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Integrated transcriptome, small RNA and degradome sequencing approaches proffer insights into chlorogenic acid biosynthesis in leafy sweet potato.

ABSTRACT: Leafy sweet potato is rich in total phenolics (TP) which play key roles in health protection, the chlorogenic acid (CGA) constitutes the major components of phenolic compounds in leafy sweet potato. Unfortunately, the mechanism of CGA biosynthesis in leafy sweet potato is unclear. To dissect the mechanisms of CGA biosynthesis, we performed transcriptome, small RNA (sRNA) and degradome sequencing of one low-CGA content and one high-CGA content genotype at two stages. A total of 2,333 common differentially expressed genes (DEGs) were identified, and the enriched DEGs were related to photosynthesis, starch and sucrose metabolism and phenylpropanoid biosynthesis. The functional genes, such as CCR, CCoAOMT and HCT in the CGA biosynthetic pathway were down-regulated, indicating that the way to lignin was altered, and two possible CGA biosynthetic routes were hypothesized. A total of 38 DE miRNAs were identified, and 1,799 targets were predicated for 38 DE miRNAs by using in silico approaches. The target genes were enriched in lignin and phenylpropanoid catabolic processes. Transcription factors (TFs) such as apetala2/ethylene response factor (AP2/ERF) and Squamosa promoter binding protein-like (SPL) predicated in silico were validated by degradome sequencing. Association analysis of the DE miRNAs and transcriptome datasets identified that miR156 family negatively targeted AP2/ERF and SPL. Six mRNAs and six miRNAs were validated by qRT-PCR, and the results showed that the expression levels of the mRNAs and miRNAs were consistent with the sequencing data. This study established comprehensive functional genomic resources for the CGA biosynthesis, and provided insights into the molecular mechanisms involving in this process. The results also enabled the first perceptions of the regulatory roles of mRNAs and miRNAs, and offered candidate genes for leafy sweet potato improvements.

SUBMITTER: Liu Y

PROVIDER: S-EPMC7822329 | biostudies-literature | 2021

REPOSITORIES: biostudies-literature

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Integrated transcriptome, small RNA and degradome sequencing approaches proffer insights into chlorogenic acid biosynthesis in leafy sweet potato.

Liu Yi Y Su Wenjin W Wang Lianjun L Lei Jian J Chai Shasha S Zhang Wenying W Yang Xinsun X

PloS one 20210122 1

Leafy sweet potato is rich in total phenolics (TP) which play key roles in health protection, the chlorogenic acid (CGA) constitutes the major components of phenolic compounds in leafy sweet potato. Unfortunately, the mechanism of CGA biosynthesis in leafy sweet potato is unclear. To dissect the mechanisms of CGA biosynthesis, we performed transcriptome, small RNA (sRNA) and degradome sequencing of one low-CGA content and one high-CGA content genotype at two stages. A total of 2,333 common diffe ...[more]

PMID: 33481815

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Project description:BackgroundStarch biosynthesis in endosperm is a key process influencing grain yield and quality in maize. Although a number of starch biosynthetic genes have been well characterized, the mechanisms by which the expression of these genes is regulated, especially in regard to microRNAs (miRNAs), remain largely unclear.ResultsSequence data for small RNAs, degradome, and transcriptome of maize endosperm at 15 and 25 d after pollination (DAP) from inbred lines Mo17 and Ji419, which exhibit distinct starch content and starch granule structure, revealed the mediation of starch biosynthetic pathways by miRNAs. Transcriptome analysis of these two lines indicated that 33 of 40 starch biosynthetic genes were differentially expressed, of which 12 were up-regulated in Ji419 at 15 DAP, one was up-regulated in Ji419 at 25 DAP, 14 were up-regulated in Ji419 at both 15 and 25 DAP, one was down-regulated in Ji419 at 15 DAP, two were down-regulated in Ji419 at 25 DAP, and three were up-regulated in Ji419 at 15 DAP and down-regulated in Ji419 at 25 DAP, compared with Mo17. Through combined analyses of small RNA and degradome sequences, 22 differentially expressed miRNAs were identified, including 14 known and eight previously unknown miRNAs that could target 35 genes. Furthermore, a complex co-expression regulatory network was constructed, in which 19 miRNAs could modulate starch biosynthesis in endosperm by tuning the expression of 19 target genes. Moreover, the potential operation of four miRNA-mediated pathways involving transcription factors, miR169a-NF-YA1-GBSSI/SSIIIa and miR169o-GATA9-SSIIIa/SBEIIb, was validated via analyses of expression pattern, transient transformation assays, and transactivation assays.ConclusionOur results suggest that miRNAs play a critical role in starch biosynthesis in endosperm, and that miRNA-mediated networks could modulate starch biosynthesis in this tissue. These results have provided important insights into the molecular mechanism of starch biosynthesis in developing maize endosperm.

Project description:BACKGROUND: Sweet potato (Ipomoea batatas L. [Lam.]) ranks among the top six most important food crops in the world. It is widely grown throughout the world with high and stable yield, strong adaptability, rich nutrient content, and multiple uses. However, little is known about the molecular biology of this important non-model organism due to lack of genomic resources. Hence, studies based on high-throughput sequencing technologies are needed to get a comprehensive and integrated genomic resource and better understanding of gene expression patterns in different tissues and at various developmental stages. METHODOLOGY/PRINCIPAL FINDINGS: Illumina paired-end (PE) RNA-Sequencing was performed, and generated 48.7 million of 75 bp PE reads. These reads were de novo assembled into 128,052 transcripts (? 100 bp), which correspond to 41.1 million base pairs, by using a combined assembly strategy. Transcripts were annotated by Blast2GO and 51,763 transcripts got BLASTX hits, in which 39,677 transcripts have GO terms and 14,117 have ECs that are associated with 147 KEGG pathways. Furthermore, transcriptome differences of seven tissues were analyzed by using Illumina digital gene expression (DGE) tag profiling and numerous differentially and specifically expressed transcripts were identified. Moreover, the expression characteristics of genes involved in viral genomes, starch metabolism and potential stress tolerance and insect resistance were also identified. CONCLUSIONS/SIGNIFICANCE: The combined de novo transcriptome assembly strategy can be applied to other organisms whose reference genomes are not available. The data provided here represent the most comprehensive and integrated genomic resources for cloning and identifying genes of interest in sweet potato. Characterization of sweet potato transcriptome provides an effective tool for better understanding the molecular mechanisms of cellular processes including development of leaves and storage roots, tissue-specific gene expression, potential biotic and abiotic stress response in sweet potato.

Project description:In various plant species, many transcription factors (TFs), such as MYB, bHLH, and WD40, have been identified as regulators of anthocyanin biosynthesis in underground organs. However, the regulatory elements of anthocyanin biosynthesis in the tuberous roots of sweet potato have not been elucidated yet. Here, we selected the purple-fleshed sweet potato cultivar "Zhezi1" (ZZ P ) and its spontaneous yellow-fleshed mutant "Xinli" (XL Y ) to investigate the regulatory mechanism of the anthocyanin biosynthesis in the tuberous roots of sweet potato. By analyzing the IbMYB1 genotype in ZZ P and XL Y , we found that the IbMYB1-2, a MYB TF involved in anthocyanin biosynthesis, was missing in the XL Y genome, which might lead to an extreme decrease in anthocyanins in XL Y . A comparative transcriptome analysis of ZZ P and XL Y was conducted to find the TFs involved in anthocyanin biosynthesis in ZZ P and XL Y . The anthocyanin structural genes were significantly enriched among the differentially expressed genes. Moreover, one MYB activator (IbMYB1), one bHLH (IbbHLH2), three WRKY activator candidates (IbWRKY21, IbWRKY24, and IbWRKY44), and two MYB repressors (IbMYB27 and IbMYBx-ZZ) were highly expressed in ZZ P accompanied with anthocyanin structural genes. We also tested the expression of these TFs in six purple- and two orange-fleshed sweet potato cultivars. Interestingly, most of these TFs were significantly positively correlated with anthocyanin contents in these cultivars. The function of the anthocyanin biosynthesis repression of IbMYB27 and IbMYBx-ZZ was verified through transient co-transformation with IbMYB1 into tobacco leaves. Further functional verification of the above TFs was conducted by Y2H, BiFC, and dual-luciferase assays. These tests showed that the MYB-bHLH-WD40/MYB-bHLH-WD40-WRKY complex activated the promoter of anthocyanin structural gene IbDFR and promoters for IbWRKY44, IbMYB27, and IbMYBx-ZZ, indicating reinforcement and feedback regulation to maintain the level of anthocyanin accumulation in the tuberous roots of purple-fleshed sweet potato. These results may provide new insights into the regulatory mechanism of anthocyanin biosynthesis and accumulation in underground organs of sweet potatoes.

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Integrated transcriptome, small RNA and degradome sequencing approaches proffer insights into chlorogenic acid biosynthesis in leafy sweet potato.

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Integrated transcriptome, small RNA and degradome sequencing approaches proffer insights into chlorogenic acid biosynthesis in leafy sweet potato.

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