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Assessing the activation/inhibition of tyrosine kinase‐related pathways with a newly developed platform
Author(s) -
Kagiwada Harumi,
Kiboku Takayuki,
Matsuo Hitomi,
Kitazawa Masashi,
Fukui Kazuhiko,
Horimoto Katsuhisa
Publication year - 2021
Publication title -
proteomics
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.26
H-Index - 167
eISSN - 1615-9861
pISSN - 1615-9853
DOI - 10.1002/pmic.202000251
Subject(s) - phosphorylation , signal transduction , microbiology and biotechnology , tyrosine phosphorylation , biology , kinase , epidermal growth factor , protein phosphorylation , receptor tyrosine kinase , phosphorylation cascade , protein kinase a , chemistry , biochemistry , receptor
The phosphorylation of cellular proteins plays a crucial role in the transduction of various signals from outside the cell into the nucleus. The signals are transduced by phosphorylation chain reactions within multiple pathways; however, determining which pathways are responsible for each defined signal has proven challenging. To estimate the activity of each pathway, we developed a phosphorylation array platform comprising a protein array with 1200 proteins belonging to 376 signalling pathways and an analytical method to estimate pathway activity based on the phosphorylation levels of proteins. The performance of our system was assessed by reconstructing kinase–substrate relationships, as well as by estimating pathway activity upon epidermal growth factor (EGF) stimulation and the pharmacological inhibition of epidermal growth factor receptor (EGFR). As a result, kinase–substrate relationships were reliably reconstructed based on the precise measurement of phosphorylation levels of constituent proteins on the array. Furthermore, the pathway activities associated with EGF stimulation and EGFR inhibition were successfully traced through the related pathways from the outer membrane to the nucleus along a time course. Thus, our phosphorylation array system can effectively assess the activity of specific signalling pathways that are perturbed by extracellular stimuli, such as various drugs.