Identification and subcellular localization of the Na+/H+ exchanger and a novel related protein in the endocrine pancreas and adrenal medulla

Abstract

C exchangers (NHE) constitute a family of membrane antiporters that contribute to the regulation of cellular pH and volume in many tissues, including pancreatic islets. We investigated the molecular identity of NHE in rodent and human endocrine pancreas, and determined its cellular and subcellular localization. NHE1 was the most abundantly expressed isoform in rat islets, and was also expressed in mouse and human islets. By western blot, an antiserum raised against the C-terminus end of NHE1 confirmed the presence of a w100 kDa protein corresponding to NHE1 in islets and unexpectedly unveiled the existence of a w65 kDa cross-reactive NHE1-related protein. By immunohistochemistry, the antiserum labelled the membranes of pancreatic acini and ducts, but also diffusely stained the cytoplasm of insulin, glucagon and somatostatin cells as well as endocrine cells of the adrenal medulla. Electron microscopy localized the NHE1 immunoreactivity in the membrane of secretory granules, an unexpected finding supported by a decrease in immunohistochemical signal in degranulated b-cells. Islets of Slc9A1 swe/swe mice, which lack full NHE1 protein, were found to express an mRNA corresponding to the 3 0 end of NHE1 as well as the w65 kDa protein. They still showed the cytoplasmic labelling but no plasma membrane was stained. We conclude that both the full-length and the shorter-splice variant of NHE1 are expressed in all cell types of the endocrine pancreas and in the adrenal medulla of rodents and humans. The complete protein is addressed to the plasma membrane and the shorter one to the membrane of secretory granules where its function remains to be established.

Glucose-induced Cytosolic pH Changes in β-Cells and Insulin Secretion Are Not Causally Related

“…We confirm (35) that the corresponding mRNA is absent from their islets and show that no significant compensatory expression of another isoform occurred. Our observations that DMA had no effect on pH c in islets from Under basal conditions (3 mM glucose), the lack of NHE1 did not affect islet pH c in HCO 3 Ϫ medium but resulted in a marked lowering of pH c in HEPES medium.…”

“…NHE1 (encoded by Slc9A1) was the most expressed among the isoforms associated with the plasma membrane (NHE1-NHE5 and NHE10). Using radioactive RT-PCR, we recently showed that NHE1 is strongly expressed, whereas NHE2 and NHE5 are weakly expressed, and NHE3 and NHE4 are not detected in rat islets (35).…”

“…We recently showed that NHE1 is strongly expressed in rat islets and is present in mouse islets (35). Here, we first compared the expression of NHE isoforms in mouse islets.…”

Physiology, Pharmacology and Pathophysiology of the pH Regulatory Transport Proteins NHE1 and NBCn1: Similarities, Differences, and Implications for Cancer Therapy

“…[28]; ChIP sequencing data available on the Human Genome Browser; and section II.7 for further information on regulation of NHE1 expression). Although a few studies have reported NHE1 splice variants in rodent and human tissues [31][32][33], a single variant seems to be the only or by far the predominant form of NHE1 in most tissues.…”

Role of Na/H exchange in insulin secretion by islet cells

“…These findings suggested unspecific, NHE1-independent effects of EIPA and DMA in islets, possibly involving KEY POINTS inhibition of another amiloride-sensitive NHE or direct inhibition of K ATP channels [22,23]. In addition to full-length plasmalemmal NHE1, b cells were found to express a shorter splice variant of NHE1 that localizes specifically to insulincontaining LDCVs [7]. Although full-length NHE1 protein ($100 kDa) was absent from islets in NHE1 swe/swe mutant mice, the shorter splice variant ($65 kDa) was still present.…”