**7. Mycorrhiza-derived non-coding RNAs and cross-kingdom signaling during symbiosis**

Recent investigations have established non-coding RNAs as one of the central mediators of cross-kingdom communication between the plant and microbes [86, 88, 112–119]. These non-coding RNAs can move from donor organism to recipient organism, and target the specific host mRNAs for degradation. Sometimes sRNAs also trigger the production of secondary sRNA and thereby modulate the host defense and metabolic pathways [87, 120, 121]. Most of the studies focus on the plant-parasite or plant-pathogen (fungi and oomycetes) interactions [86, 88, 112, 120], however, such processes have been rarely explored in case of plant-mycorrhiza associations. Emerging body of evidences suggest that many plant miRNAs show differential expression patterns during AM symbiosis, nevertheless, their functions and crosskingdom mobility remains unclear [80, 83, 111, 122, 123]. Mewalal et al. [124] identified several sRNAs from *Polpulus* spp. which were responsive to mutualistic/symbiotic interaction with mycorrhizal fungi like *Laccaria bicolor* and *Rhyzophagus irregularis*. Interestingly, they did not find any Populus RNAs interacting with *R. irregularis*, however, some of the miRNAs could interact with *L. bicolor*. Further the study revealed that these miRNAs can potentially target multiple host mRNAs encoding for vesicular transport and transcription regulatory proteins along with several uncharacterized proteins.

On the other hand, at present, very little information is available regarding the non-coding RNA biogenesis machinery and their functions in the development of arbuscular mycorrhizal (AM) and ectomycorrhizal (ECM) fungi or while interacting with the host plants. However, the successful application of host-induced gene silencing (HIGS) and virus-induced gene silencing (VIGS) approaches [65, 125–128] indicates that AMF, like pathogenic fungi, also possess functional RNAi machinery.

#### *Role of Non-Coding RNAs in Plant Nutrition through Mycorrhizal Interactions DOI: http://dx.doi.org/10.5772/intechopen.108517*

An *in silico* study identified putative RNAi machinery including a Dicer-like (DCL) gene, Argonaute-like (AGO-like) and RNA-dependent RNA polymerase (RdRp) gene families in *R. irregularis*, and validated their transcript-level expression [129]. An unsual expansion of AGO-like (5 members) and RdRp (21 members) gene families was observed in *R. irregularis.* Authors postulated that 15 out of 21RdRp genes, could be the product of a recent gene expansion event. The study also characterized the fungal sRNA and microRNA-like sequences, and predicted 237 transcripts of *Medicago truncatula* as their potential targets including a few known mRNAs that are modulated during AMF colonization. For instance, some of the *M. truncatula* mRNAs that are potentially targeted by *Rir*-sRNAs encode for the nuclear-binding leucine-rich repeat (*NBS-LRR*) type disease resistance gene, Non-specific phospholipase C4 (*NPC4*), *MtVapyrin* (Ankyrin repeat RF-like protein) and DREPP plasma membrane protein (*MtDREPP*) [129]. The homologs of NBS-LRR and NCP4 proteins from rice and arabidopsis, respectively, are involved in the plant immunity [130, 131], thus repression of these genes may allow AM colonization without triggering the robust host defense responses. MtVapyrin plays crucial role in arbuscule formation [132–134]. The down-regulation of *MtDREPP* has been reported in mycorrizal roots [135]. Though, further experimental validation is required, these findings indicate the possible existence of non-coding RNA-mediated post-transcriptional regulation and cross-kingdom gene silencing by AMF.

Another study by Silvestri et al. [136] identified the small RNA population from AMF *Gigaspora margarita* and showed their origin from different genetic sources such as endobacteria, RNA viruses and non-integrated DNA sequences from mitoviruses. Intriguingly, the extracellular vesicles (EVs), that are deployed in delivering the sRNA molecules to the other interacting partner [112, 120, 137], have also been observed in the peri-arbuscular interface of *R. irregularis* during the whole lifespan of arbuscules. This indicates the crucial role of EVs in cross-kingdom communication and nutrient exchange during AMF symbiosis [138]. More recently, a breakthrough discovery demonstrated that an ECM fungus *Pisolithus microcarpus* encodes 11 miRNAs, six of them were found induced during host colonization process. Notably, the miRNA (*Pmic\_miR-8*) enters the plant cell and partakes in cross-kingdom gene silencing at some stage in symbiotic interaction with host plant *Eucalyptus grandis* [139]. The inhibition of *Pmic\_miR-8* resulted in less developed Hartig nets, whereas, supplementation showed increased Hartig net depth in host tissue. Further the study showed that *Pmic\_miR-8* may target the host NB-ARC (nucleotide-binding adaptor shared by APAF-1, R proteins, and CED-4) domain containing transcripts, indicating its potential role in modulating host signaling to stabilize the mutualistic association. As the CC (coiled-coil) nucleotide binding and leucine-rich repeat domain immune receptors (CC-NLR) are the largest category of NLRs, thus *Pmic\_miR-8* may target several plant genes belonging to this class. Importantly, this is the first study which established the cross-kingdom gene silencing by mycorrhizal fungi and its role in beneficial interactions with host.

### **8. Conclusion and future prospects**

In the last decade, the non-coding RNAs have emerged as one of the key regulators of diverse plant process including their development, response to abiotic/biotic stress, and nutrient uptake. A significant advancement has been made to understand the crucial roles of non-coding RNAs in plant-microbe interactions, particularly

pathogenic interactions. Nutrient uptake via mycorrizal association is an important aspect of plants lifestyle and the studies suggest the extensive involvement of noncoding RNAs in regulating the plant nutrient status via affecting symbiosis. Notably, the sRNA-mediated regulatory mechanisms during mycorrhizal symbiosis have mainly focused on plant's perspective. These studies have provided crucial insights on understanding how the mycorrizal colonization proceeds and how the host plants fine-tune the extent of fungal colonization so that it does not turns pathogenic. On the contrary, the sRNA-mediated regulation of symbiosis from the fungal perspective remains infancy. Moreover, an integrated view of both the organisms (plant and fungus) will be required to appropriately comprehend the beneficial relationships. To understand ncRNA-molecular interaction networks occurring at plant host-AM symbiosis interface, experimental evidences and rewriting of dynamics of interaction, sensing, uptake, transport, assimilation and homeostasis of nutrients regulation are required. Extensive investigations for ncRNAs mediated regulation of crosstalk between AM and host plant are also needed for teeming knowledge voids. We anticipate that the recent discovery of cross-kingdom gene silencing by ECM fungus *Pisolithus microcarpus* and AM-like model EM fungus *Serendipita indica* would pave the way for future investigations of non-coding RNA-mediated regulatory networks in mycorriza growth and development as well as during host interactions, and would trigger novel research ideas among plant scientists. The better understanding of these regulatory circuits would aid in improving the nutritional status of plants in order to combat the elevating global quality-food demand.
