Activation of GH signaling and GH-independent stimulation of growth in zebrafish by introduction of a constitutively activated GHR construct

• Published in: Transgenic research 2011-06, Vol.20 (3), p.557-567
• Main Authors: Ishtiaq Ahmed, A. S, Xiong, Feng, Pang, Shao-Chen, He, Mu-Dan, Waters, Michael J, Zhu, Zuo-Yan, Sun, Yong-Hua
• Format: Article
• Language: English
• Subjects: Animal Genetics and Genomics; Animals; Animals, Genetically Modified - embryology; Animals, Genetically Modified - genetics; Animals, Genetically Modified - growth & development; Biological and medical sciences; Biomedical and Life Sciences; Biomedical Engineering/Biotechnology; Biotechnology; Danio rerio; Dimerization; fish; Freshwater; Fundamental and applied biological sciences. Psychology; Gene Expression Regulation, Developmental; gene overexpression; gene transfer; Gene Transfer Techniques; Genetic Engineering; Genetic technics; Growth Hormone - genetics; Growth Hormone - metabolism; insulin-like growth factor I; Life Sciences; Methods. Procedures. Technologies; Molecular Medicine; Original Paper; Plant Genetics and Genomics; protein products; Receptors, Somatotropin - genetics; Receptors, Somatotropin - metabolism; Signal Transduction; somatotropin; Transcriptional Activation; transgenic animals; Transgenic animals and transgenic plants; Transgenics; Zebrafish - embryology; Zebrafish - genetics; Zebrafish - growth & development
• ISSN: 0962-8819; 1573-9368
• DOI: 10.1007/s11248-010-9439-9

Growth hormone (GH) gene transfer can markedly increase growth in transgenic fish. In the present study we have developed a transcriptional assay to evaluate GH-signal activation (GHSA) in zebrafish embryos. By analyzing the transcription of c-fos and igf1, and the promoter activity of spi2.1, in zebrafish embryos injected with different constructs, we found that overexpression of either GH or growth hormone receptor (GHR) resulted in GHSA, while a synergetic overexpression of GH and GHR gave greater activation. Conversely, overexpression of a C-terminal truncated dominant-negative GHR (ΔC-GHR) efficiently blocked GHSA epistatic to GH overexpression, demonstrating the requirement for a full GHR homodimer in signaling. In view of the importance of signal-competent GHR dimerization by extracellular GH, we introduced into zebrafish embryos a constitutively activated GHR (CA-GHR) construct, which protein products constitutively dimerize the GHR productively by Jun-zippers to activate downstream signaling in vitro. Importantly, overexpression of CA-GHR led to markedly higher level of GHSA than the synergetic overexpression of GH and GHR. CA-GHR transgenic zebrafish were then studied in a growth trial. The transgenic zebrafish showed higher growth rate than the control fish, which was not achievable by GH transgenesis in these zebrafish. Our study demonstrates GH-independent growth by CA-GHR in vivo which bypasses normal IGF-1 feedback control of GH secretion. This provides a novel means of producing growth enhanced transgenic animals based on molecular protein design.