Figure 2: Parkin interacts with GluK2.
From: Parkin regulates kainate receptors by interacting with the GluK2 subunit
(a) Representative western blottings of co-immunoprecipitations in HEK293T cells transfected with FLAG-parkin and Myc-GluK2a. The data show that FLAG-parkin co-immunoprecipitates with Myc-GluK2a and that Myc-GluK2a co-immunoprecipitates with FLAG-parkin. The image is representative of five independent experiments. (b) Representative western blottings of co-immunoprecipitations between endogenous parkin and GluK2 in whole mouse brain lysates and whole human brain lysates. The data show that GluK2 co-immunoprecipitates with parkin. The image is representative of three independent experiments. (c) Western blottings of co-immunoprecipitation experiments from HEK293T cells transfected with FLAG-parkin and Myc-GluK2a mutants truncated at aa 896, 868 and 850. Our terminology for these truncation mutants indicates the position at which a stop codon was introduced. All the mutants were able to bind parkin. The data show that the parkin-GluK2a interaction is unaffected by the loss of the last 58 aa of the GluK2a tail. The image is representative of four independent experiments. (d) Pull-down assay using biotinylated peptides spanning the GluK2a C terminus and lysates from human brain or HEK293T cells expressing FLAG-parkin. GluK2a peptides that bind parkin are indicated in red. The image is representative of five independent experiments. (e) Pull-down assay using biotinylated GluK2a peptides and lysates from HEK293T cells expressing FLAG-parkin or amino-terminally flagged parkin domains (FLAG-Ubl-linker; FLAG-R1; FLAG-R2). GluK2a peptides pulled down FLAG-Ubl-linker of parkin. The image is representative of five independent experiments.
