PT  - JOURNAL ARTICLE
AU  - Stefan Kusch
AU  - Mansi Singh
AU  - Hannah Thieron
AU  - Pietro D. Spanu
AU  - Ralph Panstruga
TI  - Site-specific analysis reveals candidate cross-kingdom small RNAs, tRNA and rRNA fragments, and signs of fungal RNA phasing in the barley-powdery mildew interaction
AID  - 10.1101/2022.07.26.501657
DP  - 2022 Jan 01
TA  - bioRxiv
PG  - 2022.07.26.501657
4099  - http://biorxiv.org/content/early/2022/08/11/2022.07.26.501657.short
4100  - http://biorxiv.org/content/early/2022/08/11/2022.07.26.501657.full
AB  - The establishment of host-microbe interactions requires molecular communication between both partners, which involves the mutual transfer of noncoding small RNAs. Previous evidence suggests that this is also true for the barley powdery mildew disease, which is caused by the fungal pathogen Blumeria hordei. However, previous studies lacked spatial resolution regarding the accumulation of small RNAs upon host infection by B. hordei. Here, we analysed site-specific small RNA repertoires in the context of the barley-B. hordei interaction. To this end, we dissected infected leaves into separate fractions representing different sites that are key to the pathogenic process: epiphytic fungal mycelium, infected plant epidermis, isolated haustoria, a vesicle-enriched fraction from infected epidermis, and extracellular vesicles. Unexpectedly, we discovered enrichment of specific 31- to 33-base long 5’-terminal fragments of barley 5.8S ribosomal RNA (rRNA) in extracellular vesicles and infected epidermis, as well as particular B. hordei tRNA fragments in haustoria. We describe canonical small RNAs from both the plant host and the fungal pathogen that may confer cross-kingdom RNA interference activity. Interestingly, we found first evidence of phased small RNAs (phasiRNAs) in B. hordei, a feature usually attributed to plants, which may be associated with the post-transcriptional control of fungal coding genes, pseudogenes, and transposable elements. Our data suggests a key and possibly site-specific role for cross-kingdom RNA interference and noncoding RNA fragments in the host-pathogen communication between B. hordei and its host barley.Competing Interest StatementThe authors have declared no competing interest.AgoArgonauteDclDicer-likeEPIepidermis (colonized by B. hordei but epiphytic fungal mycelium removed)dsRNAdouble-stranded RNAEVextracellular vesicleFDRfalse discovery rateGOgene ontologyHAUhaustoria of B. hordeimilRNAmicro RNA-likemiRNAmicro RNAMYCmycelium of B. hordeiphasiRNAphased siRNAP40microsomes from B. hordei-infected epidermisRNAiRNA interferencerDNAribosomal DNArRFribosomal RNA-derived small RNA fragmentrRNAribosomal RNAsRNAsmall RNAsiRNAsmall interfering RNAtasiRNAtrans-acting short interfering RNATPMtranscripts per milliontRFtransfer RNA-derived small RNA fragmenttRNAtransfer RNA