Integrated Analysis of Seed microRNA and mRNA Transcriptome Reveals Important Functional Genes and microRNA-Targets in the Process of Walnut (Juglans regia) Seed Oil Accumulation

Abstract

Walnut (Juglans regia) is known as a promising woody oil crop with abundant polyunsaturated fatty acids in its kernel. However, the regulation mechanism of walnut oil accumulation and fatty acid metabolism is still poorly understood, which restricted the breeding and genetic improvement of high-quality oil-bearing walnuts. To reveal the molecular mechanism of walnut oil accumulation, considering the potential regulation of microRNA (miRNA) in seed development, in this study, the oil content of walnut kernel on the 80th, 100th and 120th day after flowering (DAF) was tested and the corresponding proportions are 11.51%, 40.40% and 53.20%. Between DAF of 80th~120th, the content of stearic acid and oleic acid tended to increase, but the proportion of other fatty acids tended to decrease. Meanwhile, comparative transcriptome and sRNA-seq analysis on three stages (80th, 100th and 120th DAF), found 204 conserved miRNAs and 554 novel miRNAs in walnut kernels, among which 104 key genes related to walnut oil accumulation were screened. The phospholipid:diacylglycerol acyltransferase metabolic pathway may contribute more to oil accumulation in walnut. 16 miRNA-mRNA regulatory modules related to walnut oil accumulation and fatty acid synthesis were constructed. 8 known miRNAs and 9 novel miRNAs regulate 28 genes involved in fatty acid (FA) metabolism and lipid synthesis. Among them, jre-miRn105, jre-miRn434, jre-miR477d and jre-miR156a.2 are key miRNAs that regulate walnut FA synthesis. Jre-miRn411 and jre-miR399a.1 are closely related to oil accumulation. These data provide new insights and lay the foundation for subsequent studies on walnut FA synthesis and oil accumulation.

High-Throughput Sequencing Reveals the Regulatory Networks of Transcriptome and Small RNAs During the Defense Against Marssonina brunnea in Poplar

“…The DEM (differentially expressed miRNA) selection was based on |log2 fold change| ≥ 1 and p -value < 0.05 using DESeq. Finally, sequences for target genes of DEMs were predicted by psRobot_tar and TargetFinder (Bo and Wang, 2005 ; Zhao et al, 2020 ). The analysis script of these two programs was shown in Supplementary Table 7 .…”

Identification of miRNA–mRNA Regulatory Modules Involved in Lipid Metabolism and Seed Development in a Woody Oil Tree (Camellia oleifera)

“…Target genes that contain a sequence with almost complete complementarity to the miRNA are cleaved by the RISC at a specific site opposite to the 10th or 11th nucleotide in the miRNA [ 8 ]. Studies have confirmed the key roles of miRNAs in various biological and metabolic processes in plants, such as fatty acid biosynthesis [ 9 , 10 ], lipid metabolism [ 11 , 12 ], growth and development [ 13 ], responses to stresses [ 14 ], and cellular proliferation and differentiation [ 15 ]. As of 3 December 2021, a total of 38,589 miRNA sequences have been deposited in the database, miRBase [ 16 , 17 ], and many conserved and novel miRNAs have been identified in soybeans [ 18 ], Arabidopsis [ 19 ], Brassica napus [ 20 ], rice [ 21 ], peonies [ 22 ], and maize [ 23 ].…”

“…As of 3 December 2021, a total of 38,589 miRNA sequences have been deposited in the database, miRBase [ 16 , 17 ], and many conserved and novel miRNAs have been identified in soybeans [ 18 ], Arabidopsis [ 19 ], Brassica napus [ 20 ], rice [ 21 ], peonies [ 22 ], and maize [ 23 ]. In walnuts, jre-miRn105, jre-miRn434, jre-miR477d, and jre-miR156a.2 are key miRNAs that regulate FA synthesis, and jre-miRn411 and jre-miR399a.1 are closely related to oil accumulation [ 10 ]. MiRNAs control and influence a variety of physiological processes by regulating target genes.…”

“…Currently, miRNA–mRNA regulatory modules controlling multiple biological and metabolic processes have been revealed via the functional analysis of woody oil plants, such as hazel [ 36 ], oil palm [ 37 ], olive [ 38 ], peony [ 22 ], walnut [ 10 ], and yellowhorn [ 11 ]. In developing sea buckthorn seeds, 19 (4 known and 15 novel) and 22 (6 known and 16 novel) miRNAs were found to be involved in lipid biosynthesis and seed size, respectively [ 39 ], and an integrated analysis of the mRNA and miRNA transcriptomes and the qRT-PCR revealed key miRNAs and their targets (miR164d- ARF2 , novelmiRNA-108- ACC , novelmiRNA-23- GPD 1, novelmiRNA-58- DGAT 1, and novelmiRNA-191- DGAT 2) that are potentially involved in the seed size and lipid biosynthesis in the seeds of sea buckthorn.…”

Insights Into Walnut Lipid Metabolism From Metabolome and Transcriptome Analysis

“…Huang et al (2021) found 108 genes related to lipid synthesis, including 60 genes for the fatty acid synthesis pathway, 33 for the triglyceride synthesis pathway, seven genes for the formation of oil bodies, and eight transcription factors. By analyzing the miRNA and mRNA transcriptome data of walnut kernels at different developmental stages, 104 miRNAs related to oil accumulation were found (Zhao et al, 2020). Lipid synthesis is the result of the interaction of a multilayer network.…”