Transcriptional heterogeneity ofCryptococcus gattiiVGII compared with non-VGII lineages underpins key pathogenicity pathways
Authors: Farrer Rhys A. , Ford Christopher B. , Rhodes Johanna , Delorey Toni , May Robin C. , Fisher Matthew C. , Cloutman-Green Elaine , Balloux Francois , Cuomo Christina A. 
Published: August 2018

Cryptococcus gattiiis a pathogenic yeast of humans and other animals, which causes disease predominantly in immunocompetent hosts. Infection begins when aerosolized yeast or spores enter the body, triggering an immune response, including engulfment by macrophages. To understand the early transcriptional signals in both the yeast and its mammalian host, we performed a time-course dual RNA-seq experiment for four lineages ofC. gattii(VGI-IV) interacting with mouse macrophages at 1hr, 3hr and 6hr post infection. Comparison ofin vitrotoex vivogene expression indicates lineage VGII is transcriptionally divergent to non-VGII lineages, including differential expression of genes involved in capsule synthesis, capsule attachment and ergosterol production. Various paralogs demonstrate sub-functionalisation between lineages including an upregulation of capsule biosynthesis-related geneCAP2, and downregulation ofCAP1in VGIII. Isolates also compensate for lineage-specific gene-losses by over-expression of genetically similar paralogs, including an over-expression of capsule geneCAS3in VGIV having lostCAS31. Differential expression of one in fiveC. gattiigenes was detected following co-incubation with mouse macrophages; all isolates showed high induction of oxidative-reduction functions and a downregulation of capsule attachment genes. We also show that VGII switches expression of two laccase paralogs (fromLAC1toLAC2) during co-incubation of macrophages. Finally, we found that mouse macrophages respond to all four lineages ofC. gattiiby upregulating FosB/Jun/Egr1 regulatory proteins at early time points. This study highlights the evolutionary breadth of expression profiles amongst the lineages ofC. gattiiand the diversity of transcriptional responses at this host-pathogen interface.

Importance

The transcriptional profiles of related pathogens and their response to host induced stresses underpin their pathogenicity. Expression differences between related pathogens during host interaction can indicate when and how these genes contribute to virulence, ultimately informing new and improved treatment strategies for those diseases. In this paper, we compare the transcriptional profiles of five isolates representing four lineages ofC. gattiiin rich media. Our analyses identified key processes including cell capsule, ergosterol production and melanin that are differentially expressed between lineages, and we find that VGII has the most distinct profile in terms of numbers of differentially expressed genes. All lineages have also undergone sub-functionalisation for various paralogs including capsule biosynthesis and attachment genes. Most genes appeared down-regulated during co-incubation with macrophages, with the largest decrease observed for capsule attachment genes, which appears coordinated with a stress response, as all lineages also upregulated oxidative stress response genes. Furthermore, VGII upregulated many genes that are linked to ergosterol biosynthesis and switched expression of the laccaseLAC1toLAC2 ex vivo. Finally, we saw a pronounced increase in the FosB/Jun/Egr1 regulatory proteins at early time points in bone marrow derived macrophages, marking a role in the host response toC. gattii. This work highlights the dynamic roles of keyC. gattiivirulence genes in response to macrophages.