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. 2003 Jul;23(14):4917–4928. doi: 10.1128/MCB.23.14.4917-4928.2003

FIG. 2.

FIG. 2.

CD2AP interacts with p85. (A) myc.p85 was coexpressed with FLAG-tagged CD2AP (F.CD2AP) or the control protein GFP. After immunoprecipitation (IP) with anti-FLAG antibody, coprecipitating myc.p85 was detected with a p85-specific antibody (top). The middle and lower gels show protein expression in the lysates. (B and C) The interaction of p85 occurs with the amino-terminal portion of CD2AP, a region containing all three SH3 domains and a highly tyrosine-phosphorylated segment (top). The middle and lower gels show protein expression in the lysates. (D and E) To demonstrate endogenous interaction, freshly isolated kidneys were lysed in CHAPS lysis buffer and subjected to immunoprecipitation with control antibody (Ab) (anti-HA antibody) and anti-85 polyclonal antibody or anti-CD2AP polyclonal antibody. Prior to immunoprecipitation, the lysates were precleared extensively to ensure specific coprecipitation. Coprecipitating CD2AP (D) or p85 (E) protein was detected with a specific antiserum. (F) As previously shown, endogenous nephrin coprecipitated with CD2AP.