Project description:RNA-seq from Neurospora crassa at 5 time points of light induction, with 2 replicates for each, totalling 10 samples RNA-seq from Neurospora crassa at 5 time points of light induction, with 2 replicates for each, totalling 10 samples
Project description:Cryptochromes were identified in plants and animals where they function as either photoreceptors or circadian clock components. In the filamentous fungus Neurospora, the biological function of cryptochrome has not yet been explored. Here, we demonstrate that Neurospora crassa cryptochrome (Nc cry) is a DASH-type of cryptochrome, capable of binding FAD and MTHF, whose transcript and protein levels are both strongly induced by blue light in a wc-1 dependent manner. Although the Nc cry transcript is circadian-regulated and antiphasic to frq, knockout strains of Nc cry appears to have a normal clock phenotype. Whole genome microarray and RT-QPCR analysis confirm that Nc cry is not involved in the signal transduction of either early or late light responses and seems to have no transcriptional regulatory activity under our laboratory conditions. Our study concludes that the only cryptochrome in Neurospora crassa is dispensable for the well-characterized blue light sensing cascade and is not part of the circadian clock system. Keywords: light response
Project description:Cryptochromes were identified in plants and animals where they function as either photoreceptors or circadian clock components. In the filamentous fungus Neurospora, the biological function of cryptochrome has not yet been explored. Here, we demonstrate that Neurospora crassa cryptochrome (Nc cry) is a DASH-type of cryptochrome, capable of binding FAD and MTHF, whose transcript and protein levels are both strongly induced by blue light in a wc-1 dependent manner. Although the Nc cry transcript is circadian-regulated and antiphasic to frq, knockout strains of Nc cry appears to have a normal clock phenotype. Whole genome microarray and RT-QPCR analysis confirm that Nc cry is not involved in the signal transduction of either early or late light responses and seems to have no transcriptional regulatory activity under our laboratory conditions. Our study concludes that the only cryptochrome in Neurospora crassa is dispensable for the well-characterized blue light sensing cascade and is not part of the circadian clock system. Keywords: light response Two-color microarray. Alexa Fluor 555 was consistently used to label cDNA synthesized from reference RNA, which is a mixture containing equal amounts of RNA samples harvested from different circadian time points and light treatment durations. The same batch of pooled RNA was used as a reference for each array experiment. Alexa Fluor 647 was used exclusively to label cDNA representing sample RNA.
Project description:Light represents an important environmental cue, which exerts considerable influence on the metabolism of fungi. Studies with the biotechnological fungal workhorse Trichoderma reesei (Hypocrea jecorina) have revealed an interconnection between transcriptional regulation of cellulolytic enyzmes and the light response. The filamentous fungus, Neurospora crassa, has been used as a model organism to study light and circadian rhythm biology. We therefore investigated whether light also regulates transcriptional regulation of cellulolytic enzymes in N. crassa. We show that the N. crassa photoreceptor genes wc-1, wc-2 and vvd are involved in regulation of cellulase gene expression, indicating that this phenomenon is conserved among filamentous fungi. Genome wide analysis of photoreceptor mutants and evaluation of results by analysis of mutant strains identified several candidate genes likely to play a role in light modulated cellulase gene expression.
Project description:Multi-targeting priming (MTP) for genome-wide gene expression assays provides selective targeting of multiple sequences and counter-selection against undesirable sequences. We demonstrated superior performance of two MTPs compared to oligo-dT microarray profling and RNA tag sequencing the response of Saccharomyces cerevisiae to nitrogen deficiency and profiling Neurospora crassa early sexual development. Priming with MTPs in addition to oligo-dT resulted in higher sensitivity, a greater number of well-measured genes, more genes significantly differentially expressed, and a greater power to detect meager differences. Neurospora crassa mat A FGSC#2489 2 developmental stages and oligo(dT) primers.
Project description:To determine the genes directly and indirectly under the control of the Grainy-head homolog (GHH) transcription factor in Neurospora crassa Three different sample types (Aerial Hyphae & Conidia; Mycelia; or Whole Colonies) of both wild-type (FGSC #2489) and grainy-head homolog (FGSC #13563) strains of Neurospora crassa were subjected to transcriptome analyses to determine the genes differentially expressed in the ghh background compared to wild type.
Project description:Many fungi form complex three-dimensional fruiting bodies, within which the meiotic machinery for sexual spore production has been considered to be largely conserved over evolutionary time. Indeed, much of what we know about meiosis in plant and animal taxa has been deeply informed by studies of meiosis in Saccharomyces and Neurospora. Nevertheless, the genetic basis of fruiting body development and its regulation in relation to meiosis in fungi is barely known, even within the best studied multicellular fungal model Neurospora crassa. We characterized morphological development and genome-wide transcriptomics in the closely related species Neurospora crassa, Neurospora tetrasperma, and Neurospora discreta, across eight stages of sexual development. Despite diverse life histories within the genus, all three species produce vase-shaped perithecia. Transcriptome sequencing provided gene expression levels of 2479 orthologous genes among all three species. Expression of key meiosis genes and sporulation genes, corresponded to developmental differences among these Neurospora species during sexual development. Screening N. crassa knockout crosses of genes selected for their expression differences across species, eight genes, whose functions were previously unknown, are found to be critical for the successful formation of perithecia. The absence of these genes in mutant crosses resulted in either no perithecium formation or in arrested development at an early stage. Our results provide insight into the genetic basis of Neurospora sexual reproduction, which is also of great importance with regard to other multicelluar ascomycetes, including fungal pathogens closely related to Neurospora in the Sordariomycetes, such as Fusarium spp, Magnaporthe oryzae, and Nectria haematococca mRNA were sampled and compared from eight time points across sexual reproduction in three Neurospora species
Project description:Histones are not statically embedded, but are constantly exchanged outside of DNA replication. This study reports the characterization and validation of a histone turnover reporter strain of Neurospora crassa, and the method employed. This strain utilizes FLAG-tagged histone H3 under the control of a light-inducible promoter. This study also preliminarily explores histone turnover defects at constitutive heterochromatin with the loss of the heterochromatin proteins DIM-2, HDA-1, DIM-5, and HPO.
Project description:Light represents an important environmental cue, which exerts considerable influence on the metabolism of fungi. Studies with the biotechnological fungal workhorse Trichoderma reesei (Hypocrea jecorina) have revealed an interconnection between transcriptional regulation of cellulolytic enyzmes and the light response. The filamentous fungus, Neurospora crassa, has been used as a model organism to study light and circadian rhythm biology. We therefore investigated whether light also regulates transcriptional regulation of cellulolytic enzymes in N. crassa. We show that the N. crassa photoreceptor genes wc-1, wc-2 and vvd are involved in regulation of cellulase gene expression, indicating that this phenomenon is conserved among filamentous fungi. Genome wide analysis of photoreceptor mutants and evaluation of results by analysis of mutant strains identified several candidate genes likely to play a role in light modulated cellulase gene expression. Three deletion strains (delta-wc-1 (FGSC 11712), delta-wc-2 (FGSC 11124) and delta-vvd (FGSC 11556)) and the wild type strain (FGSC 2489) at two different timepoints (28h or 40h) were analyzed. Cy3 and Cy5 dye swaps were performed.