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Multiple microRNAs Regulate the Floral Development and Sex Differentiation in the Dioecious Cucurbit Coccinia grandis (L.) Voigt

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Abstract

Despite having a small genome and heteromorphic sex chromosomes, the molecular basis of sex differentiation in the dioecious cucurbit, Coccinia grandis, largely remains unclear. Increasing evidences indicate a vital regulatory role of microRNAs in the reproductive development of plants. In this study, we used a combination of high-throughput small RNA sequencing and computational analysis to identify sex-specific miRNAomes from male and female buds of C. grandis. Ninety-eight conserved miRNAs from 22 families and 44 novel miRNAs specific to C. grandis were detected. Comparative profiling together with Northern blot and qRT-PCR analysis revealed 41 significantly differentially expressed (DE) miRNAs, of which 16 could be fundamental to the regulation of sexual dimorphism. One hundred six target genes were predicted for 35 DE miRNAs that were significantly involved in flower organogenesis, phytohormone signaling, metabolism, transcription regulation, and DNA methylation. Temporal analysis of a set of 16 target genes at three stages of the bud development revealed a reciprocal alteration in their expression pattern with the complementary miRNAs. Further, the miR167a-1, miR393a, miR398b, and miRn9 mediated down-regulation of four predicted targets associated with reproductive organ development in C. grandis was confirmed through transient co-expression in Nicotiana benthaminana. Taken together, the present study represents the first report suggesting that multiple miRNA-mediated gene silencing cascade could be involved in regulating the molecular basis of sex differentiation in C. grandis.

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Acknowledgements

We thank DST-FIST, Govt. of India, for the research infrastructure facilities provided to Centre of Biotechnology, Siksha O Anusandhan University.

Funding

This research is supported by grants from Dept. of Biotechnology (DBT), Govt. of India (grant no. BT/PR3919/PBD/16/959/2011). JNM is supported by research fellowships from Dept. of Biotechnology (DBT), Govt. of India.

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Authors and Affiliations

  1. Functional Genomics Laboratory, Centre of Biotechnology, Siksha O Anusandhan University, Kalinga Nagar, Ghatikia, Bhubaneswar, Odisha, 751003, India

    Jatindra Nath Mohanty, Subodh Kumar Chand & Raj Kumar Joshi

  2. Department of Biotechnology, Rama Devi Women’s University, Vidya Vihar, Bhubaneswar, Odisha, 751022, India

    Raj Kumar Joshi

Authors
  1. Jatindra Nath Mohanty
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  2. Subodh Kumar Chand
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  3. Raj Kumar Joshi
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Contributions

RKJ conceived, designed, and supervised the research work. JNM performed the experiments and SKC analyzed the data. JNM and SKC wrote the manuscript. RKJ provided inputs on data presentation and critically reviewed the manuscript. All authors read and approved the final manuscript.

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Correspondence to Raj Kumar Joshi.

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Electronic Supplementary Material

Figure S1

Abundance of known (A) and novel (B) miRNA families identified from Coccinia grandis. (JPG 648 kb)

Figure S2

Gene ontology (GO) classification of the target genes for differentially expressed microRNAs. GO terms were assigned based on significant hits and classified into three main categories: Biological process, cellular component and molecular function. (JPG 215 kb)

Table S1

Primers and probes used in the present study. (DOCX 21 kb)

Table S2

Detailed information of conserved miRNAs isolated from Coccinia grandis. (XLSX 36 kb)

Table S3

Novel miRNAs identified from Coccinia grandis. (XLS 41 kb)

Table S4

Differentially expressed members of C. grandis miRNA families. (XLSX 15 kb)

Table S5

List of predicted target genes for the differentially expressed miRNAs. (XLSX 20 kb)

Table S6

Gene ontology (GO) based functional classification of conserved and novel miRNA targets with miRNA. (XLSX 12 kb)

Table S7

Functional classification of conserved and novel miRNA targets using Kyoto Enclycopedia of Genes and Genomes (KEGG) analysis. (XLSX 10 kb)

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Mohanty, J.N., Chand, S.K. & Joshi, R.K. Multiple microRNAs Regulate the Floral Development and Sex Differentiation in the Dioecious Cucurbit Coccinia grandis (L.) Voigt. Plant Mol Biol Rep 37, 111–128 (2019). https://doi.org/10.1007/s11105-019-01143-8

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