Convergent microevolution of Cryptococcus neoformans hypervirulence in the laboratory and the clinic

Arras, Samantha D. M.; Ormerod, Kate L.; Erpf, Paige E.; Espinosa, Monica I.; Carpenter, Alex C.; Blundell, Ross; Stowasser, Samantha R.; Schulz, Benjamin L.; Tanurdžić, Miloš; Fraser, James A.

doi:10.1038/s41598-017-18106-2

Cited by 37 publications

(45 citation statements)

References 59 publications

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“…Disruption of the sgf29 gene is not lethal, in line with studies in other organisms including S. cerevisae and C. neoformans [33,42] and is not required for Dictyostelium development. In C. neoformans, disruption of sgf29 leads to hypervirulence, and inability to form titan cells, and in Arabidopsis with salt tolerance, demonstrating important biological consequences of its loss [42][43][44]. Importantly disruption of the sgf29 gene successfully disrupts preferential H3K9/14 acetylation of H3K4me3 upon TSA treatment and also confers resistance to TSA during development.…”

Section: Overexpression Of Sgf29 Also Leads To Tsa Resistance During supporting

confidence: 78%

Methylation-directed Acetylation of Histone H3 Regulates Developmental Sensitivity to Histone Deacetylase Inhibition

Huang

Hsu

Pears

2020

Preprint

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BackgroundTreatment of cells with hydroxamate-based lysine deacetylase inhibitors (KDACis) such as Trichostatin A (TSA) can induce biological effects such as differentiation or apoptosis of cancer cells, and a number of related compounds have been approved for clinical use. TSA treatment induces rapid initial acetylation of histone 3 (H3) proteins which are already modified by tri-methylation on lysine 4 (H3K4me3) while acetylation of bulk histones, lacking this mark, is delayed. Sgf29, a subunit of the SAGA acetyltransferase complex, interacts with H3K4me3 via a tandem tudor domain (TTD) and has been proposed to target the acetyltransferase activity to H3K4me3. However the importance of acetylation of this pool of H3 in the biological consequences of KDACi treatment is not known.ResultsWe investigated the role of H3K4me3-directed acetylation in the mechanism of action of TSA on inhibiting development of the eukaryotic social amoeba Dictyostelium discoideum. Loss of H3K4me3 in strains with mutations in the gene encoding Set1 or the histone proteins confers resistance to TSA-induced inhibition of development and delays accumulation of histone acetylation on H3K9 and K14. A candidate orthologue of Sgf29 in Dictyostelium has been identified which specifically recognizes the H3K4me3 modification via its tandem Tudor domain (TTD). Disruption of the gene encoding Sgf29 delayed accumulation of H3K9Ac, abolished targeted H3K4me3-directed H3Ac and led to developmental resistance to TSA, which is dependent on a functional TTD. TSA resistance also results from overexpression of Sgf29.ConclusionPreferential acetylation of H3K4me3 histones, regulated by Sgf29 via its TTD, is important in developmental sensitivity to TSA. Levels of H3K4me3 or Sgf29 will provide useful biomarkers for sensitivity to this class of chemotherapeutic drug.

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Section: Overexpression Of Sgf29 Also Leads To Tsa Resistance During supporting

confidence: 78%

Methylation-directed Acetylation of Histone H3 Regulates Developmental Sensitivity to Histone Deacetylase Inhibition

Huang

Hsu

Pears

2020

Preprint

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“…Two genes, LMP1 and SGF29 , are dramatically affected by SNPs/indels in the H99 derivatives; LMP1 has a frameshift deletion (H99W and H99 CMO18) and SGF29 is deleted (KN99α and H99L) [ 35 ]. To determine if these genes are involved in titan cell production, we analyzed lmp1Δ and sgf29Δ deletion mutants for in vitro and in vivo titan cell formation ( Fig 7 and S8 and S9 Figs, respectively).…”

Section: Resultsmentioning

confidence: 99%

Titan cells formation in Cryptococcus neoformans is finely tuned by environmental conditions and modulated by positive and negative genetic regulators

et al. 2018

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The pathogenic fungus Cryptococcus neoformans exhibits morphological changes in cell size during lung infection, producing both typical size 5 to 7 μm cells and large titan cells (> 10 μm and up to 100 μm). We found and optimized in vitro conditions that produce titan cells in order to identify the ancestry of titan cells, the environmental determinants, and the key gene regulators of titan cell formation. Titan cells generated in vitro harbor the main characteristics of titan cells produced in vivo including their large cell size (>10 μm), polyploidy with a single nucleus, large vacuole, dense capsule, and thick cell wall. Here we show titan cells derived from the enlargement of progenitor cells in the population independent of yeast growth rate. Change in the incubation medium, hypoxia, nutrient starvation and low pH were the main factors that trigger titan cell formation, while quorum sensing factors like the initial inoculum concentration, pantothenic acid, and the quorum sensing peptide Qsp1p also impacted titan cell formation. Inhibition of ergosterol, protein and nucleic acid biosynthesis altered titan cell formation, as did serum, phospholipids and anti-capsular antibodies in our settings. We explored genetic factors important for titan cell formation using three approaches. Using H99-derivative strains with natural genetic differences, we showed that titan cell formation was dependent on LMP1 and SGF29 genes. By screening a gene deletion collection, we also confirmed that GPR4/5-RIM101, and CAC1 genes were required to generate titan cells and that the PKR1, TSP2, USV101 genes negatively regulated titan cell formation. Furthermore, analysis of spontaneous Pkr1 loss-of-function clinical isolates confirmed the important role of the Pkr1 protein as a negative regulator of titan cell formation. Through development of a standardized and robust in vitro assay, our results provide new insights into titan cell biogenesis with the identification of multiple important factors/pathways.

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“…The deletions affect the 5′ UTRs of two genes and eliminate SGF29 (CNAG_06392) almost completely (Table S1 in File S1 ). The loss of SFG29 in the KN99 lineage was recently observed by another group, which attributed the relatively high virulence of KN99 to it ( Arras et al 2017 ). The other major feature of the KN99α sequence was a 285 kb region (chromosome 11:175,000–460,000) that was highly diverged from H99α.…”

Section: Resultsmentioning

confidence: 56%

Unintended Side Effects of Transformation Are Very Rare in Cryptococcus neoformans

Friedman

Gish

Brown

et al. 2018

G3 Genes|Genomes|Genetics

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Received wisdom in the field of fungal biology holds that the process of editing a genome by transformation and homologous recombination is inherently mutagenic. However, that belief is based on circumstantial evidence. We provide the first direct measurement of the effects of transformation on a fungal genome by sequencing the genomes of 29 transformants and 30 untransformed controls with high coverage. Contrary to the received wisdom, our results show that transformation of DNA segments flanked by long targeting sequences, followed by homologous recombination and selection for a drug marker, is extremely safe. If a transformation deletes a gene, that may create selective pressure for a few compensatory mutations, but even when we deleted a gene, we found fewer than two point mutations per deletion strain, on average. We also tested these strains for changes in gene expression and found only a few genes that were consistently differentially expressed between the wild type and strains modified by genomic insertion of a drug resistance marker. As part of our report, we provide the assembled genome sequence of the commonly used laboratory strain Cryptococcus neoformans var. grubii strain KN99α.

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Convergent microevolution of Cryptococcus neoformans hypervirulence in the laboratory and the clinic

Cited by 37 publications

References 59 publications

Methylation-directed Acetylation of Histone H3 Regulates Developmental Sensitivity to Histone Deacetylase Inhibition

Methylation-directed Acetylation of Histone H3 Regulates Developmental Sensitivity to Histone Deacetylase Inhibition

Titan cells formation in Cryptococcus neoformans is finely tuned by environmental conditions and modulated by positive and negative genetic regulators

Unintended Side Effects of Transformation Are Very Rare in Cryptococcus neoformans

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