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RNA sequencing of germinating Rhizopus delemar strain RA 99-880

ABSTRACT: Rhizopus delemar is an invasive fungal pathogen, responsible for the frequently fatal disease mucormycosis. Germination, a crucial mechanism by which spores of Rhizopus delemar infect and cause disease, is a key developmental process that transforms the dormant spore state into a vegetative one. Understanding the molecular mechanisms which underpin this transformation may be key to controlling mucormycosis; however, the regulation of germination remains poorly understood. This study describes the transcriptional changes which take place over the course of germination.

ORGANISM(S): Rhizopus delemar

PROVIDER: GSE114842 | GEO | 2018/08/09

REPOSITORIES: GEO

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Project description:Mucormycosis is a life-threatening disease especially in immunocompromised patients that was caused my mucoralean fungi. The rate of mortality is tremendously increased in the last decades due to the lack of appropriate diagnostic tools, insufficient knowledge about the immune response toward the mucormycosis and unavailability of specific antifungal drugs. Several species of mucoralean fungi cause mucormycosis such as Lichtheimia, Rhizopus, and Mucor. Lichtheimia species ranks the second and third cause of mucormycosis in Europe and the USA, respectively. In this study, we investigated the receptors present on the surface of immune cells that bind to the spores of Lichtheimia. We focus on two strains of L. corymbifera (FSU:9682 and FSU:10164) using resting and heat-killed spores. Additionally, we choose alveolar macrophages (MH-S) to carry out our experiment. MH-S is the first line of defense in the lung and the major component in the innate immune system. MH-S surface proteins were biotinylated and incubated with Lichtheimia spores. The surface proteins and putative binding partners were enriched by streptavidin. LC-MS/MS analysis showed that several proteins are highly expressed in presence of Lichtheimia spores, of which the heat shock protein family A (HSPA8) was one of the most abundant proteins. FACS analysis and immunofluorescence examination confirmed that HSPA8 is highly abundant on the surface of the MH-S, but not on the surface of Lichtheimia spores. Moreover, our study showed that the intensity of HSPA8 on the surface of MH-S depends on the multiplicity of infection (MOI). Additionally, the blocking with anti-HSPA8 antibody reduced the capability of MH-S to engulf the Lichtheimia spores, but not Aspergillus fumigatus spores. This confirms that HSPA8 is specific to Lichtheimia. THis is the first study addressing the determination of surface receptors of alveolar macrophages that in the context of Mucoralean fungi.

Project description:Description Bacillus subtilis forms highly resistant, metabolically inactive spores upon nutrient limitation. These endospores pose challenges to the food and medical sectors. Spores reactivate their metabolism upon contact with germinants through germination and outgrowth, and then develop into vegetative cells. However, the mechanism of the activation of the molecular machinery that triggers spore germination and outgrowth is unclear. To gain further insight into spore germination and outgrowth, the transcriptome and proteome changes during Bacillus subtilis spore conversion to vegetative cells were analyzed. The transcriptome analysis also allowed us to trace the different functional groups of genes expressed during this conversion. For each time-point sampled, the change in the spore proteome was quantitatively monitored relative to the reference proteome of 15N metabolically labelled vegetative cells. Of the quantified proteins, 60 percent are common to vegetative cells and spores, indicating that spores have a minimal set of proteins sufficient for the resumption of metabolism upon completion of germination. The shared proteins thus represent the most basic survival kit for spore-based life. Until the phase transition, defined as the completion of germination, we observed no significant change in the proteome or the transcriptome. Our analysis identified 34 abundant mRNA transcripts in the dormant spores, at least 31 of which were rapidly degraded after the phase transition. We observed 3152 differentially expressed genes, and demonstrated with our mass spectrometry analyses the differential expression of 323 proteins. Our data show that 173 proteins from dormant spores, both proteins unique to spores and proteins shared with vegetative cells, are lost after completion of germination. Further analysis is required to functionally interpret the observed protein loss. The observed diverse timing of the synthesis of different protein sets reveals a putative core-strategy of the revival of life starting from the B. subtilis spore.

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RNA sequencing of germinating Rhizopus delemar strain RA 99-880

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