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.

Restorer of fertility like 30, encoding a mitochondrion-localized pentatricopeptide repeat protein, regulates wood formation in poplar.

Nuclear-mitochondrial communication is crucial for plant growth, particularly in the context of cytoplasmic male sterility (CMS) repair mechanisms linked to mitochondrial genome mutations. The restorer of fertility-like (RFL) genes, known for their role in CMS restoration, remain largely unexplored in plant development. In this study, we focused on the evolutionary relationship of RFL family genes in poplar specifically within the dioecious Salicaceae plants. PtoRFL30 was identified to be preferentially expressed in stem vasculature, suggesting a distinct correlation with vascular cambium development. Transgenic poplar plants overexpressing PtoRFL30 exhibited a profound inhibition of vascular cambial activity and xylem development. Conversely, RNA interference-mediated knockdown of PtoRFL30 led to increased wood formation. Importantly, we revealed that PtoRFL30 plays a crucial role in maintaining mitochondrial functional homeostasis. Treatment with mitochondrial activity inhibitors delayed wood development in PtoRFL30-RNAi transgenic plants. Further investigations unveiled significant variations in auxin accumulation levels within vascular tissues of PtoRFL30-transgenic plants. Wood development anomalies resulting from PtoRFL30 overexpression and knockdown were rectified by NAA and NPA treatments, respectively. Our findings underscore the essential role of the PtoRFL30-mediated mitochondrion-auxin signaling module in wood formation, shedding light on the intricate nucleus-organelle communication during secondary vascular development.

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.