Fig. 1

Intracellular processing of PGRN. a Primary microglia from WT and Grn ^−/− mice were labeled with [³⁵S]methionine and [³⁵S]cysteine for 24 h. Cell lysates and media were immunoprecipitated by homemade rabbit anti-PGRN antibodies and separated by 16% Tricine-SDS PAGE. The PGRN and PGRN-derived peptide (GRNs) signals were visualized by autoradiography. * indicates non-specific bands. Please note there is a weak non-specific band that is the same size as full-length PGRN in both lysate and medium. b PGRN processing in MEF cells. Equal amounts of cell lysate from primary WT and Grn ^−/− MEF cells (left) and the rabbit anti-PGRN IP products from WT and Grn ^−/− MEF cells (right) were separated on 4–12% Bis-Tris gels and blotted with sheep anti-mouse PGRN antibodies (1:1000). * indicates non-specific bands. Please note that PGRN runs slightly differently on Tricine gels and Bis-Tris gels. c PGRN processing in mouse tissues. Equal amounts of tissue lysates were separated on a 4–12% Bis-Tris gel and blotted with sheep anti-mouse PGRN antibodies (1:1000). d Brain tissue from WT and Grn ^−/− adult mice were lysed with RIPA buffer at a ratio of 1:10 (g:ml) and an equal amount of protein was separated on a 4–12% Bis-Tris gel and immunoblotted with sheep anti-mouse PGRN antibodies (1:300). e Spleen tissues from WT and Grn ^+/− (Het) adult mice were lysed with RIPA buffer at a ratio of 1:10 (g:ml) and an equal amount of protein was separated on a 4–12% Bis-Tris gel and immunoblotted with sheep anti-mouse PGRN antibodies (1:1000). The ratios between granulin peptides (GRNs) and PGRN were quantified. ns: not significant, student’s T-test