title: Proteomic analysis of novel targets associated with TrkA-mediated tyrosine phosphorylation signaling pathways in SK-N-MC neuroblastoma cells.
abstract: Tropomyosin-related kinase A (TrkA) is a receptor-type protein tyrosine kinase and exploits pleiotypic roles via nerve growth factor (NGF)-dependent or NGF-independent mechanisms in various cell types. Here, we showed that the inhibition of TrkA activity by GW441756 resulted in the suppression of tyrosine phosphorylation of cellular proteins including extracellular signal-regulated protein kinase (ERK) and c-Jun N-terminal kinase (JNK). To find novel targets associated with TrkA-mediated tyrosine phosphorylation signaling pathways, we investigated GW441756 effects on TrkA-dependent targets in SK-N-MC neuroblastoma cells by proteomic analysis. The major TrkA-dependent protein spots controlled by GW441756 were determined by PDQuest image analysis, identified by MALDI-TOF MS and MALDI-TOF/TOF MS/MS, and verified by 2DE/Western blot analysis. Thus, we found that most of the identified protein spots were modified forms in a normal condition, and their modifications were regulated by TrkA activity. Especially, our results demonstrated that the modifications of α-tubulin and heterogeneous nuclear ribonucleoproteins C1/C2 (hnRNP C1/C2) were significantly upregulated by TrkA, whereas α-enolase modification was downregulated by TrkA, and it was suppressed by GW441756, indicating that TrkA activity is required for their modifications. Taken together, we suggest here that the major novel TrkA-dependent targets such as α-tubulin, hnRNP C1/C2, and α-enolase could play an essential role in TrkA-mediated tyrosine phosphorylation signaling pathways via regulation of their posttranslational modifications.
authors: Department of Biochemistry and Institute of Health Sciences, Gyeongsang National University School of Medicine, Jinju, South Korea.
authors: Eun Joo
authors: EJ
authors: Jung
authors: Sang-Yeul
authors: SY
authors: Lee
authors: Choong Won
authors: CW
authors: Kim
mesh_terms: D015153:Blotting, Western; D045744:Cell Line, Tumor; D015536:Down-Regulation / Q000187:drug effects; D015180:Electrophoresis, Gel, Two-Dimensional; D034482:Heterogeneous-Nuclear Ribonucleoprotein Group C / Q000378:metabolism; D006801:Humans; D058990:Molecular Targeted Therapy; D009447:Neuroblastoma / Q000188:drug therapy / Q000201:enzymology / Q000378:metabolism*; D010751:Phosphopyruvate Hydratase / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D047428:Protein Kinase Inhibitors / Q000494:pharmacology*; D020543:Proteome / Q000032:analysis / Q000378:metabolism; D040901:Proteomics / Q000379:methods*; D020917:Receptor, trkA / Q000037:antagonists & inhibitors / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D053719:Tandem Mass Spectrometry; D014404:Tubulin / Q000378:metabolism; D015854:Up-Regulation / Q000187:drug effects
chemical_list: D034482:Heterogeneous-Nuclear Ribonucleoprotein Group C; D047428:Protein Kinase Inhibitors; D020543:Proteome; D014404:Tubulin; D020917:Receptor, trkA; D010751:Phosphopyruvate Hydratase