FXYD3 (Mat-8), a New Regulator of Na,K-ATPase

Abstract

Four of the seven members of the FXYD protein family have been identified as specific regulators of Na,K-ATPase. In this study, we show that FXYD3, also known as Mat-8, is able to associate with and to modify the transport properties of Na,K-ATPase. In addition to this shared function, FXYD3 displays some uncommon characteristics. First, in contrast to other FXYD proteins, which were shown to be type I membrane proteins, FXYD3 may have a second transmembrane-like domain because of the presence of a noncleavable signal peptide. Second, FXYD3 can associate with Na,K-as well as H,K-ATPases when expressed in Xenopus oocytes. However, in situ (stomach), FXYD3 is associated only with Na,KATPase because its expression is restricted to mucous cells in which H,K-ATPase is absent. Coexpressed in Xenopus oocytes, FXYD3 modulates the glycosylation processing of the ␤ subunit of X,K-ATPase dependent on the presence of the signal peptide. Finally, FXYD3 decreases both the apparent affinity for Na ؉ and K ؉ of Na,K-ATPase. INTRODUCTIONThe Na,K-ATPase is an ubiquitous enzyme consisting of an ␣ and a ␤ subunit, which is responsible for the creation and maintenance of the Na ϩ and K ϩ gradients across the cell membrane by transporting three Na ϩ out and two K ϩ into the cell. This function is crucial for cell survival and body homeostasis because the Na ϩ gradient is used as an energy source to transport ions or solutes and is at the origin of the vectorial Na ϩ reabsorption in the kidney and of action potentials in excitable tissues.Regulation of the activity and expression of Na,K-ATPase is tight and governed by a variety of mechanisms. Shortterm regulation involves protein kinases and results in modulation of the cell surface expression of the Na,K-ATPase, whereas long-term regulation, mediated by mineralocorticoid or thyroid hormone, leads to a change in the total number of Na,K-ATPase units (Therien and Blostein, 2000; for review, see Feraille and Doucet, 2001). Moreover, the existence of multiple ␣ and ␤ isoforms permits the production of isozymes with different transport properties (Crambert et al., 2000). Finally, recent experimental evidence shows that members of the FXYD protein family specifically associate with and modulate the transport properties of Na,KATPase (for review, see Crambert and Geering, 2003).The FXYD family contains seven members that share a common signature sequence encompassing the transmembrane and adjacent regions (Sweadner and Rael, 2000). So far, all characterized FXYD proteins exhibit a similar structure with a single transmembrane domain and a type I orientation that is achieved, in some, but not all cases, by the cleavage of an N-terminal signal peptide. Among the seven members (FXYD1-7), four FXYD proteins have so far been shown to regulate the Na,K-ATPase in a tissue-and isozyme-specific way. All affect the apparent affinity for extracellular K ϩ of the Na,K-ATPase, which in the case of FXYD7 (the brain-specific FXYD protein) is thought to be physiologically relevant in neuronal excitabilit...

Interaction of Mat-8 (FXYD-3) with Na+/K+-ATPase in Colorectal Cancer Cells

“…These results suggest that mMat-8-Myc is effectively sorted to the surface membrane via vesicular transport in Colon-26 cells, possibly due to its association with partner molecule(s). 5) Interestingly, the Na ϩ /K ϩ -ATPase a subunit, one of such candidate molecules, 1,2) was also detected on the surface membrane and colocalized with mMat-8-Myc (Fig. 3, lower center and right).…”

“…In each collagen-coated (Cell matrix type IV, Nitta Gelatin) well of a 12-well culture plate, 2.5ϫ10 5 and 1ϫ10 5 cells (CHO-K1 and Colon-26 cells, respectively) were seeded in 1 ml of the medium containing 7% (v/v) FBS and then cultured for 24 h. Lipofectamine TM (Invitrogen) was used for transfection of plasmid DNA (0.5 mg), the plasmid solution (5 ml) being diluted with 45 ml of the medium without antibiotics and serum [medium(Ϫ)]. Lipofectamine (2 ml and 4 ml for CHO-K1 and Colon-26 cells, respectively) was diluted up to 50 ml with medium(Ϫ).…”

“…4) Among the FXYD proteins, Mat-8 is unique in that its signal sequence is present as an additional transmembrane segment without cleavage by signal peptidase. 5) Although it has been demonstrated that Mat-8 induces chloride currents in Xenopus oocytes under hyperpolarization, 4) the physiological role of Mat-8 remains to be unveiled.…”

Interaction with the Na,K-ATPase and Tissue Distribution of FXYD5 (Related to Ion Channel)

“…Independently, human FXYD5 was identified as a cancer-associated cell membrane protein, which down-regulates E-cadherin and promotes metastasis, and was termed dysadherin (14). 4 Dysadherin has been reported to be expressed in the plasma membrane of several types of carcinoma cells but not in nontumor cells (18 -20). The apparent molecular mass of the protein detected in these studies is 50 -55 kDa, much higher than its calculated molecular mass (Ͻ20 kDa).…”

Down-Regulation of FXYD3 Expression in Human Lung Cancers

“…Mammary tumor 8 kDa (MAT8)/FXDY domain-containing ion transport regulator 3 (FXYD3), a FXYD-containing Na,K-ATPase ion channel regulator, was first identified as a protein highly expressed in murine breast tumor initiated by oncogenic ras (v-Ha-ras) or neu (her2, egfr2) [1][2][3][4][5][6][7][8][9][10][11][12][13] and later demonstrated to occur at (inner-) cellular membranes and to be involved in the regulation of chloride conductance. [11][12][13] Recent studies found FXYD3 to be overexpressed breast, prostate, and pancreatic cancers, 5,14,15 and suggested that it promotes carcinogenesis by providing a growth advantage.…”