Biology subjects

Kochanowsky, J. A.

Publications and source records attributed to Kochanowsky, J. A..

6 recordsLinked to original sources

bioRxiv2025

Defining neuronal responses to the neurotropic parasite Toxoplasma gondii

A select group of pathogens infect neurons in the brain. Prior dogma held that neurons were "defenseless" against infecting microbes, but many studies suggest that neurons can mount anti-microbial defenses. However, a knowledge gap in understanding how neurons respond in vitro and in vivo to different classes of micro-organisms remains. To address this gap, we compared a transcriptional dataset derived from primary neuron cultures (PNCs) infected with the neurotropic intracellular parasite Toxoplasma gondii with a dataset derived from neurons injected with T. gondii protein in vivo. These curated responses were then compared to the transcriptional responses of PNCs infected with the single stranded RNA viruses West Nile Virus (WNV) or Zika Virus (ZKV). These analyses highlighted a conserved response to infection associated with chemokines (Cxcl10, Ccl2) and cytokines (interferon signaling). However, T. gondii had diminished IFN- signaling in vitro compared to the viral datasets and was uniquely associated with a decrease in neuron-specific genes (Snap25, Slc17a7, Prkcg). These data underscore that neurons participate in infection-induced neuroinflammation and illustrate that neurons possess both pathogen-specific and pathogen-conserved responses. ImportanceThough neurons are commonly the target of pathogens that infect the CNS, few datasets assess the neuronal response to infection. This paucity of data is likely because neurons are perceived to have diminished immune capabilities. However, to understand the role of neurons in neuroinflammation and their immune capabilities, their responses must be investigated. Here we analyzed publicly accessible, neuron-specific datasets to compare neuron responses to a eukaryotic pathogen versus two Orthoflaviviruses. A better understanding of neuron responses to different infections will allow us to develop methods for inhibiting pathways that lead to neuron dysfunction, enhancing those that limit pathogen survival, and mitigating infection-induced damage to the CNS.

microbiology↗

bioRxiv2024

Trichomonas vaginalis extracellular vesicles suppress IFNϵ-mediated protection against host cell cytolysis

Trichomonas vaginalis is a commonly acquired sexually transmitted infection (STI) often found in symbiosis with the intracellular bacterium Mycoplasma hominis, an opportunistic pathogen of the female reproductive tract associated with bacterial vaginosis. How this symbiosis affects infection outcomes, and the host cell innate immune response is still poorly understood. Here we show that T. vaginalis extracellular vesicles down-regulate a non-canonical type I interferon, interferon-epsilon, and suppress type I interferon responses. Transcriptomic analysis reveals that infection with T. vaginalis in symbiosis with M. hominis or M. hominis alone upregulates genes involved in the type I IFN response, but infection with T. vaginalis alone does not. Finally, we show that interferon-epsilon stimulation is protective against T. vaginalis cytoadherence and cytolysis of host cells and increases the ability of neutrophils to kill the parasite. These studies provide insight into the innate immune response induced by a highly prevalent STI and its bacterial symbiont.

microbiology↗

bioRxiv2024

Toxoplasma type II effector GRA15 has limited influence in vivo

Toxoplasma gondii is an intracellular parasite that establishes a long-term infection in the brain of many warm-blooded hosts, including humans and rodents. Like all obligate intracellular microbes, Toxoplasma uses many effector proteins to manipulate the host cell to ensure parasite survival. While some of these effector proteins are universal to all Toxoplasma strains, some are polymorphic between Toxoplasma strains. One such polymorphic effector is GRA15. The gra15 allele carried by type II strains activates host NF-{kappa}B signaling, leading to the release of cytokines such as IL-12, TNF, and IL-1{beta} from immune cells infected with type II parasites. Prior work also suggested that GRA15 promotes early host control of parasites in vivo, but the effect of GRA15 on parasite persistence in the brain and the peripheral immune response has not been well defined. For this reason, we sought to address this gap by generating a new II{Delta}gra15 strain and comparing outcomes at 3 weeks post infection between WT and II{Delta}gra15 infected mice. We found that the brain parasite burden and the number of macrophages/microglia and T cells in the brain did not differ between WT and II{Delta}gra15 infected mice. In addition, while II{Delta}gra15 infected mice had a lower number and frequency of splenic M1-like macrophages and frequency of PD-1+ CTLA-4+ CD4+ T cells and NK cells compared to WT infected mice, the IFN-{gamma}+ CD4 and CD8 T cell populations were equivalent. In summary, our results suggest that in vivo GRA15 may have a subtle effect on the peripheral immune response, but this effect is not strong enough to alter brain parasite burden or parenchymal immune cell number at 3 weeks post infection.

microbiology↗

bioRxiv2022

ROP16-mediated activation of STAT6 facilitates encystment of type III Toxoplasma gondii in neurons

Toxoplasma gondii establishes a long-lived latent infection in the central nervous system (CNS) of its hosts. Reactivation in immunocompromised individuals can lead to life threatening disease. Latent infection is driven by the ability of the parasite to convert from the acute-stage tachyzoite to the latent-stage bradyzoite which resides in long-lived intracellular cysts. While much work has focused on the parasitic factors that drive cyst development, the host factors that influence encystment are not well defined. Here we show that a polymorphic secreted parasite kinase (ROP16), that phosphorylates host cell proteins, mediates efficient encystment of T. gondii in stress-induced models of encystment and primary neuronal cell cultures (PNCs) in a strain-specific manner. Using short-hairpin RNA (shRNA) knockdowns in human foreskin fibroblasts (HFFs) and PNCs from transgenic mice, we determined that ROP16s cyst enhancing abilities are mediated by phosphorylation of the host cell transcription factor STAT6. To test the role of STAT6 in vivo, we infected STAT6KO mice, finding that, compared to infected wild-type mice, infected STAT6KO mice have a decrease in cyst burden, but not overall parasite burden or dissemination to the CNS. Finally, we found a similar ROP16-dependent encystment defect in human pluripotent stem cell-derived neurons. Together, these findings identify a host cell factor (STAT6) that T. gondii manipulates in a strain-specific manner to generate a favorable encystment environment.

microbiology↗

bioRxiv2021

IFN-γ stimulated murine and human neurons mount anti-parasitic defenses against the intracellular parasite Toxoplasma gondii

Dogma holds that Toxoplasma gondii persists in neurons because neurons cannot clear intracellular parasites, even with IFN-{gamma} stimulation. As several recent studies questioned this idea, we used primary murine neuronal cultures from wild-type and transgenic mice in combination with IFN-{gamma} stimulation and parental and transgenic parasites to reassess IFN-{gamma} dependent neuronal clearance of intracellular parasites. We found that neurons respond to IFN-{gamma} and that a subset of neurons clear intracellular parasites via immunity regulated GTPases. Whole neuron reconstructions from mice infected with parasites that trigger neuron GFP expression only after full invasion revealed that [~]40% of these T. gondii-invaded neurons no longer harbor parasites. Finally, IFN-{gamma} stimulated human stem cell derived neurons showed a [~] 50% decrease in parasite infection rate when compared to unstimulated cultures. This work highlights the capability of human and murine neurons to mount cytokine-dependent anti-T. gondii defense mechanisms in vitro and in vivo.

neuroscience↗

bioRxiv2019

The ROP16III-dependent early immune response determines the sub-acute CNS immune response and type III Toxoplasma gondii survival

Toxoplasma gondii is an intracellular parasite that persistently infects the CNS and that has genetically distinct strains which provoke different acute immune responses. How differences in the acute immune response affect the CNS immune response is unknown. To address this question, we used two persistent Toxoplasma strains (type II and type III) and examined the CNS immune response at 21 days post infection (dpi). Contrary to acute infection studies, type III-infected mice had higher numbers of total CNS T cells and macrophages/microglia but fewer alternatively activated macrophages (M2s) and regulatory T cells (Tregs) than type II-infected mice. By profiling splenocytes at 5, 10 and 21 dpi, we determined that at 5 dpi type III-infected mice had more M2s while type II-infected mice had more classically activated macrophages (M1s) and these responses flipped over time. To test how these early differences influence the CNS immune response, we engineered the type III strain to lack ROP16 (III{Delta}rop16), the polymorphic effector protein that drives the type III-associated M2 response. III{Delta}rop16-infected mice showed a type II-like neuroinflammatory response with fewer infiltrating T cells and macrophages/microglia and more M2s and an unexpectedly low CNS parasite burden. At 5 dpi, III{Delta}rop16-infected mice showed a mixed inflammatory response with more M1s, M2s, T effector cells, and Tregs, and decreased rates of infection of peritoneal exudative cells (PECs). These data suggested that type III parasites need the early ROP16-associated M2 response to avoid clearance, possibly by the Immunity-Related GTPases (IRGs), IFN-{gamma} dependent proteins essential for murine defenses against Toxoplasma. To test this possibility, we infected IRG-deficient mice and found that III{Delta}rop16 parasites now maintained parental levels of PECs infection. Collectively, these studies suggest that, for the type III strain, rop16III plays a key role in parasite persistence and influences the sub-acute CNS immune response.\n\nAuthor SummaryToxoplasma is a ubiquitous intracellular parasite that establishes an asymptomatic brain infection in immunocompetent individuals. However, in the immunocompromised and the developing fetus, Toxoplasma can cause problems ranging from fever to chorioretinitis to severe toxoplasmic encephalitis. Emerging evidence suggests that the genotype of the infecting Toxoplasma strain may influence these outcomes, possibly through the secretion of Toxoplasma strain-specific polymorphic effector proteins that trigger different host cell signaling pathways. While such strain-specific modulation of host cell signaling has been shown to affect acute immune responses, it is unclear how these differences influence the sub-acute or chronic responses in the CNS, the major organ affected in symptomatic disease. This study shows that genetically distinct strains of Toxoplasma provoke strain-specific CNS immune responses and that, for one strain (type III), the acute and sub-acute immune responses and parasite survival are heavily influenced by a polymorphic parasite gene (rop16III).

microbiology↗