Protein phosphorylation by kinases with its converse dephosphorylation by phosphatases regulates most biological processes. In human cells, more than three-quarters of cellular proteins are phosphorylated. The occupancy of a given phosphorylation site reflects the balance between the activities of kinases and phosphatases. To understand the reversible nature of protein phosphorylation, we must investigate the forward and reverse reaction by connecting kinases and phosphatases on their shared substrates of interest. Protein phosphorylation is catalyzed by more than 500 protein kinases, however, most protein dephosphorylation is carried out by only seven phosphoprotein phosphatases (PPPs). While there has been tremendous progress in deciphering cellular signaling by kinases, much less is known about phosphatases. Research in my laboratory is focused on integrating phosphatases into cellular signaling networks by establishing phosphatase-substrate relationships, identifying opposing kinases, and determining regulatory inputs. My laboratory is uniquely positioned to address these outstanding challenges by combining quantitative proteomics and phosphoproteomics approaches in cells with reconstitution of minimal signaling units in vitro. PPP form multimeric holoenzymes with overlapping subunits that function as distinct entities, which hampers mechanistic studies of holoenzyme specific functions and regulation in cells. My research program directly addresses this problem by studying PPP signaling networks on a system-wide level in cells and with isolated components in vitro. By combining in-cell discovery and in vitro validation and mechanistic reduction, we will distinguish direct from indirect effects or cellular compensation mechanisms and determine the contribution of specific holoenzymes. In this application we will focus on PP6, a PPP with limited subunit diversity that nonetheless accurately reflects the combinatorial nature of PPP holoenzymes. Collectively, these findings will further our understanding of holoenzyme specific phosphatase function, substrate preferences, and regulation, and connect phosphatase and kinase biology by integrating them into functional networks. Defining reciprocal PPP ? protein kinase regulation and opposition of shared substrates will provide insights into how physiological processes are controlled by reversible phosphorylation. We will continue to develop and implement innovative approaches in proteomics and cell biology to better address these and emerging questions. We envision this work to be a resource for the phosphorylation signaling community, as well as a framework for future research into phosphatase biology. We will share our data, reagents, and experimental approaches and generate an easily accessible database for the phosphatase substrates we will identify.

Public Health Relevance

Deregulation and mutations of protein kinases and phosphatases are commonly observed in human diseases, specifically cancer. Our research program is focused on establishing phosphatase-substrate relationships, identifying opposing kinases, and determining regulatory inputs. This knowledge will help us understand how disruption of the phosphorylation balance contributes to disease, and identify new drug targets and points of therapeutic intervention.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
Unknown (R35)
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Special Emphasis Panel (ZRG1)
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Barski, Oleg
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Dartmouth College
Schools of Medicine
United States
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Nasa, Isha; Kettenbach, Arminja N (2018) Coordination of Protein Kinase and Phosphoprotein Phosphatase Activities in Mitosis. Front Cell Dev Biol 6:30
Kettenbach, Arminja N; Schlosser, Kate A; Lyons, Scott P et al. (2018) Global assessment of its network dynamics reveals that the kinase Plk1 inhibits the phosphatase PP6 to promote Aurora A activity. Sci Signal 11:
Nasa, Isha; Rusin, Scott F; Kettenbach, Arminja N et al. (2018) Aurora B opposes PP1 function in mitosis by phosphorylating the conserved PP1-binding RVxF motif in PP1 regulatory proteins. Sci Signal 11:
Lee, Mid Eum; Rusin, Scott F; Jenkins, Nicole et al. (2018) Mechanisms Connecting the Conserved Protein Kinases Ssp1, Kin1, and Pom1 in Fission Yeast Cell Polarity and Division. Curr Biol 28:84-92.e4
Zhao, Yanding; Zurawel, Ashley A; Jenkins, Nicole P et al. (2018) Comparative Analysis of Mutant Huntingtin Binding Partners in Yeast Species. Sci Rep 8:9554
Shee, Kevin; Yang, Wei; Hinds, John W et al. (2018) Therapeutically targeting tumor microenvironment-mediated drug resistance in estrogen receptor-positive breast cancer. J Exp Med 215:895-910
Choy, Meng S; Swingle, Mark; D'Arcy, Brandon et al. (2017) PP1:Tautomycetin Complex Reveals a Path toward the Development of PP1-Specific Inhibitors. J Am Chem Soc 139:17703-17706
Cullati, Sierra N; Kabeche, Lilian; Kettenbach, Arminja N et al. (2017) A bifurcated signaling cascade of NIMA-related kinases controls distinct kinesins in anaphase. J Cell Biol 216:2339-2354
Dumitru, Ana Maria G; Rusin, Scott F; Clark, Amber E M et al. (2017) Cyclin A/Cdk1 modulates Plk1 activity in prometaphase to regulate kinetochore-microtubule attachment stability. Elife 6:
Orr, Amy; Song, Hongki; Rusin, Scott F et al. (2017) HOPS catalyzes the interdependent assembly of each vacuolar SNARE into a SNARE complex. Mol Biol Cell 28:975-983

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