# GBPs as new inflammasome regulators during mammalian host defense

> **NIH NIH R01** · YALE UNIVERSITY · 2020 · $418,750

## Abstract

PROJECT SUMMARY/ABSTRACT
Inflammasomes alert the mammalian immune system to the presence of infection and tissue damage. These
cytosolic protein complexes detect danger signals or microbial products released by a wide variety of intracellular
pathogens. In the case of bacterial pathogens, a number of prokaryotic signatures are recognized including the
major cell-wall constituent of most Gram-negative species, lipopolysaccharide (LPS). Detection of LPS inside
host cells activates a “non-canonical“ inflammasome pathway where caspase-11 (Caspases 4 and 5 in humans)
act as upstream sensors to stimulate inflammasome complex assembly and processing of the pore-forming
protein, Gasdermin D (Gsdmd), further downstream. Gsdmd pores release protective cytokines and contribute
to a lytic form of cell death termed pyroptosis that may help eliminated infected host cells. How these sequential
events are co-ordinated and the host factors involved remains a major question in the field of innate immunity
and host defense. Here, we focus on members of a new 65-73kDa immune GTPase family termed Guanylate-
Binding Proteins (GBPs) that control distinct steps in the non-canonical pathway. Preliminary results suggest
Gbp2 may target cytosolic bacteria to help liberate LPS for caspase-11 detection whereas Gbp3 acts further
downstream to regulate Gsdmd trafficking to the plasma membrane. GBPs thus offer a unique opportunity to
understand how this sequential hierarchy unfolds. In Aim 1, we will test the respective contributions of Gbp2 and
Gbp3 to immunity against Gram-negative Salmonella typhimurium (Stm) infection via the non-canonical
inflammasome in vitro and in vivo. CRISPR-Cas9 deleted human and mouse cells as well as newly-created
Gbp2-/-, Gbp3-/- and GbpDchr.3H1 mice will be infected with Stm variants designed to interfere with GBP recruitment
or responsiveness to LPS. Thereafter, we will dissect the molecular mechanisms enlisted by these GBPs to
confer their intracellular functions as part of Aim 2. Here gene-deficient macrophages complemented with GBP
mutants with distinct biochemical lesions will reveal how GBPs direct the inflammasome core machinery to LPS-
positive bacteria or control downstream events such as Gsdmd trafficking and assembly on the plasma
membrane. Cell-free studies will also attempt to reconstitute the GBP “coatomer” on the bacterial outer
membrane that serves as a platform for inflammasome assembly. Collectively, our proposal examines a new set
of host factors that act at different stages within the non-canonical signaling cascade as part of a unique
functional hierarchy, helping choreograph these events with major implications for the treatment of sepsis and
Gram-negative bacterial infections.

## Key facts

- **NIH application ID:** 9972734
- **Project number:** 2R01AI108834-06
- **Recipient organization:** YALE UNIVERSITY
- **Principal Investigator:** John David MacMicking
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $418,750
- **Award type:** 2
- **Project period:** 2014-09-19 → 2024-12-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/9972734

## Citation

> US National Institutes of Health, RePORTER application 9972734, GBPs as new inflammasome regulators during mammalian host defense (2R01AI108834-06). Retrieved via AI Analytics 2026-05-22 from https://api.ai-analytics.org/grant/nih/9972734. Licensed CC0.

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