# Discovery and characterization of a novel family of GTPase activating proteins

> **NIH NIH F32** · CORNELL UNIVERSITY · 2024 · $74,284

## Abstract

PROJECT SUMMARY
Small GTPases of the Arf and Rab families are master regulators of intracellular trafficking in eukaryotic
cells that direct the key trafficking events of vesicle formation, cargo recognition, and transport. Cells exert
tight spatiotemporal control of these GTPases through a network of GTPase effector proteins (GEFs) and
GTPase activating proteins (GAPs). Perturbations to these key regulators give rise to numerous human
diseases. Despite their importance, many GEFs and GAPs have yet to be identified — meaning we lack a
comprehensive understanding of how cells manage the flow of materials between different membrane-
bound compartments. There is a critical need, then, to study the modulators of GTPase activity and
determine their roles in membrane homeostasis. The overarching goal of this proposal is to identify and
characterize novel regulators of GTPase activity. I have recently identified a novel family of GAPs using
structural prediction approaches. The specific aims of this proposal are to characterize the physiological role
of these new GAPs. Using a combination of genetic and live-cell imagining techniques, Specific Aim 1 will
focus on identifying the target substrate for each of the GAPs. I will knock out and overexpress the novel
GAPs and subsequently monitor fluorescently-tagged GTPases. The rationale being that improper
regulation of GTPases will cause their mislocalization. Aim 1 will also determine how these proteins localize
to distinct subcellular locations and how this ultimately drives their regulation of GTPases. Together, this
specific aim will elucidate the biological niches in which each of these proteins function. Specific Aim 2 will
investigate the structure and function of these novel GAPs to better understand how they act to regulate
GTPase activity. Using in vitro GAP assays, the enzymatic activity of purified GAPs will be reconstituted on
synthetic membranes — quantifying their activity and substrate specificity in vitro. In order to validate my
structural predictions, the structure of GAPs in complex with their substrate GTPase will be determined
using cryogenic electron microscopy. Crucially, these structural studies will involve proteins bound to
synthetic membranes, providing unique insights into the molecular architecture of membrane-bound
GTPase inactivation. Completion of the research in this fellowship proposal will provide valuable insight into
the regulation of the secretory pathway and, by extension, will have broad impact in cell and membrane
biology.

## Key facts

- **NIH application ID:** 10995673
- **Project number:** 1F32GM155980-01
- **Recipient organization:** CORNELL UNIVERSITY
- **Principal Investigator:** Ryan Christopher Vignogna
- **Activity code:** F32 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2024
- **Award amount:** $74,284
- **Award type:** 1
- **Project period:** 2024-07-01 → 2026-06-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10995673

## Citation

> US National Institutes of Health, RePORTER application 10995673, Discovery and characterization of a novel family of GTPase activating proteins (1F32GM155980-01). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/10995673. Licensed CC0.

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