# Role of ROP16 in Toxoplasma gondii Cyst Development

> **NIH NIH F31** · UNIVERSITY OF ARIZONA · 2021 · $46,036

## Abstract

Abstract:
In humans, clinically relevant disease with Toxoplasma gondii, a common intracellular parasite, results from
T.gondii’s tropism for and life-long persistence in the CNS. To persist, T.gondii switches from a fast replicating
form (tachyzoite) to a slow replicating, encysted form (bradyzoite). While T.gondii infection is asymptomatic in
most, T. gondii can cause severe neurologic complications in individuals with incomplete immune responses
(e.g. AIDS patients, developing fetuses) including cognitive, visual and motor deficits, and death. The treatments
for symptomatic toxoplasmosis, most commonly pyrimethamine and sulfadiazine, are poorly tolerated, have low
therapeutic indices, and are ineffective against the persistent, encysted form. The development of therapies
targeting T. gondii’s encysted form requires a deep, mechanistic understanding of how parasites encyst. Prior
studies have focused on identifying parasite genes that drive the tachyzoite/bradyzoite transition and parasite
proteins that form the cyst wall. Less work has been done on identifying host cell genes that influence
encystment, and almost no work has been done on how parasite manipulations of the host cell affect encystment.
The goal of this proposal is to address this gap by building upon my preliminary work showing that a well-known
kinase (ROP16)– which parasites injects into the cytoplasm of host cells and which shows allelic variation
between the canonical T. gondii strains (type I, II, and III)– facilitates encystment in a strain-specific manner.
Type I and III alleles (rop16I/III/ROP16I/III), which are 99% identical, cause prolonged activation of STAT3, 6, and
possibly 5, signaling pathways during acute tachyzoite infection, while the type II allele does not. To determine
if ROP16 play a role in encystment, I generated type II and type III strains that lack ROP16 (IIΔrop16 and
IIIΔrop16) and tested them in an in vitro encystment assay. Remarkably, in contrast to IIΔrop16 which showed
little change in encystment, IIIΔrop16 has a greater than 2-fold decrease in forming cysts in fibroblasts and
neurons. In addition, I found that this defect can be complemented in trans by co-infecting a host cell with
IIIΔrop16 and parental type III (WTIII) parasites, but not with type II parasites, suggesting a role for strain-specific
host cell manipulations. My overall hypothesis, therefore, is that ROP16III facilitates cyst development through
strain-specific host cell manipulations. I will address this hypothesis by: i) determining which ROP16III functions
are required for encystment by complementing the IIIΔrop16 strain with a panel of ROP16 mutants that lack
kinase activity, nuclear localization, or STAT-binding and assessing encystment (Aim 1) and ii) identifying the
ROP16III-dependent host genes that drive encystment by transcriptionally profiling cells in encysted conditions
and infected with WTIII, IIIΔrop16, or complemented parasites (Aim 2). The completion of this work...

## Key facts

- **NIH application ID:** 10106451
- **Project number:** 5F31AI147711-02
- **Recipient organization:** UNIVERSITY OF ARIZONA
- **Principal Investigator:** Joshua Andrew Kochanowsky
- **Activity code:** F31 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $46,036
- **Award type:** 5
- **Project period:** 2020-02-01 → 2022-01-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10106451

## Citation

> US National Institutes of Health, RePORTER application 10106451, Role of ROP16 in Toxoplasma gondii Cyst Development (5F31AI147711-02). Retrieved via AI Analytics 2026-05-22 from https://api.ai-analytics.org/grant/nih/10106451. Licensed CC0.

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