Regulation of Copper Homeostasis and Biotic Interactions by MicroRNA 398b in Common Bean

Abstract

MicroRNAs are recognized as important post-transcriptional regulators in plants. Information about the roles of miRNAs in common bean (Phaseolus vulgaris L.), an agronomically important legume, is yet scant. The objective of this work was to functionally characterize the conserved miRNA: miR398b and its target Cu/Zn Superoxide Dismutase 1 (CSD1) in common bean. We experimentally validated a novel miR398 target: the stress up-regulated Nodulin 19 (Nod19). Expression analysis of miR398b and target genes –CSD1 and Nod19- in bean roots, nodules and leaves, indicated their role in copper (Cu) homeostasis. In bean plants under Cu toxicity miR398b was decreased and Nod19 and CSD1, that participates in reactive oxygen species (ROS) detoxification, were up-regulated. The opposite regulation was observed in Cu deficient bean plants; lower levels of CSD1 would allow Cu delivery to essential Cu-containing proteins. Composite common bean plants with transgenic roots over-expressing miR398 showed ca. 20-fold higher mature miR398b and almost negligible target transcript levels as well as increased anthocyanin content and expression of Cu-stress responsive genes, when subjected to Cu deficiency. The down-regulation of miR398b with the consequent up-regulation of its targets was observed in common bean roots during the oxidative burst resulting from short-time exposure to high Cu. A similar response occurred at early stage of bean roots inoculated with Rhizobium tropici, where an increase in ROS was observed. In addition, the miR398b down-regulation and an increase in CSD1 and Nod19 were observed in bean leaves challenged with Sclerotinia scleortiorum fungal pathogen. Transient over-expression of miR398b in Nicotiana benthamiana leaves infected with S. sclerotiorum resulted in enhanced fungal lesions. We conclude that the miR398b-mediated up-regulation of CSD and Nod19 is relevant for common bean plants to cope with oxidative stress generated in abiotic and biotic stresses.

The Micro-RNA172c-APETALA2-1 Node as a Key Regulator of the Common Bean-Rhizobium etli Nitrogen Fixation Symbiosis

“…RNA preparations were used to detect mature miR172 in different plant tissues by low-molecular-weight RNA-gel hybridization using 15 mg of total RNA, as reported (Naya et al, 2014). Synthetic DNA oligonucleotides with antisense sequence corresponding to miR172 (59-ATGCAGCATCATCAA-GATTCT-39) and to U6 snRNA (59-CCAATTTTATCGGATGTCCCCG-39) were used as probes after radioactive labeling.…”

“…Common bean miRNAs that respond to drought, salinity, nutrient deficiencies, or metal toxicity stresses have been identified, and their target genes have been predicted or validated (Arenas-Huertero et al, 2009;Valdés-López et al, 2010;Contreras-Cubas et al, 2012). The roles of miR399 in the common bean root response to phosphorus deficiency (Valdés-López et al, 2008) and of miR398 in the regulation of copper homeostasis and response to biotic interactions (Naya et al, 2014) have been demonstrated. In this work, we analyzed the role of miR172 in common bean roots and nodules.…”

The advance of tomato disease-related microRNAs

“…In a recent research, it was observed that the expression level of miR398 decreased gradually in tomato leaves, after Phytophthora infestans (P. infestans) infection . Under pathogen attack, miR398 were down-regulated in several other plant species, while CSD1 and CSD2 were up-regulated, further detoxifying ROS (Jagadeeswaran et al 2009;Lu et al 2011;Naya et al 2014). Indeed, functional analyses have demonstrated that Nicotiana benthamiana (N. benthamiana) leaves transiently overexpressing miR398b were more susceptible to Sclerotinia sclerotiorum infection (Naya et al 2014).…”

Identification and characterization of microRNAs and their targets in high-altitude stress-adaptive plant maca (Lepidium meyenii Walp)

“…Post-transcriptional gene regulation mediated by endogenous miRNAs play a crucial role in various aspects of plant development as well as adaptation to biotic and abiotic stresses (Naya et al 2014;Khraiwesh et al 2012;Paul et al 2011;Kundu et al 2017). In plants, mature miRNAs are generated from longer stem-loop RNA precursors (Pre-miRNA) with the aid of ribonuclease III-like Dicer (DCL1) enzyme (Fukudome and Fukuhara 2017).…”