Interplay between Polo kinase, LKB1-activated NUAK1 kinase, PP1βMYPT1 phosphatase complex and the SCFβTrCP E3 ubiquitin ligase.
NUAK1 (NUAK family SnF1-like kinase-1) and NUAK2 protein kinases are activated by the LKB1 tumour suppressor and have been implicated in regulating multiple processes such as cell survival, senescence, adhesion and polarity. In the present paper we present evidence that expression of NUAK1 is controlled by CDK (cyclin-dependent kinase), PLK (Polo kinase) and the SCFβTrCP (Skp, Cullin and F-boxβTrCP) E3 ubiquitin ligase complex. Our data indicate that CDK phosphorylates NUAK1 at Ser445, triggering binding to PLK, which subsequently phosphorylates NUAK1 at two conserved non-catalytic serine residues (Ser476 and Ser480). This induces binding of NUAK1 to βTrCP, the substrate-recognition subunit of the SCFβTrCP E3 ligase, resulting in NUAK1 becoming ubiquitylated and degraded. We also show that NUAK1 and PLK1 are reciprocally controlled in the cell cycle. In G2-M-phase, when PLK1 is most active, NUAK1 levels are low and vice versa in S-phase, when PLK1 expression is low, NUAK1 is more highly expressed. Moreover, NUAK1 inhibitors (WZ4003 or HTH-01-015) suppress proliferation by reducing the population of cells in S-phase and mitosis, an effect that can be rescued by overexpression of a NUAK1 mutant in which Ser476 and Ser480 are mutated to alanine. Finally, previous work has suggested that NUAK1 phosphorylates and inhibits PP1βMYPT1 (where PP1 is protein phosphatase 1) and that a major role for the PP1βMYPT1 complex is to inhibit PLK1 by dephosphorylating its T-loop (Thr210). We demonstrate that activation of NUAK1 leads to a striking increase in phosphorylation of PLK1 at Thr210, an effect that is suppressed by NUAK1 inhibitors. Our data link NUAK1 to important cell-cycle signalling components (CDK, PLK and SCFβTrCP) and suggest that NUAK1 plays a role in stimulating S-phase, as well as PLK1 activity via its ability to regulate the PP1βMYPT1 phosphatase.
*MRC Protein Phosphorylation and Ubiquitylation Unit, College of Life Sciences, University of Dundee, Dow Street, Dundee DD1 5EH, U.K.
Sourav
S
Banerjee
†Molecular Neurobiology Laboratory, Salk Institute for Biological Studies, La Jolla, CA 92037, U.S.A.
Anna
A
Zagórska
*MRC Protein Phosphorylation and Ubiquitylation Unit, College of Life Sciences, University of Dundee, Dow Street, Dundee DD1 5EH, U.K.
Maria
M
Deak
*MRC Protein Phosphorylation and Ubiquitylation Unit, College of Life Sciences, University of Dundee, Dow Street, Dundee DD1 5EH, U.K.
David G
DG
Campbell
‡Division of Cell Signalling and Immunology, College of Life Sciences, University of Dundee, Dow Street, Dundee DD1 5EH, U.K.
Alan R
AR
Prescott
*MRC Protein Phosphorylation and Ubiquitylation Unit, College of Life Sciences, University of Dundee, Dow Street, Dundee DD1 5EH, U.K.
Dario R
DR
Alessi
D000091162:AMP-Activated Protein Kinase Kinases; D000595:Amino Acid Sequence; D000818:Animals; D002453:Cell Cycle / Q000235:genetics*; D018797:Cell Cycle Proteins / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D005786:Gene Expression Regulation; D057809:HEK293 Cells; D006801:Humans; D008969:Molecular Sequence Data; D043263:Myosin-Light-Chain Phosphatase / Q000235:genetics* / Q000378:metabolism; D011485:Protein Binding; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D011494:Protein Kinases / Q000235:genetics* / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism; D012097:Repressor Proteins / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism; D044843:SKP Cullin F-Box Protein Ligases / Q000235:genetics* / Q000378:metabolism; D016415:Sequence Alignment; D015398:Signal Transduction; D044767:Ubiquitin-Protein Ligases / Q000235:genetics* / Q000378:metabolism; D000097869:Polo-Like Kinase 1
D000091162:AMP-Activated Protein Kinase Kinases; D018797:Cell Cycle Proteins; D043263:Myosin-Light-Chain Phosphatase; D047428:Protein Kinase Inhibitors; D011494:Protein Kinases; D017346:Protein Serine-Threonine Kinases; D011518:Proto-Oncogene Proteins; D012097:Repressor Proteins; D044843:SKP Cullin F-Box Protein Ligases; D044767:Ubiquitin-Protein Ligases; D000097869:Polo-Like Kinase 1; C470845:NUAK1 protein, human; C109790:STK11 protein, human; C490372:PPP1R12A protein, human