Physiological responses and small RNAs changes in maize under nitrogen deficiency and resupply

Background Maize is an important crop in the world, nitrogen stress severely reduces maize yield. Although a large number of studies have identified the expression changes of microRNAs (miRNAs) under N stress in several species, the miRNAs expression patterns of N-deficient plants under N resupply remain unclear. Objective The primary objective of this study was to identify miRNAs in response to nitrogen stress and understand relevant physiological changes in nitrogen-deficient maize after nitrogen resupply. Methods Physiological parameters were measured to study relevant physiological changes under different nitrogen conditions. Small RNA sequencing and qRT-PCR analysis were performed to understand the response of miRNAs under different nitrogen conditions. Results The content of chlorophyll, soluble protein and nitrate nitrogen decreased than CK by 0.52, 0.49 and 0.82 times after N deficiency treatment and increased than ND by 0.52, 1.36 and 0.65 times after N resupply, respectively. Conversely, the activity of superoxide dismutase (SOD) and peroxidase (POD) increased by 0.67 and 1.64 times than CK after N deficiency, respectively, and decreased by 0.09 and 0.35 times than ND after N resupply. A total of 226 known miRNAs were identified by sRNA sequencing; 106 miRNAs were differentially expressed between the control and N-deficient groups, and 103 were differentially expressed between the N-deficient and N-resupply groups (P < 0.05). Real-time quantitative PCR (qPCR) was used to further validate and analyze the expression of the identified miRNAs. A total of 1609 target genes were identified by target prediction, and some differentially expressed miRNAs were predicted to target transcription factors and functional proteins. Gene Ontology (GO) analysis was used to determine the biological function of these targets and revealed that some miRNAs, such as miR169, miR1214, miR2199, miR398, miR408 and miR827 might be involved in nitrogen metabolism regulation. Conclusion Our study comprehensively provides important information on miRNA functions and molecular mechanisms in response to N stress. These findings may assist to improve nitrogen availability in plants.