Pervanadate induces Mammalian Ste20 Kinase 3 (MST3) tyrosine phosphorylation but not activation.
The yeast Ste20 (sterile) protein kinase, which is a serine/threonine kinase, responds to the stimulation of the G proteincoupled receptor (GPCR) pheromone receptor. Ste20 protein kinase serves as the critical component that links signaling from the GPCR/G proteins to the mitogen-activated protein kinase (MAPK) cascade in yeast. The yeast Ste20p functions as a MAP kinase kinase kinase kinase (MAP4K) in the pheromone response. Ste20-like kinases are structurally conserved from yeast to mammals. The mechanism by which MAP4K links GPCR to the MAPK pathway is less clearly defined in vertebrates. In addition to MAP4K, the tyrosine kinase cascade bridges G proteins and the MAPK pathway in vertebrate cells. Mammalian Ste20 Kinase 3 (MST3) has been categorized into the Ste20 family and has been reported to function in the regulation of cell polarity and migration. However, whether MST3 tyrosine phosphorylation regulates diverse signaling pathways is unknown. In this study, the tyrosine phosphatase inhibitor pervanadate was found to induce MST3 tyrosine phosphorylation in intact cells, and the activity of tyrosine-phosphorylated MST3 was measured. This tyrosine-directed phosphorylation was independent of MST3 activity. Parameters including protein conformation, Triton concentration and ionic concentration influenced the sensitivity of MST3 activity. Taken together, our data suggests that the serine/threonine kinase MST3 undergoes tyrosinedirected phosphorylation. The tyrosine-phosphorylated MST3 may create a docking site for the structurally conserved SH2/SH3 (Src Homology 2 and 3) domains within the Src oncoprotein. The unusual tyrosinephosphorylated MST3 may recruit MST3 to various signaling components.
Department of Nephrology, Chi-Mei Medical Center, Tainan 701, Taiwan; Department of Medical Laboratory Science and Biotechnology, Chung Hwa University of Medical Technology, Tainan 701, Taiwan.
Wei-Chih
WC
Kan
School of Pharmacy, Tsuzuki Institute for Traditional Medicine, China Medical University, Taichung 404, Taiwan.
Te-Ling
TL
Lu
Department of Microbiology and Immunology, College of Medicine, National Cheng Kung University, Tainan 701, Taiwan.
Pin
P
Ling
Department of Medical Laboratory Science and Biotechnology, Chung Hwa University of Medical Technology, Tainan 701, Taiwan.
Te-Hsiu
TH
Lee
Department of Biochemistry and Molecular Biology, College of Medicine, National Cheng Kung University, Tainan 701, Taiwan.
Chien-Yu
CY
Cho
Institute of Biopharmaceutical Sciences, National Yang-Ming University, Taipei 112, Taiwan.
Chi-Ying F
CY
Huang
Department of Biochemistry and Molecular Biology, College of Medicine, National Cheng Kung University, Tainan 701, Taiwan.
Wen-Yih
WY
Jeng
Graduate Institute of Biological Science and Technology, Chung Hwa University of Medical Technology, Tainan, Taiwan.
Yui-Ping
YP
Weng
Department of Cardiology, Chi-Mei Medical Center, Tainan, Taiwan; Department of Optometry, Chung Hwa University of Medical Technology, Tainan, Taiwan.
Chun-Yen
CY
Chiang
School of Pharmacy, Tsuzuki Institute for Traditional Medicine, China Medical University, Taichung 404, Taiwan.
Jin Bin
JB
Wu
Department of Medical Laboratory Science and Biotechnology, Chung Hwa University of Medical Technology, Tainan 701, Taiwan. Electronic address: lutejung@yahoo.com.tw.
Te-Jung
TJ
Lu
D000595:Amino Acid Sequence; D000818:Animals; D004285:Dogs; D057075:Enzyme Assays; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D005786:Gene Expression Regulation; D057809:HEK293 Cells; D006801:Humans; D007700:Kinetics; D061985:Madin Darby Canine Kidney Cells; D009994:Osmolar Concentration; D010766:Phosphorylation / Q000187:drug effects; D011487:Protein Conformation; D017346:Protein Serine-Threonine Kinases / Q000235:genetics* / Q000378:metabolism; D015398:Signal Transduction; D014443:Tyrosine / Q000378:metabolism*; D014638:Vanadates / Q000494:pharmacology*; D000099317:Hippo Kinases
D004791:Enzyme Inhibitors; D017346:Protein Serine-Threonine Kinases; D014443:Tyrosine; D014638:Vanadates; C076715:pervanadate; C109163:STK24 protein, human; D000099317:Hippo Kinases
Ionic strength; Kinase assay; MST3; Triton; Tyrosine phosphorylation