# Defining the pathways activated by Toll-like Receptors to stimulate immunity

> **NIH NIH R37** · BOSTON CHILDREN'S HOSPITAL · 2021 · $531,000

## Abstract

Project Summary
The goal of this proposal is to identify how Toll-like Receptors (TLRs) stimulate diverse cellular responses in
macrophages and dendritic cells (DCs), and to understand how these responses influence DC-based cancer
immunotherapies. The ability of TLRs to induce inflammatory gene expression has been under investigation for
twenty years, with distinct signaling pathways mediated by the MyD88 and TRIF adaptors explaining all
transcriptional responses. It has only recently become appreciated that TLRs also drive metabolic changes in
responding cells, such as the rapid induction of aerobic glycolysis. During the previous funding period, we
discovered that the TLR-induced myddosome complex contains two classes of proteins. One class is necessary
for myddosome assembly (e.g. MyD88) and represents the core of this signaling structure. The second class is
not necessary for myddosome assembly (e.g. TRAF6), but rather operates to recruit enzymes that diversify the
effector functions of the myddosome. Specifically, we identified the kinase TBK1 as a myddosome component
that is recruited by TRAF6 and is dedicated specifically to induce glycolysis. The myddosome therefore serves
as a subcellular site of signals that activate diverse cellular responses. Our understanding of how these activities
are regulated in vitro and their impact on T cell mediated protective immunity remains limited.
In addition to the myddosome, select TLRs (e.g. TLR4 and TLR3) engage the triffosome. The central triffosome
regulator is TRIF, which stimulates interferon (IFN) responses, NF-kB and MAPK activation, necroptosis and
glycolysis. While the importance of TRIF in immunity has long-been recognized, the means by which it activates
these diverse responses is unclear. This gap in knowledge is not merely an academic curiosity, as TRIF is
essential for the ability of the LPS receptor TLR4 to stimulate adaptive immunity. Understanding regulatory
events that stimulate myddosome- and TRIF-dependent responses will enable discussions of how TLRs drive
protective immunity against infection and cancer. In this application, we propose to explore myddosome
activities in vitro and in the context of cancer immunotherapies (Aim 1). In Aim 2, we offer an innovative synthetic
biology-based approach to define the mechanisms of TRIF signaling and how these mechanisms relate to those
induced by complementary innate immune pathways. Our focus on the two major signaling pathways activated
by TLRs should provide an operational view of this important family of receptors.

## Key facts

- **NIH application ID:** 10209029
- **Project number:** 2R37AI116550-05A1
- **Recipient organization:** BOSTON CHILDREN'S HOSPITAL
- **Principal Investigator:** JONATHAN C KAGAN
- **Activity code:** R37 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $531,000
- **Award type:** 2
- **Project period:** 2016-12-13 → 2026-01-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10209029

## Citation

> US National Institutes of Health, RePORTER application 10209029, Defining the pathways activated by Toll-like Receptors to stimulate immunity (2R37AI116550-05A1). Retrieved via AI Analytics 2026-06-01 from https://api.ai-analytics.org/grant/nih/10209029. Licensed CC0.

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