Materials
Recombinant human M-CSF was purchased from R&D Systems (Minneapolis, MN). Ro-31-8220, Gö-6976, Rotterin, and BAPTA/AM were obtained from Calbiochem (San Diego, CA). Low endotoxin RPMI and X-vivo serum free medium were obtained from Lonza Walkersville Inc (Walkersville, MD). Fetal bovine serum (FBS, certified <0.06 endotoxin units/ml endotoxin levels) was purchased from Atlanta Biological (Lawrenceville, GA). All other cell culture reagents were obtained through Invitrogen (Carlsbad, CA). PKCα NF-κB p65, Lamin B, β-actin, and GAPDH antibodies, anti-mouse and rabbit IgG-HRP conjugated antibodies, and PKCα siRNA were obtained from Santa Cruz Biotechnology (Santa Cruz, CA). Phospho-NFκB p65 (Ser276 or Ser536) antibodies were purchased from Cell Signaling Technology (Danvers, MA). DNA constructs pTAL-SEAP and pNF-κB-SEAP were obtained from Clontech (Mountian View, CA). Expression vectors containing CMV-p65 WT, CMV-p65 276S/A and CMV-p65 536S/A were cloned in pBa5e Puro vector or pFLAG-CMV-2 vector as described previously [49]. Kinase deficient CMV-PKCα (KD-PKCα) or wild type pCMV-PKCα (WT-PKCα) were generous gifts from Alexandra C. Newton (University of California, San Diego, CA). IκBα (NFKBIA) and GAPDH primers for real-time RT-PCR were obtained from SABiosciences (Frederick, MD). A pool of mouse or human PKCα specific siRNA were purchased from Santa Cruz Biotechnology (Santa cruz, CA).