Interactions between β-Catenin and the HSlo Potassium Channel Regulates HSlo Surface Expression

Abstract

BackgroundThe large conductance calcium-activated potassium channel alpha-subunit (Slo) is widely distributed throughout the body and plays an important role in a number of diseases. Prior work has shown that Slo, through its S10 region, interacts with β-catenin, a key component of the cytoskeleton framework and the Wnt signaling pathway. However, the physiological significance of this interaction was not clear.Methodology/Principal FindingsUsing a combination of proteomic and cell biology tools we show the existence of additional multiple binding sites in Slo, and explore in detail β-catenin interactions with the S10 region. We demonstrate that deletion of this region reduces Slo surface expression in HEK cells, which indicates that interaction with beta-catenin is important for Slo surface expression. This is confirmed by reduced expression of Slo in HEK cells and chicken (Gallus gallus domesticus leghorn white) hair cells treated with siRNA to β-catenin. HSlo reciprocally co-immunoprecipitates with β-catenin, indicating a stable binding between these two proteins, with the S10 deletion mutant having reduced binding with β-catenin. We also observed that mutations of the two putative GSK phosphorylation sites within the S10 region affect both the surface expression of Slo and the channel's voltage and calcium sensitivities. Interestingly, expression of exogenous Slo in HEK cells inhibits β-catenin-dependent canonical Wnt signaling.Conclusions and SignificanceThese studies identify for the first time a central role for β-catenin in mediating Slo surface expression. Additionally we show that Slo overexpression can lead to downregulation of Wnt signaling.

Galectin-3 gene silencing inhibits migration and invasion of human tongue cancer cells in vitro via downregulating β-catenin

“…A close correlation between Gal-3 expression and malignant metastasis/histological grade in tongue cancer has been demonstrated [12] , as well as a close correlation between Gal-3 and β-catenin in tumors [9,25] . These data suggest that β-catenin plays a key role in the invasion of several types of cancer [3][4][5][6][7][8][9][10][13][14][15][16][17] . In the SCC-4 and CAL27 cells, β-catenin was significantly inhibited at the protein level, but it was unaffected at the mRNA level.…”

“…β-Catenin enhances cancer cell adhesion and migration [13] via the Wnt/β-catenin pathway or other signaling pathways (SDF-1/CXCR4 axis [14] and the HSIo potassium channel [15] ). Colon cancer metastasis is associated with activation of the Wnt/β-catenin signaling pathway through the expression www.chinaphar.com Zhang D et al Acta Pharmacologica Sinica npg of the metastasis mediator S100A4 [16] .…”

Hair cell BK channels interact with RACK1, and PKC increases its expression on the cell surface by indirect phosphorylation

“…Similarly, we have shown ␤ 1 and ␤ 4 subunits that are expressed in low-frequency hair cells to decrease surface expression of Slo in these hair cells (9). Furthermore, we and others have shown phosphorylation to affect Slo surface expression (8,11,43). Both CDK5 and GSK3 bind to Slo and decrease its surface expression.…”

“…Both antibodies identified bands of the expected size with minimal additional bands. For experiments using PMA and monensin, chick cochlea were kept in culture in the presence of PMA, monensin, or DMSO (control) and labeled Slo detected as described previously (11) in hair cells ϳ1.5 mm from the apical end.…”

“…Both CDK5 and GSK3 bind to Slo and decrease its surface expression. CDK5 mediates its effects by direct phosphorylation while the effects of GSK3 phosphorylation are also mediated by interactions with ␤-catenin, which, in turn, promotes Slo surface expression (8,11,43). In other work we have inferred gradients in kinase activity along the tonotopic axis with increasing PKA in low-frequency hair cells and increasing PKC activity in high-frequency hair cells (21).…”