Basic Fibroblast Growth Factor Stimulates Surface Expression and Activity of Na+/H+ Exchanger NHE3 via Mechanism Involving Phosphatidylinositol 3-Kinase*
- From the ‡Department of Medicine, Division of Gastroenterology, Hepatology, and Nutrition, University of Texas Medical School, Houston, Texas 77030 and the §Departments of Medicine and Physiology, Division of Gastroenterology, The Johns Hopkins University School of Medicine, Baltimore, Maryland 21205
Abstract
Na+/H+ exchanger NHE3 is a plasma membrane (PM) protein, which contributes to Na+ absorption in the intestine. Growth factors stimulate NHE3 via phosphatidylinositol 3-kinase (PI3-K), but mechanism of this process is not clear. To examine the hypothesis that growth factors stimulate NHE3 by modulating NHE3 recycling, and that PI3-K participates in this mechanism, we used PS120 fibroblasts expressing a fusion protein of NHE3 and green fluorescent protein. At steady state, ∼25% of cellular NHE3 content was expressed at PM. Inhibition of PI3-K decreased PM expression of NHE3, which correlated with retention of the exchanger in recycling endosomal compartment. In contrast, basic fibroblast growth factor (bFGF) increased PM expression of NHE3, which was associated with a 2-fold increase in rate constant for exit of the exchanger from the recycling compartment. Qualitatively similar effects of bFGF were observed in cells pretreated with PI3-K inhibitors, but their magnitude was only ∼50% of that in intact cells. These data suggest that: (i) bFGF stimulates NHE3 by increasing PM expression of the exchanger; (ii) PI3-K mediates PM expression of NHE3 in both basal and bFGF-stimulated conditions, and (iii) not all of the effects of bFGF on NHE3 expression are mediated by PI3-K, suggesting additional regulatory mechanisms.
Footnotes
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↵* This work was supported by National Institutes of Health NIDDK Grants K08DK02557, RO1DK26523, PO1DK44484, R29DK43778, and T32DK0763205, and by the Meyerhoff Digestive Diseases Center for Epithelial Disorders. Part of this work was presented at the 100th Annual Meeting of the American Gastroenterological Association, Orlando, FL, May 16–19, 1999 (Abstract 3877), and was published in abstract form (Janecki, A. J., Janecki, M., Akhter, S., and Donowitz, M. (1999) Gastroenterology 116, G3877).The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
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↵¶ To whom correspondence should be addressed: Div. of Gastroenterology, Hepatology and Nutrition, University of Texas Medical School, 6431 Fannin, 4.234 MSB, Houston, TX 77030. Tel.: 713-500-6649; Fax: 713-500-6699; E-mail: ajaneck@heart.med.uth.tmc.edu.
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↵2 A. J. Janecki, S. Akhter, M. Donowitz, and M. Janecki, submitted for publication.
- Abbreviations:
- NHE3
-
Na+/H+ exchanger isoform 3
- bFGF
-
basic fibroblast growth factor
- eGFP
-
red-shifted variant of green fluorescent protein
- EGF
-
epidermal growth factor
- GFP
-
green fluorescent protein
- JNC
-
juxtanuclear cytoplasmic compartment
- PCR
-
polymerase chain reaction
- PM
-
plasma membrane
- PI 3-K
-
phosphatidylinositol 3-kinase
- REC
-
recycling endosomal compartment
- Tf
-
transferrin
- TfR
-
transferrin receptor
- TMA
-
tetramethylammonium
- TX-Tf
-
Texas Red-conjugated human transferrin
- VSVG
-
vesicular stomatitis virus G protein
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- Received October 27, 1999.
- Revision received December 9, 1999.
- The American Society for Biochemistry and Molecular Biology, Inc.
