# Non-conventional regulation of essential apicomplexan guanylate cyclases

> **NIH NIH K22** · UNIVERSITY OF OKLAHOMA HLTH SCIENCES CTR · 2020 · $108,000

## Abstract

Project Summary
Most protists are free-living, but some infectious forms reproduce exclusively from within host cells. One group
of obligate-intracellular protists constitute Apicomplexa, a phylum of globally-distributed parasites notorious for
inflicting severe disease and death in humans and livestock. Since pathogenesis stems from their lytic
lifecycles, I am interested in signaling pathways that all apicomplexans utilize to transmit from one cell to the
next. This project centers around cyclic GMP (cGMP), which acts as a molecular switch for apicomplexan
motility, facilitating cell to cell transmission. Although dozens of studies have focused on kinases activated by
cGMP (i.e. PKGs), little is known about how cGMP is produced in these organisms. In other eukaryotes,
guanylate cyclases (GCs) catalyze the formation of cGMP from GTP and are regulated by domains that sense
nitric oxide or peptide hormones. However, apicomplexan GCs lack known regulatory domains, instead
possess an N-terminal P-type ATPase domain of unknown function. To investigate the function of these unique
enzymes, I chose Toxoplasma gondii as a model due to its clinical importance and excellent experimental
tractability. I found that T. gondii GC localizes to the apex of the parasite where it functions as a gatekeeper for
apical protein exocytosis, the initiating and rate-limiting step of parasite motility. Conditional loss of TgGC
paralyzes parasites preventing lytic growth and spreading, phenocopying loss of TgPKG. Though TgGC
possesses dual functional domains, cell-permeable cGMP rescues TgGC depletion, further indicating that its
primary role is to synthesize cGMP. Through genetic complementation, I found that both domains of TgGC
were required, suggesting that the P-type ATPase domain may function as a regulatory domain. These P-type
ATPase domains most closely resemble P4-ATPases, which utilize energy from ATP to flip phospholipids
across membranes to maintain the lipid asymmetry needed for a variety of cellular processes. Since P4-
ATPases typically function as an α/β heterodimer with a Cdc50 chaperone, apicomplexan GCs are likely to
function in a complex with one or more other proteins. Recently I identified 13 candidate interactors of TgGC
using BioID proximity labeling, including a Cdc50 protein that is predicted to be essential based on a prior
genome-wide CRISPR screen. The goal of this K22 proposal is to understand how apicomplexan GCs are
regulated, both intramolecularly and through interacting partners. This goal will be accomplished by two
independent Specific Aims that will define the biochemical function of the P4-ATPase domain (Specific Aim 1)
as well as the contributions of TgCdc50 and other interacting proteins (Specific Aim 2) with respect to TgGC
function. This research will reveal the mechanism by which essential cGMP signaling is initiated in
apicomplexans and will serve as a springboard for my independent career as a principle investigator at an
acad...

## Key facts

- **NIH application ID:** 9989023
- **Project number:** 5K22AI144035-02
- **Recipient organization:** UNIVERSITY OF OKLAHOMA HLTH SCIENCES CTR
- **Principal Investigator:** Kevin Michael Brown
- **Activity code:** K22 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $108,000
- **Award type:** 5
- **Project period:** 2019-08-05 → 2022-07-31

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/9989023

## Citation

> US National Institutes of Health, RePORTER application 9989023, Non-conventional regulation of essential apicomplexan guanylate cyclases (5K22AI144035-02). Retrieved via AI Analytics 2026-05-23 from https://api.ai-analytics.org/grant/nih/9989023. Licensed CC0.

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