The microRNA476a-RFL module regulates adventitious root formation through a mitochondria-dependent pathway in Populus.

For woody plants, clonal propagation efficiency is largely determined by adventitious root (AR) formation at the bases of stem cuttings. However, our understanding of the molecular mechanisms contributing to AR morphogenesis in trees remains limited, despite the importance of vegetative propagation, currently the most common practice for tree breeding and commercialization. Here, we identified Populus-specific miR476a as a regulator of wound-induced adventitious rooting that acts by orchestrating mitochondrial homeostasis. MiR476a exhibited inducible expression during AR formation and directly targeted several Restorer of Fertility like (RFL) genes encoding mitochondrion-localized pentatricopeptide repeat proteins. Genetic modification of miR476a-RFL expression revealed that miR476a/RFL-mediated dynamic regulation of mitochondrial homeostasis influences AR formation in poplar. Mitochondrial perturbation via exogenous application of a chemical inhibitor indicated that miR476a/RFL-directed AR formation depends on mitochondrial regulation that acts via auxin signaling. Our results thus establish a microRNA-directed mitochondrion-auxin signaling cascade required for AR development, providing insights into the role of mitochondrial regulation in the developmental plasticity of plants.

Identification of microRNAs involved in ectomycorrhizal formation in Populus tomentosa.

The majority of woody plants are able to form ectomycorrhizal (ECM) symbioses with fungi. During symbiotic development, plants undergo a complex re-programming process involving a series of physiological and morphological changes. MicroRNAs (miRNAs) are important components of the regulatory network underlying symbiotic development. To elucidate the mechanisms of miRNAs and miRNA-mediated mRNA cleavage during symbiotic development, we conducted high-throughput sequencing of small RNAs and degradome tags from roots of Populus tomentosa inoculated with Cenococcum geophilum. This process led to the annotation of 51 differentially expressed miRNAs between non-mycorrhizal and mycorrhizal roots of P. tomentosa, including 13 novel miRNAs. Increased or decreased accumulation of several novel and conserved miRNAs in ECM roots, including miR162, miR164, miR319, miR396, miR397, miR398, novel-miR44 and novel-miR47, suggests essential roles for these miRNAs in ECM formation. The degradome analysis identified root transcripts as miRNA-mediated mRNA cleavage targets, which was confirmed using real-time quantitative PCR. Several of the identified miRNAs and corresponding targets are involved in arbuscular mycorrhizal symbioses. In summary, increased or decreased accumulation of specific miRNAs and miRNA-mediated cleavage of symbiosis-related genes indicate that miRNAs play important roles in the regulatory network underlying symbiotic development.