Regulation of Na+/H+ Exchanger 1 (NHE1) Stability in PTEN-/- Mouse Embryonic Fibroblasts

Abstract

The Na+/H+ Exchanger 1 (NHE1), a membrane antiporter, has been implicated in tumour cell survival. However, the regulation of NHE1 expression level in tumour cells is not completely understood. Interestingly, the upregulation of NHE1 protein expression in PTEN-null Mouse embryonic fibroblasts (MEFPTEN-/-) compared to PTEN-wildtype Mouse embryonic fibroblasts (MEFPTEN+/+) does not correlate with an increase in NHE1 transcription. Hence, we hypothesized that increased NHE1 expression may be due to increased stability of the protein. In the current study, we show that Heat shock protein 90 (Hsp90) confers stability to NHE1 though enhanced interaction in MEFPTEN-/- cells as compared to MEFPTEN+/+ cells. The inhibition of Hsp90 activity using Geldanamycin and silencing of Hsp90 gene induced a decrease in NHE1 protein level in MEFPTEN-/- cells while no effect was observed in control cells. In addition, NHE1 was found to interact with Akt in MEFPTEN-/- cells. Both inhibition of Akt activation and silencing Akt gene expression decreased NHE1 protein expression. Moreover, our data reveals that Akt is involved in trafficking Hsp90 from the cytosol to the plasma membrane in MEFPTEN-/- cells. Similar result was observed in MCF10CA1a, a breast carcinoma cancer cell line. Additionally, we found that NHE1 may be involved in the regulation of Akt Serine 473 phosphorylation. We show that this occurs through the complex formation between NHE1, mTOR, Rictor in both MEFPTEN-/- and MCF10CA1a viii cells. Taken together, this study establishes a possible tripartite complex between NHE1, Akt and Hsp90 that may be critical for stabilizing NHE1 expression level and its role in survival of tumour cells. We propose that our findings might be critical for the development and maintenance of tumour cells. Indeed, results from this study could establish a new strategy to consider when developing new chemotherapeutic or even chemo-preventive drugs