Extended Data Fig. 7: Cdk5 expression is induced by educated astrocytes but not cancer-associated fibroblasts or microglial cells.
A. In vitro cultured parental and paired BrM-seeking cells or patient-derived xenografts were probed for CDK5. B. Representative bright-field photograph of primary mouse astrocytes cultured in vitro. C. mRNA expression of the indicated genes in primary mouse astrocytes. N = 4 technical replicates for all genes except for Trem2, s100b, and Ndrg2 (N = 3 technical replicates). D-F. Indicated cell lines were cultured in CM from CAFs (D), BV-2 microglial cells (E) or primary mouse microglia (F) and probed for Cdk5 at indicated timepoints. G,H. EO771.GFP cells were directly co-cultured with CAFs (G), BV-2 microglial cells (G) or primary mouse microglia (H), GFP-sorted at indicated timepoints, and probed for Cdk5. I,J. 4T1 cells were treated as indicated followed by immunoblotting for Cdk5. K. Cytokine array of CM from primary mouse astrocytes, naïve or pre-educated by 4T1.Br3 CM. L. Pre-proteomics verification of educated astrocytes CM’s ability to induce Cdk5 in 4T1 cells by immunoblotting. M. Indicated cell lines were cultured in the presence of recombinant Versican at the indicated doses for 4 h or 8 h, followed by Cdk5 immunoblotting. N. 4T1 cell lysates were immunoprecipitated by Emilin-1 antibody or IgG, and immunoprecipitation efficiency was evaluated by Emilin-1 immunoblotting. PDX – patient-derived xenografts, CAF – cancer-associated fibroblast, CM – conditioned media, EV – extracellular vesicles, IgG – Immunoglobulin G.
