Expression and tissue-specific localization of nitrate-responsive miRNAs in roots of maize seedlings

Abstract Nitrogen availability seriously affects crops productivity and environment. The knowledge of post-transcriptional regulation of plant response to nutrients is important to improve nitrogen use efficiency of crop. This research was aimed at understanding the role of miRNAs in the molecular control of plant response to nitrate. A maize miRNAs-microarray platform was used to discover previously unknown nitrate-responsive miRNAs. Six mature miRNA were identified and their expression profiles were deeply studied by qPCR and in situ hybridization (ISH). To this aim a novel optimized protocol was set up for the use of DIG labelled ZNA (Zip Nucleic Acid)-modified oligonucleotides as probes for ISH. Significant differences in miRNAs’ transcripts accumulation were evidenced between nitrate-supplied and nitrate-depleted roots. Real time PCR analyses and in situ detection of miRNA, confirmed the arrays data and allowed us to evidence distinct miRNAs spatio-temporal expression patterns in maize roots. Our results suggest that a prolonged nitrate depletion may induce post-transcriptionally the expression of target genes by repressing the transcription of specific miRNAs. In particular, the repression of the transcription of miR528a/b, miR528a*/b*, miR169i/j/k, miR169i*/j*/k*, miR166j/k/n and miR408/b upon nitrate shortage could represent a crucial step integrating nitrate signals into developmental changes in maize roots.