Identification of Dynamic Proteome Changes Upon Ligand Activation of Trk-Receptors Using Two-dimensional Fluorescence Difference Gel Electrophoresis and Mass Spectrometry

• DOI: 10.1074/mcp.m400188-mcp200
• Publication Date: 2005-01-16T01:12:49Z
• Authors: Barbara Sitek; Ognjan Apostolov; Kai Stühler; Kathy Pfeiffer; Helmut E. Meyer; Angelika Eggert; Alexander Schramm
• Keywords: Trk receptor; Difference gel electrophoresis

The TrkA and TrkB tyrosine kinases are members of the neurotrophin receptor family mediate survival, differentiation, growth, apoptosis neurons in response to stimulation by their ligands, NGF BDNF, respectively. Expression levels TrkA/TrkB important prognostic factors a variety embryonal tumors including neuroblastoma, most common solid tumor childhood. Because exhibit high level sequence similarity use overlapping pathways for signal transduction, existence specific effector molecules crucial cell-type-specific is likely. To identify these effectors analyzing biological effects activation defined model, we performed proteome study using human neuroblastoma SY5Y cell line stably transfected with or cDNA. recently introduced DIGE (fluorescence two-dimensional difference gel electrophoresis) system (Amersham Biosciences, Piscataway, NJ) allowed us monitor differences protein expression between samples one gel. Proteomic changes were monitored time course 0, 0.5, 1, 6, 24 h following activation. Using MALDI mass spectrometry, identified, respectively, 22 9 differentially expressed proteins upon addition SY5Y-TrkB SY5Y-TrkA cells. Functional assignment revealed that majority involved organization maintenance cellular structures.