# T3SS Effector Regulation of Bacterial Metabolism

> **NIH NIH R21** · KANSAS STATE UNIVERSITY · 2022 · $186,498

## Abstract

Project Summary.
Many Gram-negative bacterial pathogens interact with mammalian cells by using secretion systems to inject
virulence proteins directly into infected host cells. Some of these injected protein ‘effectors’ are enzymes that
modify the structure and inhibit the function of mammalian proteins by catalyzing the addition of unusual post-
translational modifications. Type III secretion system (T3SS) effectors play essential roles in virulence and their
mechanisms have provided great insight into the functions and components of the innate immune system. T3SS
effectors are believed to be inactive until they are injected into host cells, where they then fold into their active
conformations. However, recent work with the NleB and SseK glycosyltransferases from E. coli, Citrobacter
rodentium, and Salmonella enterica has challenged that dogma. NleB glycosylates and activates the bacterial
glutathione synthetase (GshB) enzyme, resulting in enhanced glutathione production and improved C. rodentium
survival in oxidative stress conditions. SseK1 is active within Salmonella enterica, where it glycosylates and
enhances the activity of several enzymes that are critical to the ability of Salmonella to resist methylglyoxal
stress. In support of long-term goals to identify and understand the functional significance of bacterial protein
glycosylation by NleB and SseK1, two new bacterial targets of SseK1, namely NagC and CRP have been
discovered. NagC is a dual activator-repressor that controls GlcNAc uptake and metabolism. NagC also
regulates locus of enterocyte effacement (LEE) gene expression in enterohemorrhagic E. coli (EHEC). The LEE
is an important pathogenicity island that encodes the T3SS and many effector proteins. The catabolite repressor
protein [(CRP); also referred to as the catabolite activator protein (CAP)], is a global regulator that mediates the
expression of ~150 genes, including those important to GlcNAc metabolism and several T3SS components in
Salmonella. The central hypothesis to be tested is that Arg-glycosylation of NagC and CRP by SseK1 affects
Salmonella virulence gene regulation and metabolism. The specific aims are: 1) Quantify the extent to which
Arg-glycosylation of NagC and CRP affects the ability of these transcription factors to bind DNA; 2) Characterize
the impact of Arg-glycosylation of NagC and CRP on the Salmonella transcriptome. The proposed work is highly
suitable for the R21 funding mechanism because of the innovation of the research premise and novel hypothesis
to be tested. Such data will establish the framework for future investigation of the mechanistic aspects and
functional significance of T3SS effector regulation of bacterial transcription factors.

## Key facts

- **NIH application ID:** 10425770
- **Project number:** 1R21AI168453-01
- **Recipient organization:** KANSAS STATE UNIVERSITY
- **Principal Investigator:** Philip Ross Hardwidge
- **Activity code:** R21 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2022
- **Award amount:** $186,498
- **Award type:** 1
- **Project period:** 2022-04-22 → 2024-03-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10425770

## Citation

> US National Institutes of Health, RePORTER application 10425770, T3SS Effector Regulation of Bacterial Metabolism (1R21AI168453-01). Retrieved via AI Analytics 2026-05-28 from https://api.ai-analytics.org/grant/nih/10425770. Licensed CC0.

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