Identification of Nutrient-Responsive Arabidopsis and Rapeseed MicroRNAs by Comprehensive Real-Time Polymerase Chain Reaction Profiling and Small RNA Sequencing

• Published in: Plant physiology (Bethesda) Vol. 150; no. 3; pp. 1541 - 1555
• Main Authors: Pant, Bikram Datt, Musialak-Lange, Magdalena, Nuc, Przemyslaw, May, Patrick, Buhtz, Anja, Kehr, Julia, Walther, Dirk, Scheible, Wolf-Rüdiger
• Format: Journal Article
• Language: English
• Published: Rockville, MD American Society of Plant Biologists 01.07.2009
• Subjects: algorithms; Arabidopsis; Arabidopsis - drug effects; Arabidopsis - genetics; Arabidopsis - metabolism; Arabidopsis thaliana; Biological and medical sciences; Brassica napus; Brassica napus - genetics; Brassica napus - metabolism; Canola; chemistry; drug effects; Environmental Stress and Adaptation to Stress; Fundamental and applied biological sciences. Psychology; Gene Expression Profiling; Gene Expression Profiling - methods; Genes; genetics; loci; metabolism; methods; MicroRNA; MicroRNAs; MicroRNAs - genetics; MicroRNAs - physiology; Molecular Sequence Data; nitrogen; Nitrogen - pharmacology; nucleotide sequences; nutrient availability; pharmacology; Phloem; Phloem - genetics; Phloem - metabolism; phosphorus; Phosphorus - pharmacology; physiology; plant adaptation; Plant cells; Plant physiology and development; Plants; Polymerase Chain Reaction; quantitative analysis; rapeseed; reverse transcription; RNA; RNA, Plant; RNA, Plant - chemistry; RNA, Plant - physiology; sap; Seedlings; Sequence Analysis, RNA; Sequencing; Brassica napus var. oleifera; RNA; Real time; Arabidopsis thaliana; Polymerase chain reaction; MicroRNA; Plant physiology; Cruciferae; Rapeseed; Dicotyledones; Angiospermae; Nutrient; Spermatophyta; Experimental plant; Sequencing; Oil plant (vegetal)
• ISSN: 0032-0889; 1532-2548; 1532-2548
• DOI: 10.1104/pp.109.139139

Comprehensive expression profiles of Arabidopsis (Arabidopsis thaliana) MIRNA genes and mature microRNAs (miRs) are currently not available. We established a quantitative real-time polymerase chain reaction platform that allows rapid and sensitive quantification of 177 Arabidopsis primary miR transcripts (pri-miRs). The platform was used to detect phosphorus (P) or nitrogen (N) status-responsive pri-miR species. Several pri-miR169 species as well as pri-miR398a were found to be repressed during N limitation, whereas during P limitation, pri-miR778, pri-miR827, and pri-miR399 species were induced and pri-miR398a was repressed. The corresponding responses of the biologically active, mature miRs were confirmed using specific stem-loop reverse transcription primer quantitative polymerase chain reaction assays and small RNA sequencing. Interestingly, the latter approach also revealed high abundance of some miR star strands. Bioinformatic analysis of small RNA sequences with a modified miRDeep algorithm led to the identification of the novel P limitation-induced miR2111, which is encoded by two loci in the Arabidopsis genome. Furthermore, miR2111, miR169, a miR827-like sequence, and the abundances of several miR star strands were found to be strongly dependent on P or N status in rapeseed (Brassica napus) phloem sap, flagging them as candidate systemic signals. Taken together, these results reveal the existence of complex small RNA-based regulatory networks mediating plant adaptation to mineral nutrient availability.