# Elucidation of the Role of Chlamydial ClpX During Development and Differentiation

> **NIH NIH F31** · UNIVERSITY OF NEBRASKA MEDICAL CENTER · 2022 · $31,523

## Abstract

PROJECT SUMMARY
 Chlamydia trachomatis is the leading cause of bacterial sexually transmitted infection both domestically
and globally, with the rate of cases per 100,000 increasing every year. However, infected individuals often fail to
seek treatment largely due to the asymptomatic nature of approximately 70% of infections. Unchecked, chronic
infection can lead to numerous sequelae, including ectopic pregnancy and tubal factor infertility in women or
epididymitis and sterility in men, and those who do receive treatment are typically prescribed broad spectrum
antibiotics. Given the steadily increasing case rate of infections, the risk of chronic issues, and the limited options
for antibiotic stewardship, a more comprehensive understanding of chlamydial biology to produce a more
targeted therapeutic treatment is critical.
 Despite having a highly reduced genome, Chlamydia undergoes a complex developmental cycle in which
the bacteria differentiate between two functionally and morphologically distinct forms: the infectious, non-
replicative elementary body (EB) and the non-infectious, replicative reticulate body (RB). An EB initiates infection
by binding to and inducing uptake into a host cell. Within the cell, the EB undergoes primary differentiation into
an RB, which will then give rise to a population of RBs. At an unknown signal mid-developmental cycle, newly
formed RBs will undergo secondary differentiation from RB to EB. The transitions between EBs and RBs are not
mediated by division events that re-distribute intracellular proteins. Rather, both primary (EB to RB) and
secondary (RB to EB) differentiation likely require protein turnover. As such, we hypothesize that ClpX plays
a critical role during chlamydial differentiation through targeted protein degradation. To test this
hypothesis, we generated numerous constructs for overexpression of various mutant ClpX isoforms in
Chlamydia. The overarching goal of this proposal is to delineate a mechanism of ClpX-mediated degradation of
substrates through adaptor dependent or independent means, which may uncover a network of targets that
contribute to chlamydial differentiation. Previous in vitro characterization of these mutations from other labs
studying other bacterial ClpX paralogs allows us to take a more targeted assessment of ClpX function rather
than standard alanine scanning. Aim 1 will determine in which facets of chlamydial biology ClpX plays a role by
observing the effects that these mutant proteins exert on chlamydial development, differentiation state, and
morphology. The proposed experiments of Aim 2 will identify targets of chlamydial ClpX, which will provide
significant insight into its substrate recognition and processing. We have developed a chemical crosslinking
approach combined with Click-iT labeling of chlamydial proteins, specifically, to maintain complex stability when
performing affinity purification of ClpX and its adaptors and substrates. Collectively, these experiments w...

## Key facts

- **NIH application ID:** 10480772
- **Project number:** 5F31AI164815-02
- **Recipient organization:** UNIVERSITY OF NEBRASKA MEDICAL CENTER
- **Principal Investigator:** Nicholas A Wood
- **Activity code:** F31 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2022
- **Award amount:** $31,523
- **Award type:** 5
- **Project period:** 2021-07-21 → 2023-05-05

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10480772

## Citation

> US National Institutes of Health, RePORTER application 10480772, Elucidation of the Role of Chlamydial ClpX During Development and Differentiation (5F31AI164815-02). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/10480772. Licensed CC0.

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