Biology subjects

Cabo, L. F.

Publications and source records attributed to Cabo, L. F..

6 recordsLinked to original sources

bioRxiv2024

Toxoplasma gondii infection misdirects placental trophoblast lineage specification

Pregnancy is a critical point of vulnerability to infection and other insults that could compromise proper fetal development. The placenta acts as a protective and nutrient-permeable barrier to most infectious agents, but a few are capable of bypassing its defenses. Remarkably little is known about how exposure to these select pathogens might impact ongoing placental development. Here we demonstrate that Toxoplasma gondii entirely misdirects the developmental program of trophoblast stem cells. Infection of progenitor cytotrophoblasts prevents fusion and differentiation to infection-resistant syncytiotrophoblast. Rather, T. gondii elicits a unique transcriptional identity that polarizes cytotrophoblasts to the infection-permissive extravillous trophoblast fate. Strong evidence of developmental disruption is found in multiple orthogonal models, including trophoblast stem cells, trophoblast organoids, and chorionic villi. Manipulation of cell fate by the parasite is most dramatic in trophoblast organoids, where we see robust outgrowth of HLA-G(+) extravillous trophoblasts. Collectively, these data show that Toxoplasma antagonizes differentiation of an infection-resistant cell type by inducing formation of an infection-permissive cell type, therefore potentiating its own transmission to the fetus.

developmental biology↗

bioRxiv2023

Human trophoblast stem cells can be used to model placental susceptibility to Toxoplasma gondii and highlight the critical importance of the trophoblast cell surface in pathogen resistance

The placenta is a critical barrier against viral, bacterial, and eukaryotic pathogens. For most teratogenic pathogens, the precise molecular mechanisms of placental resistance are still being unraveled. Given the importance to understand these mechanisms and challenges in replicating trophoblast-pathogen interactions using in vitro models, we tested an existing stem-cell derived model of trophoblast development for its relevance to infection with Toxoplasma gondii. We grew human trophoblast stem cells (TSCT) under conditions leading to either syncytiotrophoblast (TSSYN) or cytotrophoblast (TSCYT) and infected them with T. gondii. We evaluated T. gondii proliferation and invasion, cell ultrastructure, as well as for transcriptome changes after infection. TSSYNs cells showed similar ultrastructure compared to primary cells and villous explants when analyzed by TEM and SEM, a resistance to T. gondii adhesion could be visualized on the SEM level. Furthermore, TSSYNs were highly refractory to parasite adhesion and replication, while TSCYT were not. RNA-seq data on mock-treated and infected cells identified differences between cell types as well as how they responded to T. gondii infection. We also evaluated if TSSC-derived SYNs and CYTs had distinct resistance profiles to another vertically transmitted facultative intracellular pathogen, Listeria monocytogenes. We demonstrate that TSSYNs are highly resistant to L. monocytogenes, while TSCYTs are not. Like T. gondii, TSSYN resistance to L. monocytogenes was at the level of bacterial adhesion. Altogether, our data indicate that stem-cell derived trophoblasts recapitulate resistance profiles of primary cells to T. gondii and highlight the critical importance of the placental surface in cell-autonomous resistance to teratogens.

microbiology↗

bioRxiv2022

Heme-induced genes facilitate endosymbiont (Sodalis glossinidius) colonization of the tsetse fly (Glossina morsitans) midgut

Tsetse flies (Glossina spp.) feed exclusively on vertebrate blood. After a blood meal, the enteric endosymbiont Sodalis glossinidius is exposed to various environmental stressors including high levels of heme. To investigate how S. glossinidius morsitans (Sgm, the Sodalis subspecies that resides within the gut of G. morsitans) tolerates the heme-induced oxidative environment of tsetses midgut, we used RNAseq to identify bacterial genes that are differentially expressed in cells cultured in high versus lower heme environments. Our analysis identified 436 genes that were significantly differentially expressed (> or < 2-fold) in the presence of high heme [219 heme-induced genes (HIGs) and 217 heme-repressed genes (HRGs)]. HIGs were enriched in Gene Ontology (GO) terms related to regulation of a variety of biological functions, including gene expression and metabolic processes. We observed that 11 out of 13 Sgm genes that were heme regulated in vitro were similarly regulated in bacteria that resided within tsetses midgut 24 hr (high heme environment) and 96 hr (low heme environment) after the flies had consumed a blood meal. We used intron mutagenesis to make insertion mutations in 12 Sgm HIGs and observed no significant change in growth in vitro in any of the mutant strains in high versus low heme conditions. However, Sgm strains that carried mutations in genes encoding a putative undefined phosphotransferase sugar (PTS) system component (SG2427), fucose transporter (SG0182), bacterioferritin (SG2280), and a DNA-binding protein (SGP1-0002) presented growth and/or survival defects in tsetse midguts as compared to normal Sgm. These findings suggest that the uptake up of sugars and storage of iron represent strategies that Sgm employs to successfully reside within the high heme environment of its tsetse hosts midgut. Our results are of epidemiological relevance, as many hematophagous arthropods house gut-associated bacteria that mediate their hosts competency as a vector of disease-causing pathogens. Author summaryTsetse flies feed exclusively on vertebrate blood. This nutrient source contains large quantities of heme, which can be toxic to the flys associated microorganisms. We investigated the genetic mechanisms that underlie the ability of the bacterial endosymbiont, Sodalis glossinidius, to successfully reside within tsetses heme-rich midgut. Exposure of cultured S. glossinidius to high levels of heme induced changes in the expression of genes that encode proteins involved in transcription, replication and repair of DNA, inorganic ion transport, and carbohydrate transport and metabolism processes. Changes in the expression of several of these same S. glossinidius genes also occurred within tsetses midgut following exposure to a blood meal. S. glossinidius genetically engineered to present mutations in several of these heme regulated genes were unable to successfully colonize tsetses gut. Our results provide insight into how bacteria that live in the gut of blood feeding arthropods mitigate the toxic effects of excessive heme. This information is of epidemiological relevance, as many of these bacteria influence their hosts ability to transmit disease pathogens that cause disease in humans and domesticated animals.

microbiology↗

bioRxiv2022

A transcriptional network required for Toxoplasma gondii tissue cyst formation is dispensable for long-term persistence

Cyst formation is a key feature of the T. gondii life cycle but the genetic networks that drive this process are not yet fully characterized. To identify new components of this network, we compared T. gondii to its nearest extant relative Hammondia hammondi given the critical differences between these species in the timing and efficiency of cyst formation. Using transcriptional data from critical developmental and pH exposure time points from both species, we identified the gene TGVEG_311100, which we named Regulator of Cystogenesis 1 (ROCY1), as being both necessary and sufficient for cyst formation in T. gondii. Compared to WT parasites, TGVEG{Delta}ROCY1 parasites formed significantly fewer tissue cysts in response to alkaline pH stress in vitro and cysts were nearly undetectable in mouse brains for up to 9 weeks post-infection. Overexpression of tagged ROCY1 in WT parasites was sufficient to induce cyst formation in vitro in both WT and ROCY1-deficient parasites, demonstrating that ROCY1 is both necessary and sufficient for cyst formation. Moreover this induction of cyst formation required at least 1 of 3 predicted CCCH Zinc finger domains. Mice chronically infected with {Delta}ROCY1 parasites had detectable tachyzoites in the brain for up to 37 days post-infection (while mice infected with WT parasites did not), and CNS transcriptional analyses at day 30 post-infection throughout the chronic phase of infection revealed inflammatory signatures consistent with acute infection in {Delta}ROCY1 parasites compared to WT. Despite our inability to detect brain cysts in infected mice, both WT and {Delta}ROCY1 knockout parasites reactivated after dexamethasone treatment with similar timing and magnitude for up to 5 months post infection, challenging the paradigm that long term parasite persistence in the CNS requires cyst formation. These data identify a new regulator of cyst formation in T. gondii that is both necessary and sufficient for cyst formation, and whose function relies on its conserved nucleic acid binding motif.

microbiology↗

bioRxiv2020

Toxoplasma gondii GRA28 is required for specific induction of the regulatory chemokine CCL22 in human and mouse cells.

Toxoplasma gondii is an intracellular protozoan pathogen of humans that can cross the placenta and result in adverse pregnancy outcomes and long-term birth defects. The mechanism used by T. gondii to cross the placenta are unknown but complex interactions with the host immune response are likely to play a role in dictating infection outcomes during pregnancy. Prior work showed that T. gondii infection dramatically and specifically increases the secretion of the immunomodulatory chemokine CCL22 in human placental cells during infection.. Given the important role of this chemokine during pregnancy, we hypothesized that CCL22 induction was driven by a specific T. gondii-secreted effector. Using a combination of bioinformatics and molecular genetics, we have now identified T. gondii GRA28 as the gene product required for CCL22 induction. GRA28 is secreted into the host cell where it localizes to the nucleus, and deletion of this gene results in reduced CCL22 placental cells as well as a human monocyte cell line. The impact of GRA28 on CCL22 production is also conserved in mouse immune and placental cells both in vitro and in vivo. Moreover, parasites lacking GRA28 are impaired in their ability to disseminate throughout the animal, suggesting a link between CCL22 induction and the ability of the parasite to cause disease. Overall these data demonstrate a clear function for GRA28 in altering the immunomodulatory landscape during infection of both placental and peripheral immune cells, and show a clear impact of this immunomodulation on infection outcome. AUTHOR SUMMARYToxoplasma gondii is a globally ubiquitous pathogen that can cause severe disease in HIV/AIDS patients and can also cross the placenta and infect the developing fetus. We have found that placental and immune cells infected with T. gondii secrete significant amounts of a chemokine (called "CCL22") that is critical for immune tolerance during pregnancy. In order to better understand whether this is a response by the host or a process that is driven by the parasite, we have identified a T. gondii gene that is absolutely required to induce CCL22 production in human cells, indicating that CCL22 production is a process driven almost entirely by the parasite rather than the host. Consistent with its role in immune tolerance, we also found that T. gondii parasites lacking this gene are less able to proliferate and disseminate throughout the host. Taken together these data illustrate a direct relationship between CCL22 levels in the infected host and a key parasite effector, and provide an interesting example of how T. gondii can directly modulate host signaling pathways in order to facilitate its growth and dissemination.

microbiology↗

bioRxiv2019

Thermal stress responses of Sodalis glossinidius, an indigenous bacterial symbiont of hematophagous tsetse flies

Tsetse flies (Diptera: Glossinidae) house a taxonomically diverse microbiota that includes environmentally acquired bacteria, maternally transmitted symbiotic bacteria, and pathogenic African trypanosomes. Sodalis glossinidius, which is a facultative symbiont that resides intra and extracellularly within multiple tsetse tissues, has been implicated as a mediator of trypanosome infection establishment in the flys gut. Tsetses gut-associated population of Sodalis are subjected to marked temperature fluctuations each time their ectothermic fly host imbibes vertebrate blood. The molecular mechanisms that Sodalis employs to deal with this heat stress are unknown. In this study, we examined the thermal tolerance and heat shock response of Sodalis. When grown on BHI agar plates, the bacterium exhibited the most prolific growth at 25{degrees}C, and did not grow at temperatures above 30{degrees}C. Growth on BHI agar plates at 31{degrees}C was dependent on either the addition of blood to the agar or reduction in oxygen levels. Sodalis was viable in liquid cultures for 24 hours at 30{degrees}C, but began to die upon further exposure. The rate of death increased with increased temperature. Similarly, Sodalis was able to survive for 48 hours within tsetse flies housed at 30{degrees}C, while a higher temperature (37{degrees}C) was lethal. Sodalis genome contains homologues of the heat shock chaperone protein-encoding genes dnaK, dnaJ, and grpE, and their expression was up-regulated in thermally stressed Sodalis, both in vitro and in vivo within tsetse flies. Arrested growth of E. coli dnaK, dnaJ, or grpE mutants under thermal stress was reversed when the cells were transformed with a low copy plasmid that encoded the Sodalis homologues of these genes. The information contained in this study provides insight into how arthropod vector enteric commensals, many of which mediate their hosts ability to transmit pathogens, mitigate heat shock associated with the ingestion of a blood meal.\n\nAUTHOR SUMMARYMicroorganisms associated with insects must cope with fluctuating temperatures. Because symbiotic bacteria influence the biology of their host, how they respond to temperature changes will have an impact on the host and other microorganisms in the host. The tsetse fly and its symbionts represent an important model system for studying thermal tolerance because the fly feeds exclusively on vertebrate blood and is thus exposed to dramatic temperature shifts. Tsetse flies house a microbial community that can consist of symbiotic and environmentally acquired bacteria, viruses, and parasitic African trypanosomes. This work, which makes use of tsetses commensal symbiont, Sodalis glossinidius, is significance because it represents the only examination of thermal tolerance mechanisms in a bacterium that resides indigenously within an arthropod disease vector. A better understanding of the biology of thermal tolerance in Sodalis provides insight into thermal stress survival in other insect symbionts and may yield information to help control vector-borne disease.

microbiology↗