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Activation of Meiotic Genes Mediates Ploidy Reduction during Cryptococcal Infection.

ABSTRACT: Cryptococcus neoformans is a global human fungal pathogen that causes fatal meningoencephalitis in mostly immunocompromised individuals. During pulmonary infection, cryptococcal cells form large polyploid cells that exhibit increased resistance to host immune attack and are proposed to contribute to the latency of cryptococcal infection. These polyploid titan cells can generate haploid and aneuploid progeny that may result in systemic infection. What triggers cryptococcal polyploidization and how ploidy reduction is achieved remain open questions. Here, we discovered that Cryptococcus cells polyploidize in response to genotoxic stresses that cause DNA double-strand breaks. Intriguingly, meiosis-specific genes are activated in C. neoformans and contribute to ploidy reduction, both in vitro and during infection in mice. Cryptococcal cells that activated their meiotic genes in mice were resistant to specific genotoxic stress compared to sister cells recovered from the same host tissue but without activation of meiotic genes. Our findings support the idea that meiotic genes, in addition to their conventional roles in classic sexual reproduction, contribute to adaptation of eukaryotic cells that undergo dramatic genome changes in response to genotoxic stress. The discovery has additional implications for evolution of sexual reproduction and the paradox of the presence of meiotic machinery in asexual species. Finally, our findings in this eukaryotic microbe mirror the revolutionary discoveries of the polyploidization and meiosis-like ploidy reduction process in cancer cells, suggesting that the reversible ploidy change itself could provide a general mechanism for rejuvenation to promote individual survival in response to stress.

SUBMITTER: Zhao Y

PROVIDER: S-EPMC7228024 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BACKGROUND:Cryptococcal meningitis (CM) causes an estimated 180,000 deaths annually, predominantly in sub-Saharan Africa, where most patients receive fluconazole (FLC) monotherapy. While relapse after FLC monotherapy with resistant strains is frequently observed, the mechanisms and impact of emergence of FLC resistance in human CM are poorly understood. Heteroresistance (HetR) - a resistant subpopulation within a susceptible strain - is a recently described phenomenon in Cryptococcus neoformans (Cn) and Cryptococcus gattii (Cg), the significance of which has not previously been studied in humans. METHODS:A cohort of 20 patients with HIV-associated CM in Tanzania was prospectively observed during therapy with either FLC monotherapy or in combination with flucytosine (5FC). Total and resistant subpopulations of Cryptococcus spp. were quantified directly from patient cerebrospinal fluid (CSF). Stored isolates underwent whole genome sequencing and phenotypic characterization. RESULTS:Heteroresistance was detectable in Cryptococcus spp. in the CSF of all patients at baseline (i.e., prior to initiation of therapy). During FLC monotherapy, the proportion of resistant colonies in the CSF increased during the first 2 weeks of treatment. In contrast, no resistant subpopulation was detectable in CSF by day 14 in those receiving a combination of FLC and 5FC. Genomic analysis revealed high rates of aneuploidy in heteroresistant colonies as well as in relapse isolates, with chromosome 1 (Chr1) disomy predominating. This is apparently due to the presence on Chr1 of ERG11, which is the FLC drug target, and AFR1, which encodes a drug efflux pump. In vitro efflux levels positively correlated with the level of heteroresistance. CONCLUSION:Our findings demonstrate for what we believe is the first time the presence and emergence of aneuploidy-driven FLC heteroresistance in human CM, association of efflux levels with heteroresistance, and the successful suppression of heteroresistance with 5FC/FLC combination therapy. FUNDING:This work was supported by the Wellcome Trust Strategic Award for Medical Mycology and Fungal Immunology 097377/Z/11/Z and the Daniel Turnberg Travel Fellowship.

Project description:Intrauterine Listeria monocytogenes (L. monocytogenes) infections pose a major threat during pregnancy via affecting placental immune responses. However, the underlying mechanisms of placental defense against this pathogen remain ill-defined. Therefore, this study aims to investigate the function and the mechanism of inflammasomes on against L. monocytogenes infection during pregnancy. A listeriosis murine model and cell culture system was used to investigate the role of trophoblastic nucleotide-binding oligomerization domain-like receptor pyrin domain-containing 3 (NLRP3) in orchestrating innate immune responses to L. monocytogenes infection. Caspase-1 activity was determined using a caspase-1 activity colorimetric kit. NLRP3 and apoptosis-associated speck-like protein containing a CARD (ASC) in placental tissue was detected by immunohistochemistry. NLRP3 in HTR-8/SVneo cells was also detected by immunofluorescence. The expression of interleukin 1β (IL-1β), NLRP3, ASC, and caspase-1 was detected by Western blot. We characterized the NLRP3 inflammasome in trophoblast cells according to whether L. monocytogenes infection increased the activation of caspase-1 and the release of IL-1β. For human or mouse IL-1β in the culture supernatants and mouse tissue lysates were analyzed using ELISA Kits. Trophoblast cells constitutively expressed the components of the NLRP3 inflammasome. In vitro, L. monocytogenes triggers NLRP3 inflammasome activation in trophoblast cells by inducing caspase-1 activation, increasing the NLRP3 protein levels, IL-1β maturation and secretion in HTR-8/SVneo cells. In vivo, L. monocytogenes induces fetal resorption and IL-1β processing in pregnant mice. In addition, NLRP3-deficient mice were more prone to fetal loss than their wild-type counterparts following infection with L. monocytogenes at a lower infective dose. We conclude that trophoblast cells respond to L. monocytogenes infection through the NLRP3 receptor, resulting in inflammasome activation and IL-1β production, which prevents listeriosis during pregnancy.

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Activation of Meiotic Genes Mediates Ploidy Reduction during Cryptococcal Infection.

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