# Regulation of heterotrimeric G protein signaling by subunit phosphorylation

> **NIH NIH R01** · PENNSYLVANIA STATE UNIVERSITY, THE · 2021 · $77,492

## Abstract

Summary
Heterotrimeric G protein signaling pathways are of tremendous importance to human health. Mutation of G
protein subunits causes genetic disease, developmental abnormalities, and altered infectious disease
susceptibility. Indeed, G protein pathways are the targets of approximately a third of all drugs under clinical use.
The Assmann laboratory has furthered fundamental understanding of heterotrimeric G protein signaling through
elucidation of G protein-mediated signaling cascades and phenotypes. The emphasis of the parent award is on
mechanisms of phospho-regulation of G protein signaling that are evolutionarily conserved but have been oft
overlooked in mammalian systems. In this research, the model plant Arabidopsis is used as a facile system to
investigate kinase-mediated phosphorylation of the canonical G protein α (Gα) subunit, GPA1, and to explicate
the downstream signaling impacts of this phosphorylation; in particular, how phosphorylation biases interactions
with downstream effector proteins. In parallel, relevance to human health is demonstrated through assessment
of the impacts of analogous phosphorylation events on human Gα subunits in vitro, using BODIPY-GTP binding
and hydrolysis assays on the corresponding human phosphomimic mutants. This supplement requests funding
for the purchase of a BioTek Synergy Neo2 plate reader to increase the throughput and reliability of the BODIPY-
GTP activity assays and to allow the implementation of orthogonal methods, particularly transcreener assays,
that will allow independent validation of these biochemical data. The dual monochromators of the Synergy Neo2
plate reader will allow for assay versatility with a multitude of fluorophores. The speed and sensitivity of the
Synergy Neo2 far outpaces that of the extant obsolete Flx800 plate reader, allowing for finer timescale
measurements of a greater number of Gα variants. In addition, the capability of the Synergy Neo2 to assay
luciferase activity will facilitate assessment of biased signaling arising from phosphorylation-dependent protein-
protein interactions, as the Synergy Neo2 has the capability for high-throughput measurements of protein-protein
interaction using the split-luciferase method. In summary, the proposed state-of-the-art plate reader will provide
reliable, sensitive, and rapid data acquisition as well as entirely new capabilities to probe the molecular effects
of G protein phosphorylation. Finally, the instrumentation will be available to other NIH-funded researchers at
Penn State in the Biology and Chemistry Departments.

## Key facts

- **NIH application ID:** 10387072
- **Project number:** 3R01GM126079-04S1
- **Recipient organization:** PENNSYLVANIA STATE UNIVERSITY, THE
- **Principal Investigator:** Sarah M Assmann
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $77,492
- **Award type:** 3
- **Project period:** 2018-04-01 → 2023-02-28

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10387072

## Citation

> US National Institutes of Health, RePORTER application 10387072, Regulation of heterotrimeric G protein signaling by subunit phosphorylation (3R01GM126079-04S1). Retrieved via AI Analytics 2026-05-25 from https://api.ai-analytics.org/grant/nih/10387072. Licensed CC0.

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