# Mechanisms of Phosphorylation Signaling by Phosphoprotein Phosphatases

> **NIH NIH R35** · DARTMOUTH COLLEGE · 2024 · $442,800

## Abstract

Abstract
 Reversible protein phosphorylation is a major regulatory mechanism that controls most cellular processes,
including mitosis. A ‘tug of war’ between kinases and phosphatases controls protein activity and function. The
precise coordination of both activities is essential for the accurate execution of cell division. Despite substantial
progress in deciphering kinase-mediated phosphorylation and its functional consequences, much less is known
about phosphatases and how dephosphorylation is regulated. The long-term goal of our research program is to
uncover the mechanisms that coordinate and integrate PPPs, kinases, and their shared substrates in the
signaling networks that control cell division.
 The majority of cellular dephosphorylation is carried out by the 7 catalytic phosphatase subunits of the
phosphoprotein phosphatase (PPP) family. Despite the apparent simplicity suggested by the small number of
catalytic PPP enzymes, complexity and specificity arise through the formation of holoenzymes. Each
holoenzyme functions as a distinct entity. The non-catalytic subunits modulate the activity of the catalytic
subunits, enabling substrate specificity, dictating subcellular localization, and ensuring appropriate regulation.
Combinatorially, there are several hundred functional PPP holoenzymes.
 There are key gaps in knowledge and challenges in studying PPP function and mechanisms of
dephosphorylation. Although hundreds of thousands of phosphorylated sites on proteins have been identified,
only ~450 have been matched to be dephosphorylated by a specific PPP holoenzyme. This is in part due to the
lack of holoenzyme-specific inhibitors and the complexity of PPP holoenzymes.
 To overcome these obstacles and fill these gaps in knowledge, we will (i) identify holoenzyme-specific
substrates; (ii) dissect mechanisms of substrate recruitment and site-specific dephosphorylation; and (iii)
determine regulatory inputs governing specific PPP holoenzymes. We have developed new cell biological and
biochemical approaches to investigate PPP signaling. We plan to map the PPP substrate space
comprehensively, providing a foundation for studying PPPs for our lab and the signaling community by sharing
our findings broadly and openly via a user-friendly database that we will maintain. The mechanisms that PPPs
use to recruit substrates and catalyze site-specific dephosphorylation are still emerging. We will continue to
dissect these mechanisms to predict and systematically dissect PPP-substrate relationships in cells. Finally,
determining regulatory inputs that govern PPP holoenzyme formation, activity, and function is needed to
understand their role in regulating mitosis. We combine quantitative measurements of the proteome and
phosphoproteome with the reconstitution of minimal signaling units in vitro and in cells to precisely determine
phosphatase function and regulation in cell division.

## Key facts

- **NIH application ID:** 10841603
- **Project number:** 5R35GM119455-09
- **Recipient organization:** DARTMOUTH COLLEGE
- **Principal Investigator:** Arminja Nadine Kettenbach
- **Activity code:** R35 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $442,800
- **Award type:** 5
- **Project period:** 2016-08-05 → 2026-05-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10841603

## Citation

> US National Institutes of Health, RePORTER application 10841603, Mechanisms of Phosphorylation Signaling by Phosphoprotein Phosphatases (5R35GM119455-09). Retrieved via AI Analytics 2026-05-29 from https://api.ai-analytics.org/grant/nih/10841603. Licensed CC0.

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