PT  - JOURNAL ARTICLE
AU  - Jun Adachi
AU  - Akemi Kakudo
AU  - Yoko Takada
AU  - Junko Isoyama
AU  - Narumi Ikemoto
AU  - Yuichi Abe
AU  - Ryohei Narumi
AU  - Satoshi Muraoka
AU  - Daigo Gunji
AU  - Yasuhiro Hara
AU  - Ryohei Katayama
AU  - Takeshi Tomonaga
TI  - Systematic identification of ALK substrates by integrated phosphoproteome and interactome analysis
AID  - 10.26508/lsa.202101202
DP  - 2022 Aug 01
TA  - Life Science Alliance
PG  - e202101202
VI  - 5
IP  - 8
4099  - https://www.life-science-alliance.org/content/5/8/e202101202.short
4100  - https://www.life-science-alliance.org/content/5/8/e202101202.full
SO  - Life Sci. Alliance2022 Aug 01; 5
AB  - The sensitivity of phosphorylation site identification by mass spectrometry has improved markedly. However, the lack of kinase–substrate relationship (KSR) data hinders the improvement of the range and accuracy of kinase activity prediction. In this study, we aimed to develop a method for acquiring systematic KSR data on anaplastic lymphoma kinase (ALK) using mass spectrometry and to apply this method to the prediction of kinase activity. Thirty-seven ALK substrate candidates, including 34 phosphorylation sites not annotated in the PhosphoSitePlus database, were identified by integrated analysis of the phosphoproteome and crosslinking interactome of HEK 293 cells with doxycycline-induced ALK overexpression. Furthermore, KSRs of ALK were validated by an in vitro kinase assay. Finally, using phosphoproteomic data from ALK mutant cell lines and patient-derived cells treated with ALK inhibitors, we found that the prediction of ALK activity was improved when the KSRs identified in this study were used instead of the public KSR dataset. Our approach is applicable to other kinases, and future identification of KSRs will facilitate more accurate estimations of kinase activity and elucidation of phosphorylation signals.