The Journal of Neuroscience, September 27, 2006, ():
Vestibular Hair Bundles Control pH with (Na+, K+)/H+ Exchangers NHE6 and NHE9
J. Neurosci. Hill et al.
26: 9944
Supplemental data
Files in this Data Supplement:
- supplemental material
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Supplemental Figure 1. ClustalW alignment of mouse NHE9 with NHE1 and NHE6-8.
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Supplemental Figure 2. Antibody against NHE9 detects a single band in frog brain. Proteins of microsomes expressing mouse NHE9 fused to GFP (0.24 µg protein) and frog brain (2.3 mg wet weight) were separated by SDS-PAGE, transferred to a blotting membrane, and detected with anti-NHE9. As noted in the text, NHE9 runs anomalously during SDS-PAGE; the fastest migrating major GFP-NHE9 band is at ~65 kD; the band detected in frog brain runs at ~40 kD, consistent with NHE9 lacking the ~25 kD GFP tag.
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Supplemental Figure 3. pH dependence of salt-sensitive yeast growth. Yeast strain AX lacking NHX1 and NHA1 was transformed with vector alone or with plasmids expressing yeast Nhx1, mutant Y361F Nhx1, NHE6 or NHE9. Strains were grown in APG medium adjusted to the pH indicated and supplemented with 667 mM KCl. Unlike yeast Nhx1, mammalian NHE6 and NHE9 only improve salt-sensitive growth over the AX control above pH 7 (*).
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Supplemental Figure 4. Model for cation flow in hair cells. Transduction channels (yellow) let in K+ as the main current-carrying ion but also admit substantial amounts of Ca2+. Ca2+ is removed from stereocilia by the PMCA2v/a (frog) or PMCA2w/a (mammal) Ca2+ pumps; one H+ is transported into stereocilia for every Ca2+ transported out. H+ entering stereocilia are proposed to be removed by NHE6 and NHE9 Na+(K+)/H+-exchangers. K+ entering by counter-transport will be removed by basolateral K+ channels; the K+ flux attributable to K+/H+-exchange will be minimal compared to direct entry via transduction channels. NHE1 maintains H+ homeostasis on hair-cell basolateral membranes.
