Monitoring neuronal adaption to Toxoplasma gondii infection on the whole transcriptome level in the central and enteric nervous system

Principle Investigator

Scientific interest within the context of the graduate college:

Neuro-immune interactions.

Project description:

Introduction: Toxoplasma gondii, a ubiquitous intracellular parasite with a profound ability to infect virtually all warm-blooded animals, is responsible for a significant public health concern. In humans, infection can lead to toxoplasmosis, which, while often asymptomatic in healthy individuals, poses severe risks to immunocompromised patients and pregnant women.¹ Moreover, emerging evidence suggests T. gondii’s potential triggering of various neurological disorders, including schizophrenia and other psychiatric conditions, underscoring the critical need to understand the parasite’s interaction with the nervous system.² The central and enteric nervous systems are intricate networks that regulate numerous vital functions, from cognition and sensory processing in the central nervous system (CNS) to gastrointestinal motility and secretion in the enteric nervous system (ENS).³ T. gondii’s ability to invade and persist in these neural environments suggests significant adaptations by both the parasite and host neurons. Investigating these adaptations, particularly at the transcriptional level, provides insights into the mechanisms of pathogenesis, neuronal resilience, and potential neuropathology associated with infection.

We aim to monitor the transcriptional changes occurring in neurons during T. gondii infection. To this end, we have already established the respective lines and protocols for bulk sequencing of purified RNA from sort-purified GFP⁺ neuronal nuclei from the T. gondii-infected brain and gut tissue of the Snap25^Cre/+ x INTACT system, in which the GFP expression is only activated in neurons. To monitor alterations in enteric neurons’ composition and transcriptional changes in subsets of enteric neurons, we will perform RNA sequencing from brains and gut tissue. To this end, we will separately sort-purify GFP⁺ nuclei from Snap25^Cre/+ x INTACT system to obtain highly pure neuronal nuclei. These neuronal cell nuclei will be analyzed using the RNA-sequencing platform, which we have already implemented for single-nuclei sequencing. Together, these experiments will identify genes in neurons regulated in the context of T. gondii infection.

Based on our data detecting a type II interferon signature in the ENS in various inflammation models, we will perform bulk RNA sequencing of the ENS and CNS, in which we have deleted the Ifngr1 specifically in neurons by the Snap25^Cre/+ x Ifngr1^fl/fl INTACT system and which allow for sort-purification based on the nuclear GFP signal. This experimental setup will allow us to study the potentially protective gene expression signature in neurons induced by IFN-γ. Furthermore, these experiments will enable us measure if neuron-specific deletion of the IFN-γ receptor results in increased T.gondii burden in various organs. Therefore, these experiments will reveal how IFN-γ-signaling in neurons contributes to anti-toxoplasma immunity.

Taken together, these datasets will yield correlations of differentially expressed genes of ENS and CNS neurons and their relative abundances, exposing disease-relevant signaling circuits triggered in neurons. This global perspective is essential for uncovering the molecular underpinnings of T. gondii’s impact on neuronal function and survival, which could lead to the development of targeted therapies and interventions.^1,2

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