FIGURE 11.

Molecular interaction between L-PGDS and MARCKS. NIH3T3 cells were treated with buffer as a control or L-PGDS protein (1 μg/ml) for 24 h, and cross-linked using 1% formaldehyde. Proteins cross-linked with L-PGDS were immunoprecipitated followed by SDS-PAGE separation and silver staining (A). Coimmunoprecipitated proteins with anti-L-PGDS antibody were identified by LC-MS/MS. In a separate experiment, NIH3T3 fibroblast cells or mixed glial cells treated with L-PGDS protein (100 ng/ml) were immunoprecipitated with anti-L-PGDS antibody. Immunoprecipitates were run on a SDS-PAGE gel and analyzed for the expression of MARCKS by Western blot analysis (B). α-Tubulin in input was also detected as a control. Immunoprecipitation with recombinant protein G-agarose alone without L-PGDS antibody was also used as a control, where no specific band was detected (data not shown). The results are representative of more than three independent experiments. WB, Western blot.