Transcription profile of dormant and germinating Aspergillus niger conidia
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ABSTRACT: Genome-wide analysis was performed to assess the transcriptional landscape of germinating A. niger conidia using GeneChips. The metabolism of storage compounds during conidial germination was also examined and compared to the transcript levels from associated genes. The transcriptome of dormant conidia was shown to be highly differentiated from that of germinating conidia and major changes in response to environmental shift occurred within the first hour of germination. The breaking of dormancy was associated with increased transcript levels of genes involved in the biosynthesis of proteins, RNA turnover and respiratory metabolism. Increased transcript levels of genes involved in metabolism of nitrate and proline at the onset of germination implies their use as sources of nitrogen. Dormant conidia contained transcripts of genes involved in fermentation, gluconeogenesis and the glyoxylate cycle. The presence of such transcripts in dormant conidia may indicate the generation of energy from non-carbohydrate substrates during starvation-induced conidiation or for maintenance purposes during dormancy. The immediate onset of metabolism of internal storage compounds after the onset of germination, and the presence of transcripts of relevant genes, suggest that conidia are primed for the onset of germination. one biological replicate for each sample, each sample contained pooled RNA from three independent replicates
Project description:Genome-wide analysis was performed to assess the transcriptional landscape of germinating A. niger conidia for first hour using next generation RNA-sequencing. The transcriptome of dormant conidia was shown to be highly differentiated from that of germinating conidia. The breaking of dormancy was associated with increased transcript levels of genes involved in the biosynthesis of proteins, RNA turnover and respiratory metabolism. Increased transcript levels of genes involved in metabolism of nitrate and proline at the onset of germination implies their use as sources of nitrogen. The transcriptome of dormant conidia contained a significant component of antisense transcripts that changed during germination. Dormant conidia contained transcripts of genes involved in fermentation, gluconeogenesis and the glyoxylate cycle. The presence of such transcripts in dormant conidia may indicate the generation of energy from non-carbohydrate substrates during starvation-induced conidiation or for maintenance purposes during dormancy. The immediate onset of metabolism of internal storage compounds after the onset of germination, and the presence of transcripts of relevant genes, suggest that conidia are primed for the onset of germination. For some genes, antisense transcription is regulated in the transition from resting conidia to fully active germinants. Duplicate samples from each time point: dormant conidia T0 (DA 21 + DA 22) and conidia germinated for one hour T1 (DA 23 + DA 24)
Project description:Conidia of Aspergillus niger are characterized by a dormant state and are moderate stress-resistant. Upon contact with a moist substrate, germination of conidia occurs by changing from a dormant stabilized state towards a growing vegetative cell. The RNA expression levels of dormant conidia and conidia that were in various stages of germination were studied. The RNA composition of dormant conidia was substantially different than all the subsequent stages of germination. This indicates that the distinct morphological changes that occur during germination are not correlated with the highest change in the transcriptome. Samples of germinating conidia of Aspergillus niger N402 were taken in triple at 0h (dormant), 2h, 4h, 6h and 8h after inoculation in CM.
Project description:Genome-wide analysis was performed to assess the transcriptional landscape of germinating A. niger conidia for first hour using next generation RNA-sequencing. The transcriptome of dormant conidia was shown to be highly differentiated from that of germinating conidia. The breaking of dormancy was associated with increased transcript levels of genes involved in the biosynthesis of proteins, RNA turnover and respiratory metabolism. Increased transcript levels of genes involved in metabolism of nitrate and proline at the onset of germination implies their use as sources of nitrogen. The transcriptome of dormant conidia contained a significant component of antisense transcripts that changed during germination. Dormant conidia contained transcripts of genes involved in fermentation, gluconeogenesis and the glyoxylate cycle. The presence of such transcripts in dormant conidia may indicate the generation of energy from non-carbohydrate substrates during starvation-induced conidiation or for maintenance purposes during dormancy. The immediate onset of metabolism of internal storage compounds after the onset of germination, and the presence of transcripts of relevant genes, suggest that conidia are primed for the onset of germination. For some genes, antisense transcription is regulated in the transition from resting conidia to fully active germinants.
Project description:Genome-wide analysis was performed to assess the transcriptional landscape of germinating A. niger conidia using GeneChips. The metabolism of storage compounds during conidial germination was also examined and compared to the transcript levels from associated genes. The transcriptome of dormant conidia was shown to be highly differentiated from that of germinating conidia and major changes in response to environmental shift occurred within the first hour of germination. The breaking of dormancy was associated with increased transcript levels of genes involved in the biosynthesis of proteins, RNA turnover and respiratory metabolism. Increased transcript levels of genes involved in metabolism of nitrate and proline at the onset of germination implies their use as sources of nitrogen. Dormant conidia contained transcripts of genes involved in fermentation, gluconeogenesis and the glyoxylate cycle. The presence of such transcripts in dormant conidia may indicate the generation of energy from non-carbohydrate substrates during starvation-induced conidiation or for maintenance purposes during dormancy. The immediate onset of metabolism of internal storage compounds after the onset of germination, and the presence of transcripts of relevant genes, suggest that conidia are primed for the onset of germination.
Project description:Conidia of Aspergillus niger are characterized by a dormant state and are moderate stress-resistant. Upon contact with a moist substrate, germination of conidia occurs by changing from a dormant stabilized state towards a growing vegetative cell. The RNA expression levels of dormant conidia and conidia that were in various stages of germination were studied. The RNA composition of dormant conidia was substantially different than all the subsequent stages of germination. This indicates that the distinct morphological changes that occur during germination are not correlated with the highest change in the transcriptome.
Project description:The impact of natamycin on Aspergillus niger was analysed during 8 h of germination of conidia. Polarization, germ tube formation, and mitosis were inhibited in the presence of 3 M-BM-5M and 10 M-BM-5M of the anti fungal compound natamycin, while at 10 M-BM-5M also isotropic growth was affected. Natamycin did not have an effect on the decrease of internal microviscosity and the concomitant reduction on mannitol and trehalose levels. However, it did abolish the increase of intracellular levels of glycerol and glucose during the 8 h period of germination. Natamycin hardly affected the changes that occur in the RNA profile during the first 2 h of germination. During this time period, genes related to transcription, protein synthesis, energy and cell cycle and DNA processing were particularly up regulated. On the other hand, when 8 h old germlings were compared, genes involved in ergosterol biosynthesis were down regulated while genes related to endocytosis and metabolism of compatible solutes, and genes encoding protective proteins were up regulated. Samples of germinating conidia of Aspergillus niger N402 were taken in triplo at 0h (dormant), 2h, and 8h after inoculation in CM. These were compared with conidia that germinated in the presence of either 3 M-BM-5M or 10 M-BM-5M natamycin.
Project description:The rice pathogen, Magnaporthe oryzae, undergoes a complex developmental process leading to formation of an appressorium prior to plant infection. In an effort to better understand phosphoregulation during appressorium development, a mass spectrometry-based study was undertaken. Phosphosites were identified from phosphoproteins from mycelia, conidia, germlings, and appressoria of the wild type and a ckpA mutant. The cyclic-AMP dependent protein kinase A (CPKA) is required for initial perception of the leaf surface, development of functional appressoria, and timely mobilization of storage reserves in conidia, including glycogen and lipid bodies.
Project description:The outer layers of the fungal cell wall are the first barrier facing external challenges. In theinvasive morphotype of the airborne pathogen Aspergillus fumigatus, i.e. conidia, the outer cell wall consists of α-(1,3)-glucan, melanin and proteinaceous rodlets made up by the RodA hydrophobin. The hydrophobicity of the RodA rodlets facilitates air dispersal of conidia and progression within lungs. When conidia reach the alveolae, these are phagocytosed by host cells and killed upon germination, which requires melanin and rodlets elimination to occur. The dormant condition of the conidia protects them against activation of the host killing pathways. The outer cell wall of conidia is thus essential for maintaining the viability of the fungus. To study the role of α-(1,3)-glucan, melanin, and RodA rodlets in conidia protection, multiple mutants without two or the three major components of the outer layer were constructed and analyzed. Deletions led to the exposure of new molecules on the conidial surface. Multiple gene deletions did not always lead to logical additivity of the phenotypes. Unexpected compensatory cell wall surface modifications were indeed observed, such as the synthesis of the mycelial virulence factor galactosaminogalactan, the presence of chitin and glycoproteins or specific changes in permeability. In spite of significant modifications, sensitivity to killing by phagocytic host cells of the multiple mutants involving melanin changed little compared to the sensitive parent strain lacking melanin (ΔpksP), further indicating the importance of melanin in protecting conidia against killing by monocytes.
Project description:The response of AM transcription to exposure to conidia is expected to provide information about the mechanism by which these cells prevent conidial germination and therefore invasive aspergillosis. Experiment Overall Design: Conidia from A. fumigatus or polystyrene beads were instilled into lungs of C57Bl/6 or gp91phox-/- mice to test the response of AM in a normal and immune compromised host to determine if different responses contributed to increased susceptibility of invasive aspergillosis. Following in vivo incubation for 0, 2, or 4 hours, AM RNA was collected and prepared for hybridization to mouse Affymetrix GeneChip arrays.
Project description:Aspergillus fumigatus is a filamentous fungus capable to cause an important disease known as invasive aspergillosis. Challenge different cell lines is a crucial strategy to undercover new virulence factors involved in the infection process. In this research, RAW 264.7 macrophages and A549 lung epithelial cells were challenge using A. fumigatus conidia in a MOI of 10 and 5 respectively. When these conidia reach 30% of germination, the total RNA was isolated (A. fumigatus alone (Control), A. fumigatus vs RAW 264.7, and A. fumigatus vs A549) and purified using the RNeasy Plant Mini Kit (Qiagen). The AWAFUGE (Agilent Whole A. fumigatus Genome Expression 44K v.1) hybridization was carried out following previously publish protocols (A-MEXP-2352 and E-MTAB-5314).