A Light-Inducible Strain for Genome-Wide Histone Turnover Profiling in Neurospora crassa
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ABSTRACT:
PROVIDER: PRJNA601092 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR10883648.fastq.gz | Fastqsanger.gz | |||
| SRR10883649.fastq.gz | Fastqsanger.gz | |||
| SRR10883650.fastq.gz | Fastqsanger.gz | |||
| SRR10883651.fastq.gz | Fastqsanger.gz | |||
| SRR10883652.fastq.gz | Fastqsanger.gz |
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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.
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Project description:To better understand the roles of WC-1, WC-2 and other putative photoreceptors in mediation of light signals and to identify new components in light sensing cascades, we used microarrays with genome-wide coverage to characterize light-inducible transcriptional changes in Neurospora crassa. After unsupervised hierarchical clustering of 90 microarrays, 316 genes, representing about 3.21% of the total genome, showed robust and consistent light responses. As expected, WC-1 and WC-2 knockout strains both exhibited severe impairments in response to a continuous white light stimulus, confirming their dominant roles in light sensing cascades. Knockout strains of 5 newly-identified light-inducible transcription factors were further characterized for their potential function in mediating light signals. Interestingly, one early light-inducible transcription factor, Submerged Protoperithecia-1 (SUB-1), appeared to be essential in mediating a large number of late light response, providing a direct molecular connection between early and late light responses. Furthermore, a novel motif, which is overrepresented among late light responsive genes, was demonstrated to be necessary for full induction of late light responses in vivo. Taken together, these results reveal a simple hierarchical light-sensing cascade in Neurospora, which is initially invoked by light-activated White Collar Complex (WCC) binding to Light Response Elements (LREs) of the primary target genes, including SUB-1. Subsequently, the induction of SUB-1 and other activators activate downstream targets. Thus, light responses of well-defined timing and amplitude can be orchestrated to meet the differential needs of underlying biological processes. Keywords: time course, light response
2009-03-09 | GSE8932 | GEO
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
2014-08-01 | E-GEOD-53534 | biostudies-arrayexpress
Project description:To better understand the roles of WC-1, WC-2 and other putative photoreceptors in mediation of light signals and to identify new components in light sensing cascades, we used microarrays with genome-wide coverage to characterize light-inducible transcriptional changes in Neurospora crassa. After unsupervised hierarchical clustering of 90 microarrays, 316 genes, representing about 3.21% of the total genome, showed robust and consistent light responses. As expected, WC-1 and WC-2 knockout strains both exhibited severe impairments in response to a continuous white light stimulus, confirming their dominant roles in light sensing cascades. Knockout strains of 5 newly-identified light-inducible transcription factors were further characterized for their potential function in mediating light signals. Interestingly, one early light-inducible transcription factor, Submerged Protoperithecia-1 (SUB-1), appeared to be essential in mediating a large number of late light response, providing a direct molecular connection between early and late light responses. Furthermore, a novel motif, which is overrepresented among late light responsive genes, was demonstrated to be necessary for full induction of late light responses in vivo. Taken together, these results reveal a simple hierarchical light-sensing cascade in Neurospora, which is initially invoked by light-activated White Collar Complex (WCC) binding to Light Response Elements (LREs) of the primary target genes, including SUB-1. Subsequently, the induction of SUB-1 and other activators activate downstream targets. Thus, light responses of well-defined timing and amplitude can be orchestrated to meet the differential needs of underlying biological processes. Keywords: time course, light response Two-color microarray, totally 135 arrays were used to characterize light response in different photoreceptor knockout strains. 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.
2009-03-08 | E-GEOD-8932 | biostudies-arrayexpress
Project description:RNA-seq from Neurospora crassa at 5 time points of light induction, with 2 replicates for each, totalling 10 samples
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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.
2011-10-12 | GSE32871 | GEO
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.
2011-10-11 | E-GEOD-32871 | biostudies-arrayexpress
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
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Project description:In chromatin, nucleosomes are composed of ∼146 bp of DNA wrapped around a histone octamer, and are highly dynamic structures subject to remodeling and exchange. Histone turnover has previously been implicated in various processes including the regulation of chromatin accessibility, segregation of chromatin domains, and dilution of histone marks. Histones in different chromatin environments may turnover at different rates, possibly with functional consequences. Neurospora crassa sports a chromatin environment that is more similar to that of higher eukaryotes than yeasts, which have been utilized in the past to explore histone exchange. We constructed a simple light-inducible system to profile histone exchange in N. crassa on a 3xFLAG-tagged histone H3 under the control of the rapidly inducible vvd promoter. After induction with blue light, incorporation of tagged H3 into chromatin occurred within 20 min. Previous studies of histone turnover involved considerably longer incubation periods and relied on a potentially disruptive change of medium for induction. We used this reporter to explore replication-independent histone turnover at genes and examine changes in histone turnover at heterochromatin domains in different heterochromatin mutant strains. In euchromatin, H3-3xFLAG patterns were almost indistinguishable from that observed in wild-type in all mutant backgrounds tested, suggesting that loss of heterochromatin machinery has little effect on histone turnover in euchromatin. However, turnover at heterochromatin domains increased with loss of trimethylation of lysine 9 of histone H3 or HP1, but did not depend on DNA methylation. Our reporter strain provides a simple yet powerful tool to assess histone exchange across multiple chromatin contexts.
| S-EPMC7337079 | biostudies-literature
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.
2010-06-05 | E-GEOD-14909 | biostudies-arrayexpress