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Functional characterization of the Japanese flounder (Paralichthys olivaceus) Sox2 gene promoter

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Abstract

Sox2 has essential roles in early embryogenesis and the development of the central nervous system. Sox2 is also necessary in maintaining the identity of progenitor cells. In our study, a 1.8-kb fragment of the 5′ flanking region of Paralichthys olivaceus Sox2 (Po-Sox2) gene was cloned and functionally characterized. The activity and specificity of Po-Sox2 promoter were analyzed by comparing various deletion mutants for their ability to direct luciferase and GFP expression in flounder brain cell line. Results indicated that the basal promoter is located between −978 and −442 bp, and the region from −1370 to −978 bp enhances the promoter activity. The regulatory elements in the −1370 to −442 bp fragment were further investigated. Many binding sites of transcription factors closely related to neurogenesis and stem cell properties were found in this region. Mutational analysis indicated that Nanog, Pax6, p53, and POU binding sites play functional roles in the transcription of Po-Sox2 gene, whereas NF-Y binding sites did not affect this gene. In vivo studies using transient transgenic zebrafish embryos showed that the Po-Sox2 promoter region can drive GFP expression in brain, yolk syncytial layer, and notochord. Our results provide valuable information in understanding the molecular regulatory mechanisms of Po-Sox2 gene during neurogenesis and embryonic development.

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Abbreviations

GFP:

Green fluorescent protein

CNS:

Central nervous system

NSCs:

Neural stem cells

FBCs:

Japanese flounder brain cells

FBC:

Flounder brain cell line

PCR:

Polymerase chain reaction

CMV:

Cytomegalovirus

PTU:

1-Phenyl 2-thiourea

TF:

Transcription factor

TSS:

Transcriptional start site

UTR:

Untranslated region

dph:

Days post-hatching

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Acknowledgments

This study was supported by grants from the National High Technology Research and Development Program of China (No. 2012AA10A401) and China Postdoctoral Science Foundation (2015M551960). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

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Authors and Affiliations

  1. Key Laboratory of Marine Genetics and Breeding (Ocean University of China), Ministry of Education, Qingdao, 266003, China

    Wei Liu, Huayu Song, Aoyun Li, Xinxin Du, Yuezhong Liu, Yan He, Quanqi Zhang & Jie Qi

  2. College of Marine Life Science, Ocean University of China, No. 5 Yushan Road, Qingdao, 266003, China

    Jie Qi

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  1. Wei Liu
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  2. Huayu Song
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  3. Aoyun Li
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  5. Yuezhong Liu
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  6. Yan He
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  7. Quanqi Zhang
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Correspondence to Jie Qi.

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Liu, W., Song, H., Li, A. et al. Functional characterization of the Japanese flounder (Paralichthys olivaceus) Sox2 gene promoter. Fish Physiol Biochem 42, 1275–1285 (2016). https://doi.org/10.1007/s10695-016-0216-4

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  • DOI: https://doi.org/10.1007/s10695-016-0216-4

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