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Small RNA sequencing identifies cucumber miRNA roles in waterlogging-triggered adventitious root primordia formation

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Abstract

The formation of adventitious roots (ARs) is a key morphological adaptation of cucumber (Cucumis sativus L.) to waterlogging stress. MicroRNAs (miRNAs) constitute a group of non-coding small RNAs (sRNA) that play crucial roles in regulating diverse biological processes, including waterlogging acclimation. However, which specific miRNAs and how they are involved in waterlogging-triggered de novo AR primordia formation are not fully known. Here, Illumina sRNA sequencing was applied to sequence six sRNA libraries generated from the waterlogging-tolerant cucumber Zaoer-N after 48 h of waterlogging and the control. A total of 358 cucumber miRNAs, 312 known and 46 novel, were obtained. Among them, 23 were differentially expressed, with 10 and 13 being up- and downregulated, respectively. A qPCR expression study confirmed that the identified differentially expressed miRNAs were credible. A total of 657 putative miRNA target genes were predicted for the 23 miRNAs using an in silico approach. A gene ontology enrichment analysis revealed that target genes functioning in cell redox homeostasis, cytoskeleton, photosynthesis and cell growth were over-represented. In total, 58 of the 657 target genes showed inverse expression patterns compared with their respective miRNAs through a combined analysis of sRNA- and RNA-sequencing-based transcriptome datasets using the same experimental design. The target gene annotation included a peroxidase, a GDSL esterases/lipase and two heavy metal-associated isoprenylated plant proteins. Our results provide an important framework for understanding the unique miRNA patterns seen in responses to waterlogging and the miRNA-mediated formation of de novo AR primordia in cucumber.

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Acknowledgements

This research was supported by the National Natural Science Foundation of China (Grant Nos. 31171978, 31672176 and 31801883) and the Natural Science Foundation of the Jiangsu Higher Education Institutions of China (18KJB210014). We thank Lesley Benyon, PhD, from Liwen Bianji, Edanz Group China (www.liwenbianji.cn/ac), for editing the English text of a draft of this manuscript.

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Authors and Affiliations

  1. School of Horticulture and Plant Protection, Yangzhou University, Yangzhou, 225009, Jiangsu, China

    Xuewen Xu, Kaixuan Wang, Jiawei Pan & Xuehao Chen

  2. Joint International Research Laboratory of Agriculture & Agri-Product Safety, Yangzhou University, Yangzhou, 225009, Jiangsu, China

    Xuewen Xu, Jiawei Pan & Xuehao Chen

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  2. Kaixuan Wang
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XC conceived and designed the experiment. XX and KW performed the experiments. XX and JP interpreted the data and wrote the manuscript. All authors reviewed and approved this submission.

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Correspondence to Xuehao Chen.

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11033_2019_5084_MOESM1_ESM.xlsx

Online Resource 1 (XLSX 11 kb). Distribution of cucumber small RNAs in the six sequencing libraries. NAT, natural antisense transcripts. C1, C2 and C3 represent the three replications of control hypocotyl samples; W1, W2 and W3 represent the three replications of waterlogged hypocotyl samples

11033_2019_5084_MOESM2_ESM.xlsx

Online Resource 2 (XLSX 78 kb). Known and novel miRNAs identified in cucumber hypocotyls from the six libraries. C1, C2 and C3 represent the three replications of control hypocotyl samples; W1, W2 and W3 represent the three replications of waterlogged hypocotyl samples. TPM represents transcripts per kilobase million

11033_2019_5084_MOESM3_ESM.xlsx

Online Resource 3 (XLSX 74 kb). List of cucumber miRNAs and their target genes predicted by the online psRNATarget server

Online Resource 4 (XLSX 85 kb). GO enrichment analysis of the 657 predicted target genes

11033_2019_5084_MOESM5_ESM.xlsx

Online Resource 5 (XLSX 17 kb). List of the fragments per kilobase million (FPKM) values of the 58 negatively regulated miRNAs target genes that were identified by RNA-seq. C1, C2 and C3 represent the three replications of control hypocotyl samples; W1, W2 and W3 represent the three replications of waterlogged hypocotyl samples

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Xu, X., Wang, K., Pan, J. et al. Small RNA sequencing identifies cucumber miRNA roles in waterlogging-triggered adventitious root primordia formation. Mol Biol Rep 46, 6381–6389 (2019). https://doi.org/10.1007/s11033-019-05084-z

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  • DOI: https://doi.org/10.1007/s11033-019-05084-z

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