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First published online May 29, 2009
Journal of Experimental Biology 212, 1781-1793 (2009)
Published by The Company of Biologists 2009
doi: 10.1242/jeb.029918
Embryonic temperature affects muscle fibre recruitment in adult zebrafish: genome-wide changes in gene and microRNA expression associated with the transition from hyperplastic to hypertrophic growth phenotypes
1 School of Biology, University of St Andrews, St Andrews, Fife KY16 8LB,
UK
2 School of Biological Sciences, University of East Anglia, Norwich, Norfolk NR4
7TJ, UK
* Author for correspondence (e-mail: iaj{at}st-and.ac.uk)
Accepted 12 March 2009
We investigated the effects of embryonic temperature (ET) treatments (22,
26 and 31°C) on the life-time recruitment of fast myotomal muscle fibres
in zebrafish Danio rerio L. reared at 26/27°C from hatching. Fast
muscle fibres were produced until 25 mm total length (TL) at 22°C ET, 28
mm TL at 26°C ET and 23 mm TL at 31°C ET. The final fibre number (FFN)
showed an optimum at 26°C ET (3600) and was 19% and 14% higher than for
the 22°C ET (3000) and 31°C ET (3100) treatments, respectively.
Further growth to the maximum TL of 
48 mm only involved fibre
hypertrophy. Microarray experiments were used to determine global changes in
microRNA (miRNA) and mRNA expression associated with the transition from the
hyperplasic myotube-producing phenotype (M+, 10–12 mm TL) to
the hypertrophic growth phenotype (M–, 28–31 mm TL) in
fish reared at 26–27°C over the whole life-cycle. The expression of
miRNAs and mRNAs obtained from microarray experiments was validated by
northern blotting and real-time qPCR in independent samples of fish with the
M+ and M– phenotype. Fourteen down-regulated and
15 up-regulated miRNAs were identified in the M– phenotype
together with 34 down-regulated and 30 up-regulated mRNAs (>2-fold;
P<0.05). The two most abundant categories of down-regulated genes
in the M– phenotype encoded contractile proteins (23.5%) and
sarcomeric structural/cytoskeletal proteins (14.7%). In contrast, the most
highly represented up-regulated transcripts in the M–
phenotype were energy metabolism (26.7%) and immune-related (20.0%) genes. The
latter were mostly involved in cell–cell interactions and cytokine
pathways and included β-2-microglobulin precursor (b2m), an
orthologue of complement component 4, invariant chain-like protein 1
(iclp), CD9 antigen-like (cd9l), and tyrosine kinase,
non-receptor (tnk2). Five myosin heavy chain genes that were
down-regulated in the M– phenotype formed part of a tandem
repeat on chromosome 5 and were shown by in situ hybridisation to be
specifically expressed in nascent myofibres. Seven up-regulated miRNAs in the
M– phenotype showed reciprocal expression with seven mRNA
targets identified in miRBase Targets version 5
(http://microrna.sanger.ac.uk/targets/v5/),
including asporin (aspn) which was the target for four miRNAs. Eleven
down-regulated miRNAs in the M– phenotype had predicted
targets for seven up-regulated genes, including dre-miR-181c which had five
predicted mRNA targets. These results provide evidence that miRNAs play a role
in regulating the transition from the M+ to the M–
phenotype and identify some of the genes and regulatory interactions
involved.
Key words: Danio rerio, microRNA, developmental plasticity, temperature, muscle growth, muscle hyperplasia, gene expression, myosin heavy chains, β-2-microglobulin
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