**6. miRNA in symbiosis: regulation through repression**

Emphasis on existence and importance of small ncRNA, especially miRNA started with the discovery of its association with regulation of gene expression in Petunia [101, 102]. Owing genome wide studies and high-throughput sequencing efforts, till date thousands of miRNAs have been characterized throughout the kingdom of life. Most of the characterized miRNA associated with symbiosis are either involved in nutrient signaling, exchange and homeostasis or development of nodule/arbuscules or both [103, 104] are illustrated in **Figure 1**. Based on morphological analysis of host, AM Symbiosis (AMS) can be sub-categorized into four major stages: (1) pre-contact signaling, (2) contact establishment between plant root and fungal hyphae, 3) intra-radical proliferation, and 4) arbuscule formation [105]. A non-canonical form of miRNA171, which is found repressed under phosphate starvation, regulates an important transcription factor (TF) involved in common symbiotic signaling pathway, NSP1/2 (NODULE SIGNALING PATHWAY 1/2) during mycorrhizal symbiosis.

#### **Figure 1.**

*Illustration of cross-talk between myc-LCO (mycorrhizal lipo-chito-oligosaccharide) factor induced common pathway signaling, miRNAs and regulatory hormones during AM symbiosis. Solid and dotted lines represents direct and indirect interconnections respectively.*

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

The interaction of NSP2 with the mycorrhizal-specific GRAS TF, RAM1 (Required for Arbuscular Mycorrhization 1) regulates RAM2, and DWARF27 that are part of cutin and strigo-lactone biosynthetic pathway, respectively, as these two pathways facilitate AM inoculation. miRNA393, also identified as a N- and Pi-responsive miRNA, is involved in the homeostasis of auxin signaling and thus inhibits the auxin receptor TRANSPORT INHIBITOR RESPONSE1/AUXIN SIGNALING F-BOX PROTEIN (TIR1/ABF), also mediates the repression of root growth regulatory factors (GRFs) to affect fungal colonization and arbuscule development. miRNA396b also investigated to perform a significant role in root colonization and development during mycorrhizal symbiosis by targeting six GRFs and a TF in *Medicago truncaluta* [106].

For initiating rhizobia symbiosis, interactions between miRNAs, Nod factor signaling, and hormone regulation through NSP2 is controlled by a nodule-specific miRNA171. The NUCLEAR FACTOR (NF)-YA gene, a TF necessary for the nodule initiation and maintenance of meristem, is negatively regulated by miRNA169 [107]. The combination of cytokinin-responsive miRNA172 genes, Nod factor, and various

#### **Figure 2.**

*Illustration of cross-talk between Nod factor induced common pathway signaling, miRNAs and regulatory hormones during rhizobial symbiosis. Solid and dotted lines represents direct and indirect interconnections respectively.*

AP2 (APETALA2) targets has been implicated in rhizobia infection, stimulation of nodule organogenesis, N2 fixation, and delaying senescence in nodule cells.

The auto-regulation of the nodulation (AON) pathway, which governs the number of nodules that are formed in host plants, is one mechanism by which miRNA172 can work. In this pathway, leucine-rich repeat receptor-like kinase (LRR-LRK) receptors recognize NF- or nitrate-induced Clavata3/Embryo Surrounding Region-Relatedpeptides, resulting in an inhibitory signal (including CK) to cells, for establishing new nodules. By suppressing certain squamosa promoter-binding protein-like (SPL) TFs that stimulate miRNA172 production, miRNA156 antagonizes the action of miRNA172 [108].

Through the repression of nodulin gene (specifically ENOD93), miRNA393j-3p restricts nodules [109] whereas, miRNA1512 and miRNA1515 over-expression was discovered to be linked to increased nodule formation [75]. Finally, miRNA160 and miRNA167 cleave the transcripts of multiple auxin response factors (ARFs) that play key roles in the auxin response and pre-requisite for nodule initiation [110]. The detailed regulation mechanism is shown in **Figure 2**. miRNA390encourages the formation of a transacting small interfering (tasi)RNA, that represses ARF3 and ARF4during rhizobia colonization and nodule growth [78], it is known to combine auxin and ethylene signals. While, ARF10, ARF16, ARF17 and ARF6, ARF8 are directly regulated/targeted by miRNA160 and miRNA167, respectively [111] for auxin-responsive root development in both cases.
