Small RNA profiling for identification of microRNAs involved in regulation of seed development and lipid biosynthesis in yellowhorn

Abstract

Background: Yellowhorn (Xanthoceras sorbifolium), an endemic woody oil-bearing tree, has become economically important and is widely cultivated in northern China for bioactive oil production. However, the regulatory mechanisms of seed development and lipid biosynthesis affecting oil production in yellowhorn are still elusive. MicroRNAs (miRNAs) play crucial roles in diverse aspects of biological and metabolic processes in seeds, especially in seed development and lipid metabolism. It is still unknown how the miRNAs regulate the seed development and lipid biosynthesis in yellowhorn. Results: Here, based on investigations of differences in the seed growth tendency and embryo oil content between high-oil-content and low-oil-content lines, we constructed small RNA libraries from yellowhorn embryos at four seed development stages of the two lines and then profiled small RNA expression using high-throughput sequencing. A total of 249 known miRNAs from 46 families and 88 novel miRNAs were identified. Furthermore, by pairwise comparisons among the four seed development stages in each line, we found that 64 miRNAs (53 known and 11 novel miRNAs) were differentially expressed in the two lines. Across the two lines, 15, 11, 10, and 7 differentially expressed miRNAs were detected at 40, 54, 68, and 81 days after anthesis, respectively. Bioinformatic analysis was used to predict a total of 2654 target genes for 141 differentially expressed miRNAs (120 known and 21 novel miRNAs). Most of these genes were involved in the fatty acid biosynthetic process, regulation of transcription, nucleus, and response to auxin. Using quantitative real-time PCR and an integrated analysis of miRNA and mRNA expression, miRNA-target regulatory modules that may be involved in yellowhorn seed size, weight, and lipid biosynthesis were identified, such as miR172b-ARF2 (auxin response factor 2), miR7760-p3_1-AGL61 (AGAMOUS-LIKE 61), miR319p_1-FAD2–2 (omega-6 fatty acid desaturase 2–2), miR5647-p3_1-DGAT1 (diacylglycerol acyltransferase 1), and miR7760-p5_1-MED15A (Mediator subunit 15a). Conclusions: This study provides new insights into the important regulatory roles of miRNAs in the seed development and lipid biosynthesis in yellowhorn. Our results will be valuable for dissecting the post-transcriptional and transcriptional regulation of seed development and lipid biosynthesis, as well as improving yellowhorn in northern China.