Dataset Information

Histoplasma

ABSTRACT: Histoplasma samples from India, Raw sequence reads

PROVIDER: PRJNA1063273 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR27481863_1.fastq.gz	Fastqsanger.gz
	SRR27481863_2.fastq.gz	Fastqsanger.gz
	SRR27481864_1.fastq.gz	Fastqsanger.gz
	SRR27481864_2.fastq.gz	Fastqsanger.gz
	SRR27481865_1.fastq.gz	Fastqsanger.gz

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Project description:During infection, Histoplasma capsulatum exists primarily as an intracellular pathogen of phagocytes where it overcomes host defense mechanisms and replicates in the phagosome. This intracellular proliferation of Histoplasma yeasts requires the function of the fungal peroxisome for multiple, incompletely defined roles, with early pathogenesis functions dependent on the proper trafficking of peroxisome matrix proteins carrying a type 1 peroxisome-targeting signal (PTS1). To better understand the potential roles of peroxisomes during Histoplasma infection, we identified the constituents of the peroxisome proteome of pathogenic phase Histoplasma yeasts through proximity labeling using a peroxisome-targeted TurboID biotin ligase. Comparative proteomics using peroxisome- or cytosol-localized TurboID identified 90 high confidence peroxisome proteins. To validate the peroxisome proteome, a subset of nine putative peroxisome proteins were localized to the peroxisome using fluorescent protein fusions. To empirically define which matrix proteins of the peroxisome proteome depend on the PTS1 or PTS2 import pathways, peroxisome proteomes were also defined for Histoplasma yeasts lacking the Pex5 or Pex7 cytosolic PTS1 or PTS2 receptors. This assigned over half of the proteins to either the PTS1- or PTS2-dependent peroxisome import pathways and refined the Histoplasma PTS1 tripeptide consensus sequence. The peroxisomal proteome of Histoplasma yeast during macrophage infection was ultimately determined to provide relevance to Histoplasma’s primary pathogenic niche. The identities of the proteins comprising the peroxisome proteome include several enzymes for canonical peroxisome-localized biochemical pathways (e.g., fatty acid utilization and siderophore biosynthesis) but also represent several potential novel functions for the organelle. The defined peroxisome proteome of pathogenic-phase Histoplasma yeasts, particularly the set of proteins imported via the PTS1 import pathway, thus establishes the important foundation for understanding how peroxisomes promote intracellular pathogenesis.

Project description:The fungal pathogen Histoplasma capsulatum is thought to be the most common cause of fungal respiratory infections in immunocompetent humans, yet little is known about its biology. Here we provide the first genome-wide studies to experimentally validate its genome annotation. A functional interrogation of the Histoplasma genome provides critical support for continued investigation into the biology and pathogenesis of H. capsulatum and related fungi. We employed a three-pronged approach to provide a functional annotation for the H. capsulatum G217B strain. First, we probed high-density tiling arrays with labeled cDNAs from cells grown under diverse conditions. These data defined 6,172 transcriptionally active regions (TARs), providing validation of 6,008 gene predictions. Interestingly, 22% of these predictions showed evidence of anti-sense transcription. Additionally, we detected transcription of 264 novel genes not present in the original gene predictions. To further enrich our analysis, we incorporated expression data from whole-genome oligonucleotide microarrays. These expression data included profiling under growth conditions that were not represented in the tiling experiment, and validated an additional 2,249 gene predictions. Finally, we compared the G217B gene predictions to other available fungal genomes, and observed that an additional 254 gene predictions had an ortholog in a different fungal species, suggesting that they represent genuine coding sequences. These analyses yielded a high confidence set of validated gene predictions for H. capsulatum. The transcript sets resulting from this study are a valuable resource for further experimental characterization of this ubiquitous fungal pathogen. The data is available for interactive exploration at http://histo.ucsf.edu. The non-redundant genome sequence of Histoplasma capsulatum G217B was tiled over a set of 93 CombiMatrix arrays, which were then hybridized with labeled cDNA samples from yeast-form (red channel) or mycelial-form (green channel) Histoplasma. Due to low yields from the mycelial samples, only the red channel intensities were analyzed (and the red foreground intensities are, therefore, reported in the VALUE column for each sample). Rather than normalizing intensities across arrays, each probe was evaluated as detected or undetected relative to the negative control intensities on the corresponding array, and the density of detected probes as a function of genome position was used for the remaining analysis.

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Histoplasma

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