3

Organic anion transporter 3 (OAT3) plays a vital role in removing a broad array of anionic drugs from kidney, thereby avoiding their possibly toxic side effects in the body. We earlier demonstrated that OAT3 is subjected to a specific type of post-translational modification called SUMOylation. SUMOylation is a dynamic event, where de-SUMOylation is catalyzed by a class of SUMO-specific proteases. In the present investigation, we assessed the role of SUMO-specific protease Senp2 in OAT3 SUMOylation, expression and function. We report here that overexpression of Senp2 in COS-7 cells led to a reduced OAT3 SUMOylation, which correlated well with a decreased OAT3 expression and transport activity. Such phenomenon was not observed in cells overexpressing an inactive mutant of Senp2. Furthermore, transfection of cells with Senp2-specific siRNA to knockdown the endogenous Senp2 resulted in an increased OAT3 SUMOylation, which correlated well with an enhanced OAT3 expression and transport activity. Coimmunoprecipitation experiments showed that Senp2 directly interacted with OAT3 in the kidneys of rats. Together these results provided first demonstration that Senp2 is a significant regulator for OAT3-mediated organic anion/drug transport.     

Involvement of rat organic anion transporter 3 (rOAT3) in cephaloridine-induced nephrotoxicity: in comparison with rOAT1

This study was performed to elucidate the possible involvement of organic anion transporter 3 (OAT3) in cephaloridine (CER)-induced nephrotoxicity and compare the substrate specificity between rOAT3 and rat OAT1 (rOAT1) for various cephalosporin antibiotics, using proximal tubule cells stably expressing rOAT3 (S2 rOAT3) and rOAT1 (S2 rOAT1). S2 rOAT3 exhibited a CER uptake and a higher susceptibility to CER cytotoxicity than did mock, which was recovered by probenecid. Various cephalosporin antibiotics significantly inhibited both estrone sulfate uptake in S2 rOAT3 and para-aminohippuric acid uptake in S2 rOAT1. The Ki values of CER, cefoperazone, cephalothin and cefazolin for rOAT3- and rOAT1-mediated organic anion transport ranged from 0.048 to 1.14 mM and from 0.48 to 1.32 mM, respectively. These results suggest that rOAT3, at least in part, mediates CER uptake and CER-induced nephrotoxicity as rOAT1. There was some difference of affinity between rOAT3 and rOAT1 for cephalosporin antibiotics.     

Cloning of the human thiamine transporter, a member of the folate transporter family.

We have isolated a cDNA from human placenta, which, when expressed heterologously in mammalian cells, mediates the transport of the water-soluble vitamin thiamine. The cDNA codes for a protein of 497 amino acids containing 12 putative transmembrane domains. Northern blot analysis indicates that this transporter is widely expressed in human tissues. When expressed in HeLa cells, the cDNA induces the transport of thiamine (K(t) = 2.5 +/- 0.6 microM) in a Na(+)-independent manner. The cDNA-mediated transport of thiamine is stimulated by an outwardly directed H(+) gradient. Substrate specificity assays indicate that the transporter is specific to thiamine. Even though thiamine is an organic cation, the cDNA-induced thiamine transport is not inhibited by other organic cations. Similarly, thiamine is not a substrate for the known members of mammalian organic cation transporter family. The thiamine transporter gene, located on human chromosome 1q24, consists of 6 exons and is most likely the gene defective in the metabolic disorder, thiamine-responsive megaloblastic anemia. At the level of amino acid sequence, the thiamine transporter is most closely related to the reduced-folate transporter and thus represents the second member of the folate transporter family.     

Regulation of human organic anion transporter 4 by parathyroid hormone-related protein and protein kinase A

Human organic anion transporter 4 (hOAT4) belongs to a family of organic anion transporters that play critical roles in the body disposition of clinically important drugs, including anti-human immunodeficiency virus therapeutics, anti-tumor drugs, antibiotics, antihypertensives, and anti-inflammatories. hOAT4 is abundantly expressed in the kidney and placenta. In the current study, we examined the regulation of hOAT4 by parathyroid hormone-related protein (PTHrP) and protein kinase A (PKA) in kidney COS-7 cells. PTHrP induced a time- and concentration-dependent stimulation of hOAT4 transport activity. The stimulation of hOAT4 activity by PTHrP mainly resulted from an increased cell surface expression without a change in total cell expression of the transporter. Activation of PKA by Bt2-cAMP also resulted in a stimulation of hOAT4 activity through an increased cell surface expression of the transporter. However, PTHrP-induced stimulation of hOAT4 activity could not be prevented by treating hOAT4-expressing cells with the PKA inhibitor H89. We concluded that both PTHrP and activation of PKA stimulate hOAT4 activity through redistribution of the transporter from intracellular compartments to the cell surface. However, PTHrP regulates hOAT4 activity by mechanisms independent of PKA pathway.     

Impaired organic anion transport in kidney and choroid plexus of organic anion transporter 3 (Oat3 (Slc22a8)) knockout mice

To begin to develop in vivo model systems for the assessment of the contributions of specific organic anion transporter (OAT) family members to detoxification, development, and disease, we carried out a targeted disruption of the murine organic anion transporter 3 (Oat3) gene. Surviving Oat3(-/-) animals appear healthy, are fertile, and do not exhibit any gross morphological tissue abnormalities. No Oat3 mRNA expression was detected in kidney, liver, or choroid plexus (CP) of Oat3(-/-) mice. A distinct phenotype manifested by a substantial loss of organic anion transport capacity in kidney and CP was identified. Uptake sensitive to inhibition by bromosulfophthalein or probenecid was observed for taurocholate, estrone sulfate, and para-aminohippurate in renal slices from wild-type mice, whereas in Oat3(-/-) animals transport of these substances was greatly reduced. No discernable differences in uptake were observed between hepatic slices from wild-type and Oat3(-/-) littermates, suggesting Oat3 does not play a major role in hepatic organic anion uptake. Cellular accumulation of fluorescein was reduced by approximately 75% in CP from Oat3(-/-) mice. However, capillary accumulation of fluorescein-methotrexate was unchanged, indicating the effects of Oat3 loss are restricted to the entry step and that Oat3 is localized to the apical membrane of CP. These data indicate a key role for Oat3 in systemic detoxification and in control of the organic anion distribution in cerebrospinal fluid.     

IRIP, a new ischemia/reperfusion-inducible protein that participates in the regulation of transporter activity

We report the identification and characterization of a new ischemia/reperfusion-inducible protein (IRIP), which belongs to the SUA5/YrdC/YciO protein family. IRIP cDNA was isolated in a differential display analysis of an ischemia/reperfusion-treated kidney RNA sample. Mouse IRIP mRNA was expressed in all tissues tested, the highest level being in the testis, secretory, and endocrine organs. Besides ischemia/reperfusion, endotoxemia also activated the expression of IRIP in the liver, lung, and spleen. The transporter regulator RS1 was identified as an IRIP-interacting protein in yeast two-hybrid screening. The interaction between IRIP and RS1 was further confirmed in coimmunoprecipitation assays. A possible role of IRIP in regulating transporter activity was subsequently investigated. IRIP overexpression inhibited endogenous 1-methyl-4-phenylpyridinium (MPP+) uptake activity in HeLa cells. The activities of exogenous organic cation transporters (OCT2 and OCT3), organic anion transporter (OAT1), and monoamine transporters were also inhibited by IRIP. Conversely, inhibition of IRIP expression by small interfering RNA or antisense RNA increased MPP+ uptake. We measured transport kinetics of OCT2-mediated uptake and demonstrated that IRIP overexpression significantly decreased V(max) but did not affect K(m). On the basis of these results, we propose that IRIP regulates the activity of a variety of transporters under normal and pathological conditions.     

Characterization of the transport properties of cloned rat multidrug resistance-associated protein 3 (MRP3).

We have previously cloned rat MRP3 as an inducible transporter in the liver (Hirohashi, T., Suzuki, H., Ito, K., Ogawa, K., Kume, K., Shimizu, T., and Sugiyama, Y. (1998) Mol. Pharmacol. 53, 1068-1075). In the present study, the function of rat MRP3 was investigated using membrane vesicles isolated from LLC-PK1 and HeLa cell population transfected with corresponding cDNA. The ATP-dependent uptake of both 17beta estradiol 17-beta-D-glucuronide ([3H]E217betaG) and glucuronide of [14C] 6-hydroxy-5, 7-dimethyl-2-methylamino-4-(3-pyridylmethyl) benzothiazole (E3040), but not that of [3H]leukotriene C4 and [3H]2, 4-dinitrophenyl-S-glutathione, was markedly stimulated by MRP3 transfection in both cell lines. The Km and Vmax values for the uptake of [3H]E217betaG were 67 +/- 14 microM and 415 +/- 73 pmol/min/mg of protein, respectively, for MRP3-expressing membrane vesicles and 3.0 +/- 0.7 microM and 3.4 +/- 0.4 pmol/min/mg of protein, respectively, for the endogenous transporter expressed on HeLa cells. [3H]E217betaG had also a similar Km value for MRP3 when LLC-PK1 cells were used as the host. All glucuronide conjugates examined (E3040 glucuronide, 4-methylumbelliferone glucuronide, and naphthyl glucuronide) and methotrexate inhibited MRP3-mediated [3H]E217betaG transport in LLC-PK1 cells. Moreover, [3H]methotrexate was transported via MRP3. The inhibitory effect of estrone sulfate, [3H]2,4-dinitrophenyl-S-glutathione, and [3H]leukotriene C4 was moderate or minimal, whereas N-acetyl-2,4-dinitrophenylcysteine had no effect on the uptake of [3H]E217betaG. The uptake of [3H]E217betaG was enhanced by E3040 sulfate and 4-methylumbelliferone sulfate. Thus we were able to demonstrate that several kinds of organic anions are transported via MRP3, although the substrate specificity of MRP3 differs from that of MRP1 and cMOAT/MRP2 in that glutathione conjugates are poor substrates for MRP3.     

Activation of Protein Kinase C�� Increases OAT1 (SLC22A6)- and OAT3 (SLC22A8)-mediated Transport

Organic anion transporters (OATs) play a pivotal role in the clearance of small organic anions by the kidney, yet little is known about how their activity is regulated. A yeast two-hybrid assay was used to identify putative OAT3-associated proteins in the kidney. Atypical protein kinase Cζ (PKCζ) was shown to bind to OAT3. Binding was confirmed in immunoprecipitation assays. The OAT3/PKCζ interaction was investigated in rodent renal cortical slices from fasted animals. Insulin, an upstream activator of PKCζ, increased both OAT3-mediated uptake of estrone sulfate (ES) and PKCζ activity. Both effects were abolished by a PKCζ-specific pseudosubstrate inhibitor. Increased ES transport was not observed in renal slices from OAT3-null mice. Transport of the shared OAT1/OAT3 substrate, ρ-aminohippurate, behaved similarly, except that stimulation was reduced, not abolished, in the OAT3-null mice. This suggested that OAT1 activity was also modified by PKCζ, subsequently confirmed using an OAT1-specific substrate, adefovir. Inhibition of PKCζ also blocked the increase in ES uptake seen in response to epidermal growth factor and to activation of protein kinase A. Thus, PKCζ acted downstream of the epidermal growth factor to protein kinase A signaling pathway. Activation of transport was accompanied by an increase in V_max and was blocked by microtubule disruption, indicating that activation may result from trafficking of OAT3 into the plasma membrane. These data demonstrate that PKCζ activation up-regulates OAT1 and OAT3 function, and that protein-protein interactions play a central role controlling these two important renal drug transporters.Organic anion transporters (OATs)⁷ are members of the solute carrier 22A family and play a pivotal role in the renal clearance of small (<500 Dalton) anionic drugs, xenobiotics, and their metabolites. OAT substrates include a variety of drugs such as β-lactam antibiotics, non-steroidal anti-inflammatory drugs, diuretics, and chemotherapeutics (1). OATs are predominantly expressed in renal proximal tubule, with OATs 1–3 localized to the basolateral membrane and OAT4 and URAT1 on the apical membrane. OATs 1 and 3 are dicarboxylate exchangers, and are indirectly coupled to the sodium gradient maintained by Na,K-ATPase through sodium/dicarboxylate co-transport to drive the uphill basolateral step in renal organic anion secretion (2).Although the ionic gradients, electrophysiology, and underlying kinetics that drive transport by OATs 1 and 3 are well characterized, physiologically important interactions of these basolateral OATs with membrane or cytosolic proteins have yet to be identified (1). Nevertheless, there is clear evidence that other plasma membrane transporters do interact with protein partners, influencing a diverse array of functions including transport itself, cytoskeletal structure, vesicle formation, and trafficking, as well as signaling (3). Among the transporters with activity modulated by protein-protein interactions, particularly by the PDZ proteins, PDZK1 and NHERFs 1 and 2, are apical drug transporters of the SLC22A family, including OCTN1, OCTN2, OAT4, and URAT1 (4–6).In the present study, we have used a yeast two-hybrid assay to identify putative protein partners that interact directly with OAT3. The C-terminal 81 amino acids of OAT3 were used as bait to screen a human cDNA kidney library. Among the 23 positive clones (putative binding partners) was a clone encoding the C-terminal 141 amino acids of atypical protein kinase Cζ (PKCζ). Functional consequences of the putative OAT3/PKCζ interaction were investigated in rodent renal slices. The resulting data indicate that activation of PKCζ by insulin or epidermal growth factor (EGF) increased OAT3- and OAT1-mediated transport. Thus, PKCζ controls function of both major secretory organic anion transporters expressed at the basolateral face of the renal proximal tubule, positioning it to regulate the efficacy of renal drug elimination.