Abstract
Nitrogen (N) is essential for sugar beet (Beta vulgaris L.), a highly N-demanding sugar crop. This study investigated the morphological, subcellular, and microRNA-regulated responses of sugar beet roots to low N (LN) stress (0.5 mmol/L N) to better understand the N perception, uptake, and utilization in this species. The results showed that LN led to decreased dry weight of roots, N accumulation, and N dry matter production efficiency, along with damage to cell walls and membranes and a reduction in organelle numbers (particularly mitochondria). Meanwhile, there was an increase in root length (7.2%) and branch numbers (29.2%) and a decrease in root surface area (6.14%) and root volume (6.23%) in sugar beet after 7 d of LN exposure compared to the control (5 mmol/L N). Transcriptomics analysis was confirmed by qRT-PCR for 6 randomly selected microRNAs, and we identified 22 differentially expressed microRNAs (DEMs) in beet root under LN treatment. They were primarily enriched in functions related to binding (1125), ion binding (641), intracellular (437) and intracellular parts (428), and organelles (350) and associated with starch and sucrose metabolism, tyrosine metabolism, pyrimidine metabolism, amino sugar and nucleotide sugar metabolism, and isoquinoline alkaloid biosynthesis, as indicated by the GO and KEGG analyses. Among them, the upregulated miR156a, with conserved sequences, was identified as a key DEM that potentially targets and regulates squamosa promoter-binding-like proteins (SPLs, 104889216 and 104897537) through the microRNA-mRNA network. Overexpression of miR156a (MIR) promoted root growth in transgenic Arabidopsis, increasing the length, surface area, and volume. In contrast, silencing miR156a (STTM) had the opposite effect. Notably, the fresh root weight decreased by 45.6% in STTM lines, while it increased by 27.4% in MIR lines, compared to the wild type (WT). It can be inferred that microRNAs, especially miR156, play crucial roles in sugar beet root's development and acclimation to LN conditions. They likely facilitate active responses to N deficiency through network regulation, enabling beet roots to take up nutrients from the environment and sustain their vital life processes.