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Dynamic Regulation of Novel and Conserved miRNAs Across Various Tissues of Diverse Cucurbit Species

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Abstract

MicroRNA genes (miRNAs) encoding small non-coding RNAs are abundant in plant genomes and play a key role in regulating several biological mechanisms. Five conserved miRNAs, miR156, miR168-1, miR168-2, miR164, and miR166 were selected for analysis from the 21 known plant miRNA families that were recovered from deep sequencing data of small RNA libraries of pumpkin and squash. A total of six novel miRNAs that were not reported before were found to have precursors with reliable fold-back structures and hence considered novel and were designated as cuc_nov_miRNAs. A set of five conserved, six novel miRNAs, and five uncharacterized small RNAs from the deep sequencing data were profiled for their dynamic regulation using qPCR. The miRNAs were evaluated for differential regulation across the tissues among four diverse cucurbit species, including pumpkin and squash (Cucurbita moschata Duch. Ex Poir. and Cucurbita pepo L.), bitter melon (Momordica charantia L.), and Luffa (Loofah) (Luffa acutangula Roxb.). Expression analysis revealed differential regulation of various miRNAs in leaf, stem, and fruit tissues. Importantly, differences in the expression levels were also found in the leaves and fruits of closely related C. moschata and C. pepo. Comparative miRNA profiling and expression analysis in four cucurbits led to identification of conserved miRNAs in cucurbits. Predicted targets for two of the conserved miRNAs suggested miRNAs are involved in regulating similar biological mechanisms in various species of cucurbits.

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Acknowledgments

The authors are grateful to Dr. Jarret, Plant Genetic Resources Conservation Unit, USDA-ARS, Griffin, GA, 30223 for providing the seeds of germplasm accessions. Funding support is provided by NSF-EPSCOR no. 1003907, Gus R. Douglass Institute, and USDA-NIFA Research (2010-38821-21476).

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Authors and Affiliations

  1. Gus R. Douglass Institute, Department of Biology, West Virginia State University, Institute, WV, 25112-1000, USA

    Sumanth Manohar, Padma Nimmakayala, Yan Tomason, Aldo Almeida & Umesh K. Reddy

  2. Department of Biochemistry and Molecular Biology, Oklahoma State University, Stillwater, OK, 74078, USA

    Guru Jagadeeswaran & Ramanjulu Sunkar

  3. U. S. Vegetable Laboratory, USDA-ARS, 2875 Savannah Highway, Charleston, SC, 29414, USA

    Amnon Levi

  4. Dnipropetrovsk State Agrarian University, Dnipropetrovsk, Ukraine, 49600

    Yan Tomason

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  1. Sumanth Manohar
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  8. Umesh K. Reddy
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Corresponding author

Correspondence to Umesh K. Reddy.

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Umesh K. Reddy contributed equally with the first author

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Supplementary Fig. 1

Heatmap of expression profiles of conserved miRNAs in leaf, stem, flesh, rind, and placenta of various cucurbit species. The color white to light blue to dark blue to black represents the gradation of scale of relative miRNA expression (log2). (JPEG 44 kb)

High-resolution image (TIFF 130628 kb)

Supplementary Fig. 2

Heatmap of expression profiles of novel miRNAs in leaf, stem, flesh, rind, and placenta of various cucurbit species. The color white to light blue to dark blue to black represents the gradation of scale of relative miRNA expression (log2). (JPEG 46 kb)

High-resolution image (TIFF 115967 kb)

Supplementary Fig. 3

Heatmap of expression profiles of small RNAs in leaf, stem, flesh, rind, and placenta of various cucurbit species. The color white to light blue to dark blue to black represents the gradation of scale of relative miRNA expression (log2). (JPEG 43 kb)

High-resolution image (TIFF 128602 kb)

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Manohar, S., Jagadeeswaran, G., Nimmakayala, P. et al. Dynamic Regulation of Novel and Conserved miRNAs Across Various Tissues of Diverse Cucurbit Species. Plant Mol Biol Rep 31, 335–343 (2013). https://doi.org/10.1007/s11105-012-0506-7

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