Distinct functional properties of two electrogenic isoforms of the SLC34 Na-Pi cotransporter.

Natsuki Mizutani,Yoshifumi Okochi,Yasushi Okamura

doi:10.14814/phy2.14156

Natsuki Mizutani, Yoshifumi Okochi + Show 1 more

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https://doi.org/10.14814/phy2.14156

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Journal: Physiological reports	Publication Date: Jul 1, 2019
Citations: 4	License type: CC BY 4.0

Affiliation: Osaka University

Abstract

Inorganic phosphate (Pi) is crucial for proper cellular function in all organisms. In mammals, type II Na‐Pi cotransporters encoded by members of the Slc34 gene family play major roles in the maintenance of Pi homeostasis. However, the molecular mechanisms regulating Na‐Pi cotransporter activity within the plasma membrane are largely unknown. In the present study, we used two approaches to examine the effect of changing plasma membrane phosphatidylinositol 4,5‐bisphosphate (PI(4,5)P2) levels on the activities of two electrogenic Na‐Pi cotransporters, NaPi‐IIa and NaPi‐IIb. To deplete plasma membrane PI(4,5)P2 in Xenopus oocytes, we utilized Ciona intestinalis voltage‐sensing phosphatase (Ci‐VSP), which dephosphorylates PI(4,5)P2 to phosphatidylinositol 4‐phosphate (PI(4)P). Upon activation of Ci‐VSP, NaPi‐IIb currents were significantly decreased, whereas NaPi‐IIa currents were unaffected. We also used the rapamycin‐inducible Pseudojanin (PJ) system to deplete both PI(4,5)P2 and PI(4)P from the plasma membrane of cultured Neuro 2a cells. Depletion of PI(4,5)P2 and PI(4)P using PJ significantly reduced NaPi‐IIb activity, but NaPi‐IIa activity was unaffected, which excluded the possibility that NaPi‐IIa is equally sensitive to PI(4,5)P2 and PI(4)P. These results indicate that NaPi‐IIb activity is regulated by PI(4,5)P2, whereas NaPi‐IIa is not sensitive to either PI(4,5)P2 or PI(4)P. In addition, patch clamp recording of NaPi‐IIa and NaPi‐IIb currents in cultured mammalian cells enabled kinetic analysis with higher temporal resolution, revealing their distinct kinetic properties.

Highlights

Inorganic phosphate (Pi) is essential for a variety of cell activities, including bioenergetics and cell signaling
Absorption of Pi in both the intestine and kidney is mediated primarily by type II Na-Pi cotransporters, which consist of three members, NaPiIIa (SLC34A1), NaPi-IIb (SLC34A2), and NaPi-IIc (SLC34A3), showing different patterns of expression (Wagner et al 2014): NaPi-IIa and NaPi-IIc are localized in the apical brush border membrane of renal proximal tubule cells, whereas NaPi-IIb is found in many tissues, including the luminal brush border membrane of the small intestine and alveolar type II epithelial cells (Traebert et al 1999; Wagner et al 2014)
Physiological Reports published by Wiley Periodicals, Inc. on behalf of The Physiological Society and the American Physiological Society

Summary

Introduction

Inorganic phosphate (Pi) is essential for a variety of cell activities, including bioenergetics and cell signaling. 1,25-(OH)2-Vitamin D3, which stimulates calcium uptake in the small intestine, increases the number of NaPi-IIb in the brush border membrane of this tissue (Hattenhauer et al 1999). These cotransporter activities are regulated by membrane potential (Forster et al 1998; Hilfiker et al 1998) and extracellular pH (de la Horra et al 2000). PI(4,5)P2-sensitive ion channel activities are regulated through stimulation of Gq-coupled receptors, which in turn mediate PLC-catalyzed PI(4,5)P2 hydrolysis (Hille et al 2015). Such regulation is well characterized in cardiac cells and neurons (Cho et al 2005; Morita et al 2006). We measured mNaPi-IIa and mNaPi-IIb currents in cultured mammalian cells, Neuro 2a, using the whole-cell patch clamp technique and revealed their distinct kinetic properties

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