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. 2017 May 12;13(5):e1006791. doi: 10.1371/journal.pgen.1006791

Fig 2. NPHP5 directly interacts with USP9X.

Fig 2

A) Flag or Flag-NPHP5 was expressed in HEK293 cells. Lysates were immunoprecipitated with anti-Flag antibody and Western blotted with the indicated antibodies. IN, input. B) (Top) IRCM1 and IRCM2 are two anti-NPHP5 antibodies raised against the C- and the N-terminus of NPHP5, respectively. Grey box, calmodulin-binding domain; white box, coiled-coil domain; black box; Cep290-binding domain. (Bottom) HEK293 lysates were immunoprecipitated with anti-Flag (control), anti-NPHP5 (IRCM1 or IRCM2) or anti-USP9X antibody and Western blotted with the indicated antibodies. IN, input. C) The ability of full-length NPHP5 (1-598(FL)) and various NPHP5 truncates to interact with USP9X is presented. D) Flag (control) or the indicated fragment of Flag-tagged NPHP5 was expressed in HEK293 cells. Lysates were immunoprecipitated with anti-Flag antibody and Western blotted with the indicated antibodies. IN, input. Asterisks indicate bands corresponding to the expected recombinant proteins. E) Purified GFP, GFP-USP9X wild type or catalytically inactive mutant (C1566S) bound to beads was mixed with purified Flag-NPHP5. Proteins recovered on beads were analyzed by Western blotting with the indicated antibodies. IN, input.