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. 2012 Aug 22;32(34):11879–11889. doi: 10.1523/JNEUROSCI.1376-12.2012

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Copyright © 2012 the authors 0270-6474/12/3211879-11$15.00/0

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Figure 8. — Nope and Ppk25 interact specifically in cultured cells. A, HEK293 cells were transfected with plasmids encoding Nope-HA, Arm-HA, and, where indicated, Ppk25-Flag, and were immunoprecipitated using anti-Flag antibody. Extracts, supernatants (S), and precipitates (P) were analyzed on Western blots with anti-HA antibody. B, Same as in A, except that Nope was tagged with Flag and Ppk25 with HA. C, Extracts of cells expressing Nope-Flag, Ppk25-HA, and human β-ENaC-HA were immunoprecipitated with anti-Flag, and both supernatant and pellet were immunoblotted with anti-HA. The increased ratio of Ppk25-HA to β-ENaC-HA in the pellet compared with the supernatant indicates that Nope-Flag interacts preferentially with Ppk25-HA. D, Same as in C, except that Ppk25 was tagged with Flag and Nope with HA. Here also, the ratio of Nope-HA to β-ENaC-HA is higher in the pellet than in the supernatant.