Phosphoscan Analysis of Mantle Cell Lymphoma Cell Lines
A Zamo, TL Gu, RD Polakiewicz, A Parisi, A Bertolaso, S Barbi, G Inghirami, F Menestrina. University of Verona, Verona, Italy; Cell Signaling Technology, Inc., Danvers, MA; University of Torino, Turin, Italy
Background: Mantle cell lymphoma (MCL) constitutes about 5% of non-Hodgkin lymphoma in the western world. It shows a characteristic clinical behaviour with initial response to therapy followed by relapses and death in 3-5 years. Many studies have investigated MCL pathogenesis by high-throughput techniques such as gene expression profiling, but very few have attempted the study of MCL from a proteomic point of view. In our previous work, we have focused our attention on the phospho-proteome of several MCL cell lines by IMAC pre-fractionation and 2D-PAGE/MS.
Design: In this work we used the PhosphoScan approach, which has already been successfully used in the phospho-profiling of lung cancer and acute myeloid leukemia. This involves a protein extraction followed by a digestion. Phosphorylated peptides are then immunoprecipitated by anti-phosphotyrosine antibody followed by capillary electrophoresis and MS identification of eluted peptides. We analysed four MCL cell lines (MAVER-1, Jeko-1, Rec-1 and Granta-519).
Results: 421 unique peptides were identified, corresponding to 341 proteins. Of these, 191 were present in at least two cell lines, and were further analysed for KEGG pathway. This analysis showed that the most represented ones are Regulation of actin cytoskeleton (16 proteins, 9.04%), Focal adhesion (15 proteins, 8.47%), Fc epsilon RI signaling pathway (14 proteins, 7.91%) and B cell receptor signaling pathway (13 proteins, 7.34%). The four pathways are strongly interconnected, and suggest the activation of the B-cell receptor in these cell lines. The absence of the antigen during in vitro culture suggests that these cell have a tonic BCR signalling either by abnormalities of the signalling pathway or by self-stimulation.
Conclusions: The BCR signalling pathway is promising from a therapeutic point of view, and recent data indicate that it might be very important in the pathogenesis of DLBCL. Its importance in MCL has not been investigated so far. PhosphoScan analysis of MCL cell lines suggests that tonic BCR signalling might be one of the mechanisms driving cell survival and proliferation in MCL, but these data need to be validated by functional studies and by the extensive analysis of tissue samples.
Category: Special Category for 2009 - Pan-genomic/Pan-proteomic approaches to Cancer
Tuesday, March 10, 2009 9:30 AM
Poster Session III # 222, Tuesday Morning