Proteomic profiling of DC

We have compiled a comprehensive protein profile of DCs and obtained data on changes in protein level and post-translational modifications involved in the regulation of specific signalling pathways or cell processes.

First, we have set up a system for SILAC labelling of bone marrow derived murine dendritic cells (BM-DC). The principal method for generating BM-DC with GM-CSF was adapted from Lutz et al. (J Immunol Methods 1999, 223: 77-92). After some modifications of the protocol the FACS analysis of DC, which were generated in conditions described by Lutz et al. or in modified SILAC conditions did not show differences in the expression levels of the common DC markers CD11c, CD80, CD86. Analysis by mass spectrometry showed successful labelling. For the investigation of chemokine dependent CCR7 signalling, we have stimulated mature DCs with CCL19 or CCL21 for different time points. Western Blot analysis of phosphotyrosines showed a chemokine dependent and stimulation-time dependent phosphorylation pattern. We are now identifying and quantifying these chemokine dependent tyrosine phosphorylation events by mass spectrometry-based proteomics, as established in the optimization steps described above.

• organ type
• human bone marrow

• molecule type
• CD197,
• CCL19,
• CCL21,
• CD11c,
• CD86,
• CD80

• cell type
• dendritic cell

• organism type
• Mus musculus

created over 16 years ago (2 March 2009)    last modified over 12 years ago (9 July 2012)   [ RDF ]   [ RelFinder ]