Background Small RNAs (sRNAs) are short non-coding RNA molecules (20–30 nt) that regulate gene expression at transcriptional or post-transcriptional levels in many eukaryotic organisms, through a mechanism known as RNA interference (RNAi). Recent studies have highlighted that they are also involved in cross-kingdom communication: sRNAs can move across the contact surfaces from “donor” to “receiver” organisms and, once in the host cells of the receiver, they can target specific mRNAs, leading to a modulation of host metabolic pathways and defense responses. Very little is known about RNAi mechanism and sRNAs occurrence in Arbuscular Mycorrhizal Fungi (AMF), an important component of the plant root microbiota that provide several benefits to host plants, such as improved mineral uptake and tolerance to biotic and abiotic stress. Results Taking advantage of the available genomic resources for the AMF Rhizophagus irregularis we described its putative RNAi machinery, which is characterized by a single Dicer -like ( DCL ) gene and an unusual expansion of Argonaute -like ( AGO -like) and RNA-dependent RNA polymerase ( RdRp ) gene families. In silico investigations of previously published transcriptomic data and experimental assays carried out in this work provided evidence of gene expression for most of the identified sequences. Focusing on the symbiosis between R. irregularis and the model plant Medicago truncatula , we characterized the fungal sRNA population, highlighting the occurrence of an active sRNA-generating pathway and the presence of microRNA-like sequences. In silico analyses, supported by host plant degradome data, revealed that several fungal sRNAs have the potential to target M. truncatula transcripts, including some specific mRNA already shown to be modulated in roots upon AMF colonization. Conclusions The identification of RNAi-related genes, together with the characterization of the sRNAs population, suggest that R. irregularis is equipped with a functional sRNA-generating pathway. Moreover, the in silico analysis predicted 237 plant transcripts as putative targets of specific fungal sRNAs suggesting that cross-kingdom post-transcriptional gene silencing may occur during AMF colonization. Electronic supplementary material The online version of this article (10.1186/s12864-019-5561-0) contains supplementary material, which is available to authorized users.
We attempted to transfect six recently characterized virus species to protoplasts of Penicillium janczewskii and Chryphonectria parasitica. None of the recovered P. janczewskii colonies was positive for the transfected viruses, but Penicillium aurantiogriseum partiti-like virus 1 (PaPLV1) was detected in three distinct regenerated C. parasitica colonies. We screened the phenotype of the infected strains in up to 45 different conditions combining different media, salinity and temperatures: our results show that the infected strains grow slower than the virus- free in most of the tested conditions with the exception of halophilic stress in a specific nutrient combination media. We proceeded to characterize molecularly the population of distinct isolates of PaPLV1 infected C. parasitica through RNAseq: comparison to the viral population present in the original host - P. auratiogriseum - showed that two isolates accumulated non-synonymous mutations suggesting adaptation to the new host. RNAseq analyses identified a second genomic RNA segment and northern blot of RNA extracted from purified virus suspensions allowed establishing that PaPLV1 is at least bipartite in nature and that it forms isometric virions of circa 36-38 nm in diameter. In light of these new acquisitions, we discuss the taxonomic placement of PaPLV1 inside the Partitiviridae.
Arbuscular Mycorrhizal Fungi (AMF) are key components of the plant microbiota. AMF genetic complexity is increased by the presence of endobacteria, which live inside many species. A further component of such complexity is the virome associated to AMF, whose knowledge is still very limited. Here, by exploiting transcriptomic data we describe the virome of Gigaspora margarita. A BLAST search for viral RNA-dependent RNA polymerases sequences allowed the identification of four mitoviruses, one Ourmia-like narnavirus, one Giardia-like virus, and two sequences related to Fusarium graminearum mycoviruses. Northern blot and RT-PCR confirmed the authenticity of all the sequences with the exception of the F. graminearum-related ones. All the mitoviruses are replicative and functional since both positive strand and negative strand RNA are present. The abundance of the viral RNA molecules is not regulated by the presence or absence of Candidatus Glomeribacter gigasporarum, the endobacterium hosted by G. margarita, with the exception of the Ourmia-like sequence which is absent in bacteria-cured spores. In addition, we report, for the first time, DNA fragments corresponding to mitovirus sequences associated to the presence of viral RNA. These sequences are not integrated in the mitochondrial DNA and preliminary evidence seems to exclude integration in the nuclear genome.
RNA interference (RNAi) is a key regulatory pathway of gene expression in almost all eukaryotes. This mechanism relies on short non-coding RNA molecules (sRNAs) to recognize in a sequence-specific manner DNA or RNA targets leading to transcriptional or post-transcriptional gene silencing. To date, the fundamental role of sRNAs in the regulation of development, stress responses, defense against viruses and mobile elements, and cross-kingdom interactions has been extensively studied in a number of biological systems. However, the knowledge of the "RNAi world" in arbuscular mycorrhizal fungi (AMF) is still limited. AMF are obligate mutualistic endosymbionts of plants, able to provide several benefits to their partners, from improved mineral nutrition to stress tolerance. Here we described the RNAi-related genes of the AMF Gigaspora margarita and characterized, through sRNA sequencing, its complex small RNAome, considering the possible genetic sources and targets of the sRNAs. G. margarita indeed is a mosaic of different genomes since it hosts endobacteria, RNA viruses, and nonintegrated DNA fragments corresponding to mitovirus sequences. Our findings show that G. margarita is equipped with a complete set of RNAi-related genes characterized by the expansion of the Argonaute-like (AGO-like) gene family that seems a common trait of AMF. With regards to sRNAs, we detected populations of sRNA reads mapping to nuclear, mitochondrial, and viral genomes that share similar features (25-nt long and 5 -end uracil read enrichments), and that clearly differ from sRNAs of endobacterial origin. Furthermore, the annotation of nuclear loci producing sRNAs suggests the occurrence of different sRNA-generating processes. In silico analyses indicate that the most abundant G. margarita sRNAs, including those of viral origin, could target transcripts in the host plant, through a hypothetical cross-kingdom RNAi.
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