Mitotic phosphotyrosine network analysis reveals that tyrosine phosphorylation regulates Polo-like kinase 1 (PLK1)

Caron, Danielle; Byrne, Dominic P.; Thebault, Philippe; Soulet, Denis; Landry, Christian R.; Eyers, Patrick A.; Elowe, Sabine

doi:10.1126/scisignal.aah3525

Cited by 29 publications

(28 citation statements)

References 82 publications

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“…While phosphomimetic Eg5 is an imperfect substitution for phosphorylated protein, generating cells with hyper‐phosphorylated Eg5 is not trivial. Mitosis involves a complicated and inter‐connected network of kinase signaling that is highly regulated (Caron et al, ). Simply over‐expressing c‐Src kinase would not guarantee that Eg5 is hyper‐phosphorylated at Y211 and the interpretation of spindle phenotypes would be complicated by the effect of c‐Src overactivation on other mitotic targets, potentially including other mitotic kinases.…”

Section: Resultsmentioning

confidence: 99%

“…Their dysregulation also contributes to oncogenesis (Kim, Song, & Haura, ) and recent data points to a new role for SFKs in regulating spindle establishment and orientation (Nakayama et al, ). Other recent work suggests that phosphotyrosine (pTyr) modifications are more prevalent than previously appreciated, particularly in the kinetochore/spindle region and particularly by SFKs (Caron et al, ). To date, however, few mitotic SFK targets have been identified and none of them are known to regulate the MT cytoskeleton (Bhatt, Erdjument‐Bromage, Tempst, Craik, & Moasser, ; Fumagalli, Totty, Hsuan, & Courtneidge, ; Wang, Chen, Ding, Jin, & Liao, ).…”

Section: Introductionmentioning

confidence: 99%

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Src family kinase phosphorylation of the motor domain of the human kinesin‐5, Eg5

et al. 2017

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Spindle formation in mammalian cells requires precise spatial and temporal regulation of the kinesin-5, Eg5, which generates outward force to establish spindle bipolarity. Our results demonstrate that Eg5 is phosphorylated in cultured cells by Src family kinases (SFKs) at three sites in the motor head: Y125, Y211, and Y231. Mutation of these sites diminishes motor activity in vitro, and replacement of endogenous Eg5 with phosphomimetic Y211 in LLC-Pk1 cells results in monopolar spindles, consistent with loss of Eg5 activity. Cells treated with SFK inhibitors show defects in spindle formation, similar to those in cells expressing the non-phosphorylatable Y211 mutant, and distinct from inhibition of other mitotic kinases. We propose that this phosphoregulatory mechanism tunes Eg5 enzymatic activity for optimal spindle morphology.

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Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Src family kinase phosphorylation of the motor domain of the human kinesin‐5, Eg5

et al. 2017

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“…The PLK family are stringently regulated multifaceted modulators of mitosis and cytokinesis (Combes et al, 2017), and the close regulatory relationship between PLK1 and Aurora A in mitosis (Macurek et al, 2008, Scutt et al, 2009) led us to investigate potential redox regulation for both PLK1 and PLK4. (Caron et al, 2016a). We demonstrated dose-dependent activation and inhibition of PLK1 by DTT and H 2 O 2 respectively (Fig.…”

Section: Cys-dependent Redox Regulation Of Aurora A-related Kinasesmentioning

confidence: 57%

“…The kinase activation segment (Fig. 5A, top) contains multiple conserved residues available for post-translational phosphorylation in protein kinases (McSkimming et al, 2016, Caron et al, 2016b, Wilson et al, 2018, Cobbaut et al, 2017, and serves as a critical regulatory structure for the modulation of catalytic activity (Nolen et al, 2004). The activation loop of Aurora A itself undergoes dynamic conformational changes in response to phosphorylation and interactions with allosteric binding-partners, enabling Aurora A to transition between alternate active states (Levinson, 2018).…”

Section: The Cys-containing Regulatory Activation Segment In Epksmentioning

confidence: 99%

An evolutionary-conserved redox regulatory mechanism in human Ser/Thr protein kinases

Byrne

Shrestha

Kannan

et al. 2019

Preprint

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ONE-SENTENCE SUMMARY:The catalytic activity of Ser/Thr kinases is regulated through a conserved Cys-based redox mechanism. ABSTRACT:Reactive oxygen species (ROS) are products of oxygen metabolism, but are also recognized as endogenous physiological mediators of cellular signaling. Eukaryotic protein kinase (ePK) regulation occurs through reversible phosphorylation events in the flexible activation segment. In this study, we demonstrate that the catalytic phosphotransferase output from the mitotic Ser/Thr kinase Aurora A is also controlled by cysteine (Cys) oxidation. Reversible regulation occurs by direct modification of a conserved residue (Cys 290), which lies adjacent to Thr 288, the activating site of phosphorylation. Strikingly, redox modulation of the Cys 290-equivalent in other ePKs is predicted to be an underappreciated regulatory mechanism, since ~100 human Ser/Thr kinases possess a Cys at this position in the conserved activation loop. Using real-time enzyme assays, we confirm that the presence of the equivalent Cys residue is prognostic for redox-sensitivity amongst a cohort of human CAMK, AGC and AGC-like kinases, including AKT, AMPK, CAMK1, MAPKAP-K2/3 and SIK1-3. Our findings demonstrate that dominant Cys-based redox-switching in the activation segment represents an evolutionary-conserved mode of regulation for a significant subset of the human kinome. This finding has important implications for understanding physiological and pathological signaling responses to ROS, and emphasises the importance of multivalent activation segment regulation in ePKs.

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“…1 Our integrated approach revealed extensive Tyr phosphorylation during mitosis, particularly at the spindle and associated structures. We first determined the localization of pTyr signals in mitotic HeLa cells and the non-cancerous immortalized RPE1 cells, and we verified the presence of prominent spindle-associated Tyr phosphorylation during mitosis using a panel of different anti-pTyr antibodies.…”

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confidence: 99%

Tyr(less) kinase signaling during mitosis

Elowe

2017

Cell Cycle

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Tyrosine phosphorylation is rare, representing only about 0.5% of phosphorylations in the cell under basal conditions. While mitogenic tyrosine kinase signaling has been extensively explored, the role of phosphotyrosine signaling across the cell cycle and in particular during mitosis is poorly understood.Two recent, independent studies tackled this question from different angles to reveal exciting new insights into the role of this modification during cell division. Caron et al. 1 exploited mitotic phosphoproteomics data sets to determine the extent of mitotic tyrosine phosphorylation, and St-Denis et al. 2 identified protein tyrosine phosphatases from all subfamilies as regulators of mitotic progression or spindle formation. These studied collectively revealed that tyrosine phosphorylation may play a more prominent and active role in mitotic progression than previously appreciated. Protein phosphorylation is well recognized as a major mode of post-translational regulation. The human genome encodes over 23,000 proteins; more than two-thirds phosphorylated. 3 Phosphorylation is a crucial component of signal transduction, regulating pathways that control processes from cell division to cell death. At the molecular level, reversible phosphorylation can lead to changes in protein structure and stability, protein-protein interactions, enzyme activation, or subcellular localization. Phosphorylation is most dynamic during the mitotic stage of the cell cycle, 4 and many mitotic phosphoproteome studies have resulted in a wealth of data on serine (Ser) and threonine (Thr) phosphorylation events. However, phosphotyrosine (pTyr) sites are relatively rare, and are underrepresented even in large scale mitosis-specific phosphoproteome studies. Seminal work from many groups in the 1980s, 1990s and early 2000s has revealed remarkable insight into how tyrosine (Tyr) phosphorylation regulates many aspects of growth and development. It is an integral component of cell surface signaling by receptors such as receptor tyrosine kinases, and integrins which signal in part via intracellular kinases such as the Src-family kinases (SFKs). 5 These cell surface signals are then propagated throughout the cytoplasm and nucleus through signaling networks often involving feedbacks and crosstalks to finally elicit the appropriate response. A number of tissue-specific pTyr screens have been reported (e.g. 6-8 ) but little is known about Tyr phosphorylation dynamics throughout the cell cycle, leaving a significant gap in our understanding of cell cycle control.The best understood pTyr site regulated during mitosis is arguably the inhibitory phosphorylation of cyclin-dependent kinase (CDK1) at Tyr, 15 which is catalyzed by WEE1 and MYT1, and whose dephosphorylation serves as the basis for the switch-like activity of CDK1/cyclin B at mitotic entry. 9 Tyr phosphorylation at the spindle was clearly demonstrated almost 20 years ago when pTyr-containing epitopes, detected by the monoclonal 4G10 pTyr antibody, were observed at kinetochores and cen...

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Mitotic phosphotyrosine network analysis reveals that tyrosine phosphorylation regulates Polo-like kinase 1 (PLK1)

Cited by 29 publications

References 82 publications

Src family kinase phosphorylation of the motor domain of the human kinesin‐5, Eg5

Src family kinase phosphorylation of the motor domain of the human kinesin‐5, Eg5

An evolutionary-conserved redox regulatory mechanism in human Ser/Thr protein kinases

Tyr(less) kinase signaling during mitosis

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