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Diverse pathways generate microRNA-like RNAs and Dicer-independent small interfering RNAs in fungi

ABSTRACT: A variety of small RNAs, including the Dicer-dependent miRNAs and the Dicer-independent Piwi-interacting RNAs, associate with Argonaute family proteins to regulate gene expression in diverse cellular processes. These two species of small RNA have not been found in fungi. Here, by analyzing small RNA associated with the Neurospora Argonaute protein QDE-2, we show that diverse pathways generate miRNA-like small RNAs (milRNAs) and Dicer-independent small interfering RNAs (disiRNAs) in this filamentous fungus. Surprisingly, milRNAs are produced by at least four different mechanisms that use a distinct combination of factors, including Dicers, QDE-2, the exonuclease QIP and an RNAse III domain-containing protein MRPL3. In contrast, disiRNAs originate from loci producing overlapping sense and antisense transcripts, and do not require the known RNAi components for their production. Taken together, these results uncover several pathways for small RNA production in filamentous fungi, shedding light on the diversity and evolutionary origins of eukaryotic small RNAs.

ORGANISM(S): Neurospora crassa

PROVIDER: GSE21175 | GEO | 2010/07/01

SECONDARY ACCESSION(S): PRJNA125805

REPOSITORIES: GEO

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Diverse pathways generate microRNA-like RNAs and Dicer-independent small interfering RNAs in fungi

Project description:A variety of small RNAs, including the Dicer-dependent miRNAs and the Dicer-independent Piwi-interacting RNAs, associate with Argonaute family proteins to regulate gene expression in diverse cellular processes. These two species of small RNA have not been found in fungi. Here, by analyzing small RNA associated with the Neurospora Argonaute protein QDE-2, we show that diverse pathways generate miRNA-like small RNAs (milRNAs) and Dicer-independent small interfering RNAs (disiRNAs) in this filamentous fungus. Surprisingly, milRNAs are produced by at least four different mechanisms that use a distinct combination of factors, including Dicers, QDE-2, the exonuclease QIP and an RNAse III domain-containing protein MRPL3. In contrast, disiRNAs originate from loci producing overlapping sense and antisense transcripts, and do not require the known RNAi components for their production. Taken together, these results uncover several pathways for small RNA production in filamentous fungi, shedding light on the diversity and evolutionary origins of eukaryotic small RNAs. One small RNA library was generated using QDE-2 immunoprecipitate from Neurospora crassa.

2010-07-01 | E-GEOD-21175 | biostudies-arrayexpress

Identification of microRNA-like RNAs in the filamentous fungus Trichoderma reesei by Solexa sequencing

Project description:MicroRNAs (miRNAs) are small non-coding RNAs capable of negatively regulating gene expression. Trichoderma reesei is an industrial filamentous fungus that can secrete abundant hydrolases for cellulosic biofuels. Recently, microRNA-like RNAs (milRNAs) were discovered in several filamentous fungi rather than T. reesei. The purpose of this study was to explore the presence of milRNA in T. reesei, to characterize the differential expression of T. reesei milRNA under cellulose induction, and to reveal the target genes of milRNA involved in cellulase production. Two small RNA libraries of cellulose induction (IN) or non-induction (CON) were generated and sequenced using Solexa sequencing technology. A total of 664,463 and 529,545 unique sequences, representing 1,271 and 1,021 unique small RNAs, were obtained from the IN and CON samples, respectively. Thirteen milRNAs were finally identified in T. reesei using the hairpin structure analysis. The milRNAs profiles obtained in deep sequencing were validated by RT-qPCR assay. The miRanda program predicts a number of potential targets for T. reesei milRNAs, including several hydrolases and carbon catabolite repressor Cre1.The presence and differential expression of T. reesei milRNAs, along with their predicted targets indicate that milRNAs might play a regulatory role in cellulase induction. This work lays foundation for further functional study of fungal milRNAs and their industrial application.

2014-12-31 | GSE46679 | GEO

Identification of miRNA-like RNAs in a plant pathogen fungus Sclerotinia sclerotiorum by High-throughput sequencing

Project description:This study provides a first large-scale cloning and characterization of Sclerotinia sclerotiorum milRNAs and milRNAs candidates. Two microRNA-like RNAs (milRNAs) and 42 milRNA candidates were identified by sequence analysis. These milRNAs and candidates provide new insights into the functional roles of small RNAs and adds new resources for the study of plant pathogenic fungi. We constructed a small RNA library from Sclerotinia sclerotiorum.

2012-05-17 | E-GEOD-28730 | biostudies-arrayexpress

Identification of miRNA-like RNAs in a plant pathogen fungus Sclerotinia sclerotiorum by High-throughput sequencing

2012-05-18 | GSE28730 | GEO

Small RNAs generated in vitro by budding-yeast Dicer

Project description:The Dicer ribonuclease III (RNase III) enzymes process long double-stranded RNA (dsRNA) into the small interfering RNAs (siRNAs) that direct RNA interference. Here, we describe the structure and activity of a catalytically active fragment of Kluyveromyces polysporus Dcr1, which represents the noncanonical Dicers found in budding yeast. The crystal structure reveals a homodimer that resembles bacterial RNase III but includes a novel N-terminal domain and newly identified catalytic residues conserved throughout eukaryotic RNase III enzymes. Biochemical analyses show that Dcr1 dimers bind cooperatively along the dsRNA substrate and cleave at precise intervals based on the distance between consecutive active sites. Thus, unlike canonical Dicers, which successively remove siRNA duplexes from the dsRNA termini, Dcr1 initiates processing in the interior and works outward. The distinct mechanism of budding-yeast Dicers establishes a novel paradigm for natural protein-based molecular rulers and imparts substrate preferences with ramifications for biological function. High-throughput sequencing of small RNAs generated by in vitro processing of long dsRNA using Kluyveromyces polysporus Dicer

2011-05-11 | E-GEOD-29168 | biostudies-arrayexpress

Global transcriptional regulation by small non-coding RNAs is required for the response to injury of a filamentous fungus

Project description:The ability to respond to injury is essential for the survival of an organism and involves analogous mechanisms in animals and plants. Such mechanisms integrate coordinated genetic and metabolic reprogramming events requiring fine tuned regulation by small RNAs for adequate healing of the wounded area. We previously reported that in response to injury, the filamentous fungus Trichoderma atroviride uses molecular mechanisms very similar to those of plants and animals. These processes lead to the formation of new hyphae and the development of asexual reproduction structures (conidiophores). Here we show that regulation by sRNAs of nitrogen metabolic processes and the cellular redox balance is necessary to carry out the correct response to injury. We further discover microRNA-like RNAs (milRNAs) related with the development of conidiophores. Finally, using a loss-of-function and transcriptomic approach, we demonstrate that a milRNA plays a central role in growth and hyphal regeneration in the filamentous fungus T. atroviride, which regulates signaling and cell proliferation at the transcriptional level. Although many classes of small RNAs have been profiled in fungi, and showed to be important for contending with different types of stress, this is the first report showing a physiological function of a milRNA in fungi.

2018-12-05 | GSE123330 | GEO

micro-like RNAs and their targets engage in DCL2-deleted Valsa mali

Project description:Dicer proteins are mainly responsible for generating small RNAs (sRNAs). In previous study, two DCL proteins in Valsa mali, the pathogenic fungus of apple tree Valsa canker, were found to be associated with the pathogenicity and generation of sRNAs. In this study, we constructed two milRNA libraries using the the wild-type 03-8 (MVm) and Vm-DCL2 deletion mutant (MD2) strains and the differential expression of miRNA-like RNAs (milRNAs) were analyzed based on the deep-sequencing of the wild-type and Vm-DCL2 mutant, respectively. In total, by comparing the expression difference of milRNAs between MVm and MD2, it was found that 33 milRNAs were not detected in MD2, 28 milRNAs were both detected in MVm and MD2. By statistical analysis, one and 17 milRNAs showed up-regulated and down-regulated expression in MD2 compared with MVm, respectively (p<0.05). Importantly, 10 milRNAs were not detected in MD2, and identified to be DCL2 dependent milRNAs. We also identified the target genes of these milRNAs using degradome sequencing technology and found they were associated with protein phosphatase, isocitrate dehydrogenase, phosphoglycerate mutase, etc. The study will lay a foundation for the comprehensive analysis of pathogenic mechanisms of V. mali, and enrich cognition of the generation and function of fungal sRNA.

2021-08-07 | GSE181528 | GEO

Small RNAs generated in vitro by budding-yeast Dicer

2011-05-11 | GSE29168 | GEO

Small RNAs in budding yeast

Project description:RNAi, a gene-silencing pathway triggered by double-stranded RNA, is conserved in diverse eukaryotic species but has been lost in the model budding yeast, Saccharomyces cerevisiae. We report that RNAi is present in other budding-yeast species, including Saccharomyces castellii and Candida albicans. These species use noncanonical Dicer proteins to generate siRNAs, which mostly correspond to transposable elements and YM-BM-4 subtelomeric repeats. In S. castellii, RNAi mutants are viable but have excess YM-BM-4 mRNA levels. In S. cerevisiae, introducing Dicer and Argonaute of S. castellii restores RNAi, and the reconstituted pathway silences endogenous retrotransposons. These results identify a novel class of Dicer proteins, bring the tool of RNAi to the study of budding yeasts, and bring the tools of budding yeast to the study of RNAi. Employ high-throughput sequencing of endogenous small RNAs from the budding yeasts Saccharomyces castellii, Kluyveromyces polysporus, Candida albicans, Saccharomyces cerevisiae, and Saccharomyces bayanus.

2009-09-10 | E-GEOD-17871 | biostudies-arrayexpress

Tailing and degradation of Argonaute-bound small RNAs protect the genome from uncontrolled RNAi

Project description:RNAi is a conserved mechanism in which small RNAs induce silencing of complementary targets. How Argonaute-bound small RNAs are targeted for degradation is not well understood. We show here that the adenyl-transferase Cid14, a member of the TRAMP complex, and the uridyl-transferase Cid16 add non-templated nucleotides to Argonaute-bound small RNAs in fission yeast. The tailing of Argonaute-bound small RNAs in vitro recruits the 3’-5’ exonuclease Rrp6 to degrade small RNAs. Failure in degradation of Argonaute-bound small RNAs results in accumulation of “noise” small RNAs on Argonaute and targeting of diverse euchromatic genes by RNAi. To protect themselves from uncontrolled RNAi, cid14Δ cells exploit the RNAi machinery and silence genes essential for RNAi itself, which is required for their viability. Our data indicate that surveillance of Argonaute-bound small RNAs by Cid14/Cid16 and the exosome protects the genome from uncontrolled RNAi and reveal a rapid RNAi-based adaptation to stress conditions.

2017-05-09 | GSE95821 | GEO

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