Utilization of liquid chromatography mass spectrometry analyses to identify LKB1-APC interaction in modulating Wnt/β-catenin pathway of lung cancer cells.
STK11/LKB1, a serine/threonine protein kinase and tumor suppressor, is a key upstream kinase of adenine monophosphate-activated protein kinase, which is a kinase involved in controlling cell polarity and maintaining cellular energy homeostasis. LKB1 is mutated in a significant number of Peutz-Jeghers syndrome (PJS) cases and sporadic cancers, and is most frequently mutated in lung adenocarcinomas; however, little is known about how LKB1 is involved in lung cancer progression. In this study, immunoprecipitation-HPLC tandem mass spectrometry (IP-LC-MS/MS) was performed to identify novel proteins interacting with LKB1 in lung cancer. Interestingly, many LKB1-interacting proteins acquired from the LC-MS/MS approach were mapped, using MetaCore pathway analysis, to the cystic fibrosis transmembrane conductance regulator activation pathway. Moreover, it was determined that LKB1 directly interacts with APC, and this LKB1-APC interaction was further confirmed by reverse immunoprecipitation assays, but GSK3β was dispensable for the association of LKB1 and APC. Importantly, LKB1 binds to APC to suppress the Wnt/β-catenin signaling pathway, which is known to be involved in cell proliferation and migration. Subsequent analysis of the downstream targets of the Wnt/TCF pathway led to the identification of several Wnt-regulated genes, such as CD44, COX-2, survivin, and c-Myc, whose expression levels are downregulated by LKB1. In summary, these results demonstrate that LKB1 regulates the Wnt pathway through a direct interaction with APC to suppress the tumorigenic/metastatic potential of lung tumors.


CONCLUSIONS
LKB1 status influences the molecular circuitry (Wnt/β-catenin pathway), cellular biology, and may serve as a potential therapeutic node in genetically defined subsets of lung cancer.
Institute of Biomedical Sciences, National Sun Yat-Sen University, Kaohsiung 80424, Taiwan. khcheng@faculty.nsysu.edu.tw.
Shu-Fang
SF
Jian
Chang-Chun
CC
Hsiao
Shin-Yi
SY
Chen
Ching-Chieh
CC
Weng
Tzu-Lei
TL
Kuo
Deng-Chyang
DC
Wu
Wen-Chun
WC
Hung
Kuang-Hung
KH
Cheng
D000091162:AMP-Activated Protein Kinase Kinases; D025601:Adenomatous Polyposis Coli Protein / Q000032:analysis / Q000378:metabolism*; D000818:Animals; D049109:Cell Proliferation; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D057809:HEK293 Cells; D064593:Heterografts; D006801:Humans; D008175:Lung Neoplasms / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D016513:Mice, SCID; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000032:analysis / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000032:analysis / Q000378:metabolism; D015398:Signal Transduction; D053719:Tandem Mass Spectrometry / Q000379:methods*; D060449:Wnt Signaling Pathway; D051176:beta Catenin / Q000378:metabolism*
C555337:APC protein, human; D025601:Adenomatous Polyposis Coli Protein; C495270:CTNNB1 protein, human; D034741:RNA, Small Interfering; D051176:beta Catenin; D017346:Protein Serine-Threonine Kinases; C109790:STK11 protein, human; D000091162:AMP-Activated Protein Kinase Kinases