# Tracking the dynamics of the macrophage response to interferon-gamma at a single-cell level

> **NIH NIH F31** · UNIVERSITY OF CALIFORNIA, SAN DIEGO · 2022 · $39,681

## Abstract

Project Summary
Precise temporal regulation of inflammatory responses is required to clear infection without damaging healthy
tissue. Previous work elucidating the mechanisms involved in this regulation have focused mainly on single
time points or bulk samples of cells. However, immune cells in vivo receive complex temporal combinations of
stimuli during an immune response, respond with gene expression patterns that vary over time, and display
heterogeneity within the population. Interferon gamma (IFNγ) is a pro-inflammatory cytokine that plays key
roles in immune responses. Macrophages are immune cells that are one of the primary responders to IFNγ.
During infection, macrophages may experience multiple periods of IFNγ stimulation, and employ signaling and
gene expression networks to decode these varying stimuli into gene expression responses and diverse
functions. GBP1 and NOS2 are two IFNγ-responsive genes that have important roles in host defense against
microbes and are regulated by different network architectures and chromatin regulatory mechanisms.
Mycobacterium tuberculosis (Mtb) infection is a pressing global health issue that also depends critically on
macrophage responses to IFNγ. Mtb infection outcomes are heterogeneous on the cellular as well as the
organismal (human) level. Previous work has shown that IFNγ signaling is essential for macrophages to kill
intracellular Mtb and that this ability to kill Mtb varies between cells. This proposal uses a system that
combines endogenous fluorescent gene reporters in macrophage cell lines with long-term live-cell imaging in a
microfluidic device to simultaneously track expression kinetics of multiple genes in response to dynamic
stimuli, as well as the outcomes of Mtb infection, in the same single cells over time. This can be used to obtain
a quantitative understanding of the mechanisms underlying kinetic gene expression responses and functional
heterogeneity. In Aim 1, this system is used to quantify and model single macrophage gene expression
kinetics following dynamic IFNγ stimulus and to elucidate the mechanism of signal decoding. This is done by
applying IFNγ stimulus of varying amplitude and duration to the macrophages and simultaneously tracking
expression kinetics of three components of the GBP1 and NOS2 networks in the same single cells over time. A
mathematical model will be developed to describe these responses, predict the response to perturbation, and
will be tested using inducible promoters and inhibitors of chromatin regulators to perturb the decoding. Aim 2
investigates the connection between cell-to-cell variability in gene expression kinetics and heterogeneous Mtb
infection outcomes. This is done by infecting fluorescent reporter macrophage cell lines with Mtb marked by a
viability reporter and assaying both gene expression kinetics and infection outcomes in single cells. The
completion of these aims will provide a quantitative understanding of the mechanisms by which macroph...

## Key facts

- **NIH application ID:** 10388046
- **Project number:** 1F31AI161903-01A1
- **Recipient organization:** UNIVERSITY OF CALIFORNIA, SAN DIEGO
- **Principal Investigator:** Beverly Naigles
- **Activity code:** F31 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2022
- **Award amount:** $39,681
- **Award type:** 1
- **Project period:** 2022-01-01 → 2024-12-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10388046

## Citation

> US National Institutes of Health, RePORTER application 10388046, Tracking the dynamics of the macrophage response to interferon-gamma at a single-cell level (1F31AI161903-01A1). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/10388046. Licensed CC0.

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