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First published online December 14, 2005
Journal of Experimental Biology 209, 188-198 (2006)
Published by The Company of Biologists 2006
doi: 10.1242/jeb.01978

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Molecular cloning and mRNA expression analysis of carp embryonic, slow and cardiac myosin heavy chain isoforms

Yoshiaki Nihei¹, Atsushi Kobiyama^1,*, Daisuke Ikeda¹, Yosuke Ono¹, Satoshi Ohara¹, Nicholas J. Cole², Ian A. Johnston³ and Shugo Watabe^1,

¹ Laboratory of Aquatic Molecular Biology and Biotechnology, Graduate School of Agricultural and Life Sciences, The University of Tokyo, Bunkyo, Tokyo 113-8657, Japan
² Division of Cell and Developmental Biology, MSI/WTB Complex, University of Dundee, Dow Street, Dundee, DDI 5EH, UK
³ Gatty Marine Laboratory, Division of Environmental and Evolutionary Biology, School of Biology, University of St Andrews, St Andrews, Fife KY16 8LB, UK

Author for correspondence (e-mail: awatabe{at}mail.ecc.u-tokyo.ac.jp)

Accepted 6 November 2005

Three embryonic class II myosin heavy chains (MYHs) were cloned from the common carp (Cyprinus carpio L.), MYH_emb1, MYH_emb2 and MYH_emb3. MYH DNA clones were also isolated from the slow muscle of adult carp acclimated to 10°C (MYH_S10) and 30°C (MYH_S30). Phylogenetic analysis demonstrated that MYH_emb1 and MYH_emb2 belonged to the fast skeletal muscle MYH clade. By contrast, the sequence of MYH_emb3 was similar to the adult slow muscle isoforms, MYH_S10 and MYH_S30. MYH_emb1 and MYH_emb2 transcripts were first detected by northern blot analysis in embryos 61 h post-fertilization (h.p.f.) at the heartbeat stage, with peak expression occurring in 1-month-old juveniles. MYH_emb1 continued to be expressed at low levels in 7-month-old juveniles when MYH_emb2 was not detectable. MYH_emb3 transcripts appeared at almost the same stage as MYH_emb1 transcripts did (61 h.p.f.), and these genes showed a similar pattern of expression. Whole mount in situ hybridization analysis revealed that the transcripts of MYH_emb1 and MYH_emb2 were expressed in the inner part of myotome, whereas MYH_emb3 was expressed in the superficial compartment. MYH_S10 and MYH_S30 mRNAs were first detected at hatching. In adult stages, the expression of slow muscle MYH mRNAs was dependent on acclimation temperature. MYH_S10 mRNA was expressed at an acclimation temperature of 10 and 20°C, but not at 30°C. In contrast, MYH_S30 mRNA was strongly expressed at all acclimation temperatures. The predominant MYH transcripts found in adult slow muscle and in embryos at hatching were expressed in adult fast muscle at some acclimation temperatures but not others. A MYH DNA clone was isolated from the cardiac muscle of 10°C-acclimated adult fish (MYH_card). MYH_card mRNA was first detected at 61 h.p.f., but strong signals were only observed in the adult myocardium. The present study has therefore revealed a complex pattern of expression of MYH genes in relation to developmental stage, muscle type and acclimation temperature. None of the skeletal muscle MYHs identified so far was strongly expressed during the late juvenile stage, indicating further developmentally regulated members of the MYH II gene family remain to be discovered.

Key words: carp, Cyprinus carpio, myosin heavy chain, fast skeletal muscle, slow skeletal muscle, myocardium

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