# Proteomic interrogation of the parasite vacuole using autoSTOMP

> **NIH NIH R21** · UNIVERSITY OF VIRGINIA · 2021 · $242,250

## Abstract

Project Summary
 Toxoplasma gondii (Tg) is an incurable, obligate intracellular protozoan parasite that naturally infects
human and rodent hosts for life. Toxoplasma’s extraordinary ability to persist inside almost any nucleated cell
depends on an intact parasite vacuole membrane (PVM). Secreted parasite effectors localized to the PVM
interface subvert cell-intrinsic immune detection and import nutrients for growth. However, biochemical tools
have not been available to identify the complete sets of host and parasite components recruited to the PVM. To
recognize and clear Tg the host relies on two complementary cell autonomous immunity pathways. The
interferon-y regulated GTPases (IRG) system detects the PVM as foreign and Toll-like receptors (TLRs) prime
inflammasome cytosolic sensor recognition of Tg. Sub-cellular localization of the IRG system is a major point of
regulation, however, the precise mechanism of PVM recognition, Tg clearance and control of host cell fate
downstream of the IRG and TLR pathways are unclear. We hypothesize that immune recognition at the PVM
controls parasite fate and host cell survival. To identify the critical regulators of host and parasite survival at
the PVM we have developed a novel proteomics technology called automated spatially targeted optical micro
proteomics or autoSTOMP. AutoSTOMP uses the confocal microscope to image the PVM and attach photo-
activatable affinity purification tags to all PVM proteins so that they can be precipitated and identified by liquid
chromatography and mass spectrometry (LC-MS). This tool allows us to easily compare PVM localized proteins
across the three canonical types of Tg that differ in virulence by several logs. Here autoSTOMP will be used to
understand how immune stimulation regulates protein recruitment to the PVM and controls the fate of Tg and
host cells.

## Key facts

- **NIH application ID:** 10108771
- **Project number:** 1R21AI156153-01
- **Recipient organization:** UNIVERSITY OF VIRGINIA
- **Principal Investigator:** Sarah E. Ewald
- **Activity code:** R21 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $242,250
- **Award type:** 1
- **Project period:** 2020-11-23 → 2022-10-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10108771

## Citation

> US National Institutes of Health, RePORTER application 10108771, Proteomic interrogation of the parasite vacuole using autoSTOMP (1R21AI156153-01). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/10108771. Licensed CC0.

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