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. 2002 Mar;22(6):1778–1791. doi: 10.1128/MCB.22.6.1778-1791.2002

FIG. 7.

FIG. 7.

The N terminus of SAP97 is important for its basolateral membrane localization. MDCK cells stably expressing YFP-tagged full-length SAP97 (a to f), YFP-SAP97ΔN (g to l), and YFP-SAP97(1-158) (m to r) were stained with anti-CASK antibodies (c, i, o, f, l, and r). Cells were then stained with secondary antibodies coupled to fluorochromes and examined by confocal laser scanning microscopy. YFP-tagged protein localization was observed at an excitation wavelength of 513 nm and an emission wavelength 527 nm (a, g, m, d, j, and p). Squares, digital photomicrographs (the X-Y dimension of the Z series); rectangles, X-Z dimension (Z section). The apical side of the membrane is on the top of the Z sections and the basolateral sides are at the bottom. (B) The N-terminus-deleted SAP97 does not interact with mLin-2/CASK. HEK 293 cells were cotransfected with HA-tagged full-length mLin-2/CASK and either full-length YFP-tagged SAP97 or YFP-tagged N-terminus-deleted SAP97. Lysates and proteins immunoprecipitated (IP) with a YFP antibody were resolved on a 4 to 12% Novex gel and transferred to a nitrocellulose membrane. Subsequently, the membrane was probed with an anti-HA antibody to detect coprecipitated HA-mLin-2/CASK (top), stripped, and then probed with an anti-YFP antibody to detect both full-length YFP-SAP97 and N-terminus-deleted YFP-SAP97 (bottom).