Abstract
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 non-integrated 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.
Highlights
RNA interference (RNAi) or RNA silencing is a conserved eukaryotic pathway involved in the repression of gene expression at transcriptional or post-transcriptional level (Moazed, 2009; Wilson and Doudna, 2013; Ipsaro and Joshua-Tor, 2015)
Recent surveys of two arbuscular mycorrhizal fungi (AMF) genomes allowed a first characterization of the RNAi components (DCL, AGO-like, and RNA-dependent RNA polymerase (RdRp)) in this group of obligate biotrophs (Lee et al, 2018; Silvestri et al, 2019)
These works highlighted that the two analyzed AMF, R. irregularis and R. clarus, are equipped with a RNAi machinery characterized by 26–40 AGO-like, 3–21 RdRp, and 1–2 DCL proteins
Summary
RNA interference (RNAi) or RNA silencing is a conserved eukaryotic pathway involved in the repression of gene expression at transcriptional or post-transcriptional level (Moazed, 2009; Wilson and Doudna, 2013; Ipsaro and Joshua-Tor, 2015). The process is mediated by small RNAs (sRNAs) of about 20–30 nucleotides that direct, by sequence complementarity, the recognition and silencing of the target genetic elements. The RNAi pathway relies on three core enzymes: Dicer-like (DCL), Argonaute-like (AGO-like), and RNA-dependent RNA polymerase (RdRp). DCL are ribonuclease III (RNase III) proteins that cleave double-stranded RNAs (dsRNAs) or single-stranded hairpin RNAs producing sRNAs that are loaded onto AGO-like which, guided by sRNAs, are responsible for the silencing of the specific target sequences. RdRp generally play a dual role, both triggering the RNAi pathway and/or amplifying the silencing signals through the synthesis of dsRNAs from aberrant RNAs
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