# Regulation of the Salmonella Pathogenicity Island 1 Type III Secretion System via the hilD 3' untranslated region

> **NIH NIH R21** · UNIVERSITY OF ILLINOIS AT URBANA-CHAMPAIGN · 2023 · $186,820

## Abstract

PROJECT SUMMARY
The foodborne pathogen Salmonella is an important model organism for understanding genetic regulation
and bacterial pathogenesis. A requisite for Salmonella to cause disease is the direct injection of effector
proteins into host cells via a Type Three Secretion System (T3SS) encoded on Salmonella Pathogenicity
Island 1 (SPI1). This critical virulence factor is controlled in response to a plethora of environmental and
regulatory signals that dictate expression of the system at the proper time and place in the host. Our long-
term goal is to understand overall signal integration that allows this precise regulation. The SPI1 regulatory
circuit is controlled by three AraC-like regulators, HilD, HilC, and RtsA, which act in a complex feed-forward
regulatory loop to control expression of hilA, encoding the direct regulator of the SPI1 structural genes.
Much of the regulatory input is integrated at the level of HilD, including at hilD mRNA translation or stability.
The hilD mRNA has an unusual 300 nucleotide 3’ untranslated region (UTR) that acts as an independent
module to confer instability to the mRNA. A primary hypothesis is that the hilD 3’ UTR serves as a critical
node for integration of regulatory signals. Preliminary data show that mRNA stability is regulated by a
novel mechanism involving interaction between Rho-mediated transcriptional termination at the 3’ UTR
and RNase E-dependent degradation. Moreover, these activities are independently controlled by sRNAs.
The first aim of this proposal is to identify sRNAs and cis-acting sites that regulate via the hilD 3’ UTR.
Interacting sRNAs will be identified using an unbiased molecular technique, with base pairing confirmed
by mutagenesis. Deletion analysis will identify the site of Rho action in the 3’ UTR. The resulting hilD
mRNAs with mutations in sRNA binding sites or Rho-utilization site provide tools for further mechanistic
analyses. The second aim is to characterize the mechanism of post-transcriptional regulation via the hilD
3' UTR. The roles of Rho, RNase E, and the small RNAs in the creation and/or processing of the 3’ ends
in the hilD 3’ UTR will be monitored using tagging and deep sequence analysis. In vitro transcription will
more precisely define the action of Rho in creating terminated hilD transcripts. The interactions of these
factors will reveal the mechanistic details of this novel regulation. The SP1 T3SS regulatory circuit serves
as a paradigm for understanding the integration of host environmental signals to control a complex
virulence phenotype. Analysis of this system is critical to our understanding of this important pathogen.

## Key facts

- **NIH application ID:** 10625450
- **Project number:** 5R21AI166495-02
- **Recipient organization:** UNIVERSITY OF ILLINOIS AT URBANA-CHAMPAIGN
- **Principal Investigator:** JAMES M. SLAUCH
- **Activity code:** R21 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2023
- **Award amount:** $186,820
- **Award type:** 5
- **Project period:** 2022-05-20 → 2024-04-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10625450

## Citation

> US National Institutes of Health, RePORTER application 10625450, Regulation of the Salmonella Pathogenicity Island 1 Type III Secretion System via the hilD 3' untranslated region (5R21AI166495-02). Retrieved via AI Analytics 2026-05-24 from https://api.ai-analytics.org/grant/nih/10625450. Licensed CC0.

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