Identification of Potential Regulatory Non-Coding RNAs in Lotus Japonicus Symbiosis

Symbiosis between legumes and rhizobia is beneficial on nutrient-poor soils, as it enables the fixation of atmospheric N ₂ . To establish this symbiosis, gene expression in both the host plant and the symbiont has to be regulated. To understand the underlying RNA-mediated regulation of host gene expression, we designed experiments to identify competing endogenous networks involving circular RNA, microRNA, and linear transcripts during symbiosis, using wt and symbiosis-deficient Lotus japonicus mutants with the rhizobium Mesorhizobium loti (M. loti ).

CircRNA, miRNA, and linear transcripts were identified from Lotus japonicus wildtype and CCamK mutant ( ccamk-13; snf-1 ) seedlings without inoculation or with M. loti inoculation using deep short-read sequencing with rRNA-depletion and random primers.

Differentially expressed miRNAs showed negative correlations to predicted target genes and may regulate symbiotic processes. The symbiosis essential iron-sensor LjnsRING/BRUTUS expresses a circRNA which was upregulated in symbiotic treatments. This circRNA may act as a target mimic and contribute to nodule longevity. CircRNAs are predicted to act predominantly as trans- regulatory molecules with similar frequencies in Arabidopsis thaliania, Oryza sativa, and Lotus japonicus .

We identified novel miRNAs, long noncoding RNAs, and circRNAs, and nominated several as potential new regulatory non-coding RNAs that may act as target mimics to stabilize genes and support symbiosis.