Abstract
Cryptococcus gattii is a human and animal pathogen that infects healthy hosts and caused the Pacific Northwest outbreak of cryptococcosis. The inhalation of infectious propagules can lead to internalization of cryptococcal cells by alveolar macrophages, a niche in which C. gattii cells can survive and proliferate. Although the nutrient composition of macrophages is relatively unknown, the high induction of amino acid transporter genes inside the phagosome indicates a preference for amino acid uptake instead of synthesis. However, the presence of countable errors in the R265 genome annotation indicates significant inhibition of transcriptomic analysis in this hypervirulent strain. Thus, we analyzed RNA-Seq data from in vivo and in vitro cultures of C. gattii R265 to perform the reannotation of the genome. In addition, based on in vivo transcriptomic data, we identified highly expressed genes and pathways of amino acid metabolism that would enable C. gattii to survive and proliferate in vivo. Importantly, we identified high expression in three APC amino acid transporters as well as the GABA permease. The use of amino acids as carbon and nitrogen sources, releasing ammonium and generating carbohydrate metabolism intermediaries, also explains the high expression of components of several degradative pathways, since glucose starvation is an important host defense mechanism.
Highlights
Cryptococcus gattii, along with its sibling species Cryptococcus neoformans, are the etiological agents of cryptococcosis, a life-threatening disease that affects the lungs and central nervous system
We noted that only 65.04% of the reads aligned to genes, and 18.47% of the reads aligned to introns and intergenic regions. This led us to speculate that the current genomic annotation of C. gattii might contains errors, confirming our previous findings for the ZAP1 gene [20]
Using our RNA-Seq dataset from three different growth conditions (C. gattii WT and ZIP1 null mutant exposed to a low zinc environment for 2 h, as well as cryptococcal cells recovered from bronchoalveolar lavage (BAL)), we executed the CodingQuarry pipeline to generate new gene models for the R265 strain of C. gattii, which consisted of three steps (Figure 1)
Summary
Cryptococcus gattii, along with its sibling species Cryptococcus neoformans, are the etiological agents of cryptococcosis, a life-threatening disease that affects the lungs and central nervous system. Cryptococcal cells are considered a facultative intracellular pathogen of neutrophils and dendritic cells, defense cells which act during the early stages of murine pulmonary infection [8]. These innate immune cells have antigen-presenting activity and produce proinflammatory cytokines, inducing an adaptive immune response in the host [8,9]. The capability of Cryptococcus cells to avoid phagocytosis, as well as to survive and replicate at a high rate inside phagocytes, contributes to the dissemination of fungal cells from the lung via the bloodstream [10,11]. Phagolysosomal membrane damage, in turn, enables Cryptococcus to escape the phagolysosome, accessing the nutrients in the cytoplasm [11]
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