Biology subjects

Alrubaye, H. S.

Publications and source records attributed to Alrubaye, H. S..

3 recordsLinked to original sources

bioRxiv2025

A single-cell atlas of Toxoplasma sexual development identifies regulators of gametogenesis

Toxoplasma gondii undergoes sexual development exclusively in the feline intestine, a process critical for genetic diversity and population expansion. Recent studies have identified genes critical in suppressing presexual development and metabolic differences in felines that may promote sexual development, but to date the gene regulatory networks driving development in the cat are unknown. To investigate this, we performed single-cell transcriptomics on parasites isolated from cat intestines, using fluorescent reporter strains and flow cytometry. From 15,068 cells across two experiments, we identified rare populations, including cells that bear all of the hallmarks of gametes. Candidate genes emerging from this study were tested via CRISPR-Cas9 Perturb-seq, identifying AP2X6 as a regulator of macrogametocyte development. Our single-cell data extends what is known about gene expression changes throughout sexual development and should be useful to those in the field working towards inducing gametogenesis, mating, and oocyst production in vitro.

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↗