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The Molecular Differentiation of Anatomically Paired Left and Right Mantles of the Pacific Oyster Crassostrea gigas

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Abstract

Left-right (L-R) asymmetry is controlled by gene regulation pathways for the L-R axis, and in vertebrates, the gene Pitx2 in TGF−β signaling pathway plays important roles in the asymmetrical formation of organs. However, less is known about the asymmetries of anatomically identical paired organs, as well as the transcriptional regulation mechanism of the gene Pitx in invertebrates. Here, we report the molecular biological differences between the left and right mantles of an invertebrate, the Pacific oyster Crassostrea gigas, and propose one possible mechanism underlying those differences. RNA sequencing (RNA-seq) analysis indicated that the paired organs showed different gene expression patterns, suggesting possible functional differences in shell formation, pheromone signaling, nerve conduction, the stress response, and other physiological processes. RNA-seq and real-time qPCR analysis indicated high right-side expression of the Pitx homolog (cgPitx) in oyster mantle, supporting a conserved role for Pitx in controlling asymmetry. Methylation-dependent restriction-site associated DNA sequencing (MethylRAD) identified a methylation site in the promoter region of cgPitx and showed significantly different methylation levels between the left and right mantles. This is the first report, to our knowledge, of such a difference in methylation in spiralians, and it was further confirmed in 18 other individuals by using a pyrosequencing assay. The miRNome analysis and the TGF-β receptor/Smad inhibition experiment further supported that several genes in TGF−β signaling pathway may be related with the L/R asymmetry of oyster mantles. These results suggested that the molecular differentiation of the oyster’s paired left and right mantles is significant, TGF−β signaling pathway could be involved in establishing or maintaining the asymmetry, and the cgPitx gene as one of genes in this pathway; the different methylation levels in its promoter regions between L/R mantles was the one of possible mechanisms regulating the left-right functional differentiation.

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Acknowledgements

We thank Peter W H Holland and Jordi Paps for helpful suggestions upon critical reading of the manuscript, as well as Guofan Zhang for helpful discussion regarding the experimental design.

Funding

This research was supported by the Modern Agricultural Industry Technology System of Shandong Province, China (SDAIT-14-03), the Key R & D Program of Yantai City, China (No. 2017ZH054), the Innovation Plan for Marine/Fishery Science and Technology of Shandong Province, China (No. 2017YY03), the National Natural Science Foundation of China (Nos. 31302181, 41776152), the Shandong Provincial Natural Science Foundation, China (Nos. ZR2013CM026, ZR2017BC058), and A Project of Shandong Province Higher Educational Science and Technology Program (No. J17KA129).

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  1. Xiaotong Wang

    Present address: School of Agriculture, Ludong University, Yantai, 264025, China

  2. Lei Wei and Fei Xu contributed equally to this work.

Authors and Affiliations

  1. School of Agriculture, Ludong University, Yantai, 264025, China

    Lei Wei, Wenchao Yu, Cheng He & Qiuyun Jiang

  2. Institute of Oceanology, Chinese Academy of Sciences, Qingdao, 266071, China

    Fei Xu

  3. School of Life Sciences, Shandong University, Jinan, 250100, China

    Yuzhi Wang

  4. Changdao Enhancement and Experiment Station, Chinese Academy of Fishery Sciences, Changdao, 265800, China

    Zhongqiang Cai

  5. Qingdao OE Biotechnology Company Limited, Qingdao, 266101, China

    Xiqiang Xu

  6. Marine Biology Institute of Shandong Province, Qingdao, 266104, China

    Wen Guo

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  1. Lei Wei
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Different expressed miRNA related with TGF−β pathway. (XLSX 9 kb)

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Wei, L., Xu, F., Wang, Y. et al. The Molecular Differentiation of Anatomically Paired Left and Right Mantles of the Pacific Oyster Crassostrea gigas. Mar Biotechnol 20, 425–435 (2018). https://doi.org/10.1007/s10126-018-9806-8

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