scholarly journals Dissection of the in vitro developmental program of Hammondia hammondi reveals a link between stress sensitivity and life cycle flexibility in Toxoplasma gondii

eLife ◽  
2018 ◽  
Vol 7 ◽  
Author(s):  
Sarah L Sokol ◽  
Abby S Primack ◽  
Sethu C Nair ◽  
Zhee S Wong ◽  
Maiwase Tembo ◽  
...  
Keyword(s):  
Life Cycle ◽  
Toxoplasma Gondii ◽  
Stress Responses ◽  
Cyst Formation ◽  
Stress Sensitivity ◽  
Stress Exposure ◽  
Developmental Program ◽  
Dramatic Difference ◽  
Sequential Infection

Most eukaryotic parasites are obligately heteroxenous, requiring sequential infection of different host species in order to survive. Toxoplasma gondii is a rare exception to this rule, having a uniquely facultative heteroxenous life cycle. To understand the origins of this phenomenon, we compared development and stress responses in T. gondii to those of its its obligately heteroxenous relative, Hammondia hammondi and have identified multiple H. hammondi growth states that are distinct from those in T. gondii. Of these, the most dramatic difference was that H. hammondi was refractory to stressors that robustly induce cyst formation in T. gondii, and this was reflected most dramatically in its unchanging transcriptome after stress exposure. We also found that H. hammondi could be propagated in vitro for up to 8 days post-excystation, and we exploited this to generate the first ever transgenic H. hammondi line. Overall our data show that H. hammondi zoites grow as stringently regulated, unique life stages that are distinct from T. gondii tachyzoites, and implicate stress sensitivity as a potential developmental innovation that increased the flexibility of the T. gondii life cycle.

scholarly journals Hammondia hammondihas a developmental programin vitrothat mirrors its stringent two host life cycle

2017 ◽  
Author(s):  
Sarah L. Sokol ◽  
Abby S. Primack ◽  
Sethu C. Nair ◽  
Zhee S. Wong ◽  
Maiwase Tembo ◽  
...  
Keyword(s):  
Life Cycle ◽  
Toxoplasma Gondii ◽  
Specific Growth ◽  
Cyst Formation ◽  
Host Cells ◽  
Tissue Cyst ◽  
Life Stages ◽  
New Host ◽  
Host Life

AbstractHammondia hammondiis the nearest relative ofToxoplasma gondii,but unlikeT. gondiiis obligately heteroxenous. We have comparedH. hammondiandT. gondiidevelopmentin vitroand identified multipleH. hammondi-specific growth states. Despite replicating slower thanT. gondii,H. hammondiwas resistant to pH-induced tissue cyst formation early after excystation. However, in the absence of stressH. hammondispontaneously converted to a terminally differentiated tissue cyst stage whileT. gondiidid not. CulturedH. hammondicould infect new host cells for up to 8 days following excystation, and this period was exploited to generate stably transgenicH. hammondi. Coupled with RNAseq analyses, our data clearly show thatH. hammondizoites grow as stringently regulated life stages that are fundamentally distinct fromT. gondiitachyzoites and bradyzoites.

scholarly journals A Comparison of Stage Conversion in the Coccidian Apicomplexans Toxoplasma gondii, Hammondia hammondi, and Neospora caninum

2020 ◽  
Vol 10 ◽  
Author(s):  
Sarah L. Sokol-Borrelli ◽  
Rachel S. Coombs ◽  
Jon P. Boyle
Keyword(s):  
Life Cycle ◽  
Toxoplasma Gondii ◽  
Stress Responses ◽  
Neospora Caninum ◽  
Life Forms ◽  
New Host ◽  
Environmental Signals ◽  
Stage Conversion ◽  
Genetic Components ◽  
Species Specific

Stage conversion is a critical life cycle feature for several Apicomplexan parasites as the ability to switch between life forms is critical for replication, dissemination, pathogenesis and ultimately, transmission to a new host. In order for these developmental transitions to occur, the parasite must first sense changes in their environment, such as the presence of stressors or other environmental signals, and then respond to these signals by initiating global alterations in gene expression. As our understanding of the genetic components required for stage conversion continues to broaden, we can better understand the conserved mechanisms for this process and unique components and their contribution to pathogenesis by comparing stage conversion in multiple closely related species. In this review, we will discuss what is currently known about the mechanisms driving stage conversion in Toxoplasma gondii and its closest relatives Hammondia hammondi and Neospora caninum. Work by us and others has shown that these species have some important differences in the way that they (1) progress through their life cycle and (2) respond to stage conversion initiating stressors. To provide a specific example of species-specific complexities associated with stage conversion, we will discuss our recent published and unpublished work comparing stress responses in T. gondii and H. hammondi.

scholarly journals Analysis of Noncanonical Calcium-Dependent Protein Kinases in Toxoplasma gondii by Targeted Gene Deletion Using CRISPR/Cas9

2016 ◽  
Vol 84 (5) ◽  
pp. 1262-1273 ◽  
Author(s):  
Shaojun Long ◽  
Qiuling Wang ◽  
L. David Sibley
Keyword(s):  
Toxoplasma Gondii ◽  
Protein Kinases ◽  
Gene Deletion ◽  
Cyst Formation ◽  
Content Type ◽  
Calcium Dependent ◽  
Targeted Gene Deletion ◽  
Dependent Protein ◽  
Calcium Dependent Protein Kinases

Calcium-dependent protein kinases (CDPKs) are expanded in apicomplexan parasites, especially inToxoplasma gondiiwhere 14 separate genes encoding these enzymes are found. Although previous studies have shown that several CDPKs play a role in controlling invasion, egress, and cell division inT. gondii, the roles of most of these genes are unexplored. Here we developed a more efficient method for gene disruption using CRISPR (clustered regularly interspaced short palindromic repeats)/Cas9 (CRISPR-associated protein 9) that was modified to completely delete large, multiexonic genes from the genome and to allow serial replacement by recycling of the selectable marker using Cre-loxP. Using this system, we generated a total of 24 mutants in type 1 and 2 genetic backgrounds to ascertain the functions of noncanonical CDPKs. Remarkably, although we were able to confirm the essentiality of CDPK1 and CDPK7, the majority of CDPKs had no discernible phenotype for growthin vitroor infection in the mouse model. The exception to this was CDPK6, loss of which leads to reduced plaquing, fitness defect in a competition assay, and reduced tissue cyst formation in chronically infected mice. Our findings highlight the utility of CRISPR/Cas9 for rapid serial gene deletion and also suggest that additional models are needed to reveal the functions of many genes inT. gondii.

scholarly journals Leishmania donovani parasite requires Atg8 protein for infectivity and survival under stress

2019 ◽  
Vol 10 (11) ◽  
Author(s):  
Sagnik Giri ◽  
Chandrima Shaha
Keyword(s):  
Life Cycle ◽  
Leishmania Donovani ◽  
Protein A ◽  
Critical Event ◽  
Stress Exposure ◽  
Cellular Defense ◽  
Atg8 Protein ◽  
Insect Gut

Abstract The importance of autophagy in parasites with a digenetic life cycle like Leishmania spp. is significant. The parasite survives as promastigotes in the insect gut and as immotile amastigotes in mammals. This study demonstrates increased autophagy in Leishmania parasite during progression of in vitro life cycle and upon exposure to stress stimuli like starvation, oxidative stress, and drugs. Autophagy inhibition during stress exposure increased cell death, indicating the importance of autophagy in cellular defense against adverse conditions. Atg8 protein, a homolog of mammalian autophagy protein LC3 is expressed in Leishmania parasite but its function remains unknown. Overexpression of Atg8 (Atg8-OE) rendered the parasites resistant to stress and capable of infecting macrophages in substantial numbers; however, disruption of the Atg8 gene (ΔAtg8) resulting in suppression of Atg8 protein expression, increased susceptibility to stress and reduced the capability to cause infection. A critical event in the Leishmania parasite lifecycle is the differentiation of promastigote forms to the disease causing amastigote forms. The failure of ΔAtg8 parasites lacking Atg8 protein to differentiate into amastigotes, unlike the Atg8-OE and vector-transfected parasites, clearly indicated Atg8 involvement in a crucial event. The inability of ΔAtg8 parasites to infect macrophages in vitro was verified in an in vivo mouse model of leishmaniases where infection could not be induced by the ΔAtg8 parasites. Autophagy is known to be involved in the remodeling of damaged organelles. The accumulation of Atg8 around damaged mitochondria suggested increase of autophagy in the vicinity of the organelle. This buildup was prevented when mitochondria generated reactive oxygen species that were quenched, suggesting them as possible signaling molecules for sensing mitochondrial instability. In summary, our study provides new evidences for a crucial role of Atg8 protein in sustaining Leishmania parasite survival during life cycle and stress exposure, differentiation to amastigotes, and their infective abilities.

Toxoplasma gondii reorganizes the host cell architecture during spontaneous cyst formation in vitro

Parasitology ◽  
2017 ◽  
Vol 145 (8) ◽  
pp. 1027-1038 ◽  
Author(s):  
T. C. Paredes-Santos ◽  
E. S. Martins-Duarte ◽  
W. de Souza ◽  
M. Attias ◽  
R. C. Vommaro
Keyword(s):  
Toxoplasma Gondii ◽  
Host Cell ◽  
Cyst Wall ◽  
Cyst Formation ◽  
Acute Stage ◽  
Host Cells ◽  
Cell Cytoplasm ◽  
Chronic Stage ◽  
Host Cell Cytoplasm

AbstractToxoplasma gondii is an intracellular protozoan parasite that causes toxoplasmosis, a prevalent infection related to abortion, ocular diseases and encephalitis in immuno-compromised individuals. In the untreatable (and life-long) chronic stage of toxoplasmosis, parasitophorous vacuoles (PVs, containing T. gondii tachyzoites) transform into tissue cysts, containing slow-dividing bradyzoite forms. While acute-stage infection with tachyzoites involves global rearrangement of the host cell cytoplasm, focused on favouring tachyzoite replication, the cytoplasmic architecture of cells infected with cysts had not been described. Here, we characterized (by fluorescence and electron microscopy) the redistribution of host cell structures around T. gondii cysts, using a T. gondii strain (EGS) with high rates of spontaneous cystogenesis in vitro. Microtubules and intermediate filaments (but not actin microfilaments) formed a ‘cage’ around the cyst, and treatment with taxol (to inhibit microtubule dynamics) favoured cystogenesis. Mitochondria, which appeared adhered to the PV membrane, were less closely associated with the cyst wall. Endoplasmic reticulum (ER) profiles were intimately associated with folds in the cyst wall membrane. However, the Golgi complex was not preferentially localized relative to the cyst, and treatment with tunicamycin or brefeldin A (to disrupt Golgi or ER function, respectively) had no significant effect on cystogenesis. Lysosomes accumulated around cysts, while early and late endosomes were more evenly distributed in the cytoplasm. The endocytosis tracer HRP (but not BSA or transferrin) reached bradyzoites after uptake by infected host cells. These results suggest that T. gondii cysts reorganize the host cell cytoplasm, which may fulfil specific requirements of the chronic stage of infection.

scholarly journals Sex-specific neural responses to acute psychosocial stress in depression

2022 ◽  
Vol 12 (1) ◽  
Author(s):  
Daifeng Dong ◽  
Maria Ironside ◽  
Emily L. Belleau ◽  
Xiaoqiang Sun ◽  
Chang Cheng ◽  
...  
Keyword(s):  
Sex Differences ◽  
Exposure Time ◽  
Stress Responses ◽  
A Priori ◽  
Stress Sensitivity ◽  
Anterior Cingulate ◽  
First Episode ◽  
Stress Exposure ◽  
Frontoparietal Network ◽  
Dorsolateral Prefrontal

AbstractMajor Depressive Disorder (MDD) is characterized by increased stress sensitivity. Emerging findings in healthy adults suggest that stress responses within limbic/striatal-prefrontal regions are moderated by sex and unfold over time. Thus, we hypothesized that stress response abnormalities in MDD might be affected by sex and stress exposure time. The Montreal Imaging Stress Task was administered to 124 unmedicated patients with first-episode MDD (76 females) and 243 healthy controls (HC; 137 females) during functional magnetic resonance imaging (fMRI). Based on prior studies, amygdala, hippocampus, medial orbitofrontal cortex (mOFC), nucleus accumbens (NAc) and dorsolateral prefrontal cortex (dlPFC) were selected as a priori regions of interest. In a complementary approach, we probed the effects of stress on the frontoparietal network (FPN) and a network including the amygdala, NAc and anterior cingulate cortex (ACC). Across groups, males exhibited higher dlPFC activity and right FPN amplitude than females. Relative to female HCs, the female MDD group had less deactivation in limbic/striatal regions (amygdala, NAc, hippocampus, Amygdala-NAc-ACC network). Furthermore, unlike female HCs, the female MDD group failed to show a significant increase of deactivation over stress exposure time in the amygdala, mOFC and NAc. Our findings confirm the importance of considering sex differences when investigating neural stress responses. Case-control differences in neural stress responses observed in females (but not males) provide insights into sex differences in the etiology and pathophysiology of depression. The failure to deactivate limbic/NAc regions in depressed females point to dysfunction of adaptive stress responses over stress exposure time.

scholarly journals Toxoplasma gondii AP2XII-2 Contributes to Proper Progression through S-Phase of the Cell Cycle

mSphere ◽  
2020 ◽  
Vol 5 (5) ◽  
Author(s):  
Sandeep Srivastava ◽  
Michael W. White ◽  
William J. Sullivan
Keyword(s):  
Cell Cycle ◽  
Toxoplasma Gondii ◽  
Cell Cycle Progression ◽  
Transcriptional Repressor ◽  
Protozoan Parasite ◽  
Cyst Formation ◽  
S Phase ◽  
Tissue Cyst ◽  
Content Type

ABSTRACT Toxoplasma gondii is a protozoan parasite that causes lifelong chronic infection that can reactivate in immunocompromised individuals. Upon infection, the replicative stage (tachyzoite) converts into a latent tissue cyst stage (bradyzoite). Like other apicomplexans, T. gondii possesses an extensive lineage of proteins called ApiAP2s that contain DNA-binding domains first characterized in plants. The function of most ApiAP2s is unknown. We previously found that AP2IX-4 is a cell cycle-regulated ApiAP2 expressed only in dividing parasites as a putative transcriptional repressor. In this study, we purified proteins interacting with AP2IX-4, finding it to be a component of the recently characterized microrchidia (MORC) transcriptional repressor complex. We further analyzed AP2XII-2, another cell cycle-regulated factor that associates with AP2IX-4. We monitored parallel expression of AP2IX-4 and AP2XII-2 proteins in tachyzoites, detecting peak expression during S/M phase. Unlike AP2IX-4, which is dispensable in tachyzoites, loss of AP2XII-2 resulted in a slowed tachyzoite growth due to a delay in S-phase progression. We also found that AP2XII-2 depletion increased the frequency of bradyzoite differentiation in vitro. These results suggest that multiple AP2 factors collaborate to ensure proper cell cycle progression and tissue cyst formation in T. gondii. IMPORTANCE Toxoplasma gondii is a single-celled parasite that persists in its host by converting into a latent cyst stage. This work describes a new transcriptional factor called AP2XII-2 that plays a role in properly maintaining the growth rate of replicating parasites, which contributes to signals required for development into its dormant stage. Without AP2XII-2, Toxoplasma parasites experience a delay in their cell cycle that increases the frequency of latent cyst formation. In addition, we found that AP2XII-2 operates in a multisubunit complex with other AP2 factors and chromatin remodeling machinery that represses gene expression. These findings add to our understanding of how Toxoplasma parasites balance replication and dormancy, revealing novel points of potential therapeutic intervention to disrupt this clinically relevant process.

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