Skip to main content

Advertisement

SpringerLink
Log in

Transgenic Wuzhishan minipigs designed to express a dominant-negative porcine growth hormone receptor display small stature and a perturbed insulin/IGF-1 pathway

  • Original Paper
  • Published:
Transgenic Research Aims and scope Submit manuscript

Abstract

Growth hormone (GH) is an anabolic mitogen with widespread influence on cellular growth and differentiation as well as on glucose and lipid metabolism. GH binding to the growth hormone receptor (GHR) on hepatocytes prompts expression of insulin growth factor I (IGF-1) involved in nutritionally induced compensatory hyperplasia of pancreatic β-cell islets and insulin release. A prolonged hyperactivity of the IGF-1/insulin axis in the face of insulinotropic nutrition, on the other hand, can lead to collapse of the pancreatic islets and glucose intolerance. Individuals with Laron syndrome carry mutations in the GHR gene resulting in severe congenital IGF-1 deficiency and elevated GH serum levels leading to short stature as well as perturbed lipid and glucose metabolism. However, these individuals enjoy a reduced prevalence of acne, cancer and possibly diabetes. Minipigs have become important biomedical models for human conditions due to similarities in organ anatomy, physiology, and metabolism relative to humans. The purpose of this study was to generate transgenic Wuzhishan minipigs by handmade cloning with impaired systemic GHR activity and assess their growth profile and glucose metabolism. Transgenic minipigs featuring overexpression of a dominant-negative porcine GHR (GHRdm) presented postnatal growth retardation and proportionate dwarfism. Molecular changes included elevated GH serum levels and mild hyperglycemia. We believe that this model may prove valuable in the study of GH functions in relation to cancer, diabetes and longevity.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7
Fig. 8

Similar content being viewed by others

References

  • Allevato G, Billestrup N, Goujon L, Galsgaard ED, Norstedt G, Postel-Vinay MC, Kelly PA, Nielsen JH (1995) Identification of phenylalanine 346 in the rat growth hormone receptor as being critical for ligand-mediated internalization and down-regulation. J Biol Chem 270(29):17210–17214

    Article  CAS  PubMed  Google Scholar 

  • Amit T, Youdim MB, Hochberg Z (2000) Clinical review 112: Does serum growth hormone (GH) binding protein reflect human GH receptor function? J Clin Endocrinol Metab 85(3):927–932

    Article  CAS  PubMed  Google Scholar 

  • Amselem S, Duquesnoy P, Duriez B, Dastot F, Sobrier ML, Valleix S, Goossens M (1993) Spectrum of growth hormone receptor mutations and associated haplotypes in Laron syndrome. Hum Mol Genet 2(4):355–359

    Article  CAS  PubMed  Google Scholar 

  • Ayling RM, Ross R, Towner P, Von Laue S, Finidori J, Moutoussamy S, Buchanan CR, Clayton PE, Norman MR (1997) A dominant-negative mutation of the growth hormone receptor causes familial short stature. Nat Genet 16(1):13–14

    Article  CAS  PubMed  Google Scholar 

  • Ayling RM, Ross RJ, Towner P, Von Laue S, Finidori J, Moutoussamy S, Buchanan CR, Clayton PE, Norman MR (1999) New growth hormone receptor exon 9 mutation causes genetic short stature. Acta Paediatr 88(428):168–172, discussion 173

  • Berryman DE, List EO, Coschigano KT, Behar K, Kim JK, Kopchick JJ (2004) Comparing adiposity profiles in three mouse models with altered GH signaling. Growth Horm IGF Res 14(4):309–318

    Article  CAS  PubMed  Google Scholar 

  • Carter-Su C, King AP, Argetsinger LS, Smit LS, Vanderkuur J, Campbell GS (1996) Signalling pathway of GH. Endocr J 43(Suppl):S65–S70

    Article  CAS  PubMed  Google Scholar 

  • Cioffi JA, Wang X, Kopchick JJ (1990) Porcine growth hormone receptor cDNA sequence. Nucleic Acids Res 18(21):6451

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Colosi P, Wong K, Leong SR, Wood WI (1993) Mutational analysis of the intracellular domain of the human growth hormone receptor. J Biol Chem 268(17):12617–12623

    CAS  PubMed  Google Scholar 

  • Cunningham BC, Ultsch M, De Vos AM, Mulkerrin MG, Clauser KR, Wells JA (1991) Dimerization of the extracellular domain of the human growth hormone receptor by a single hormone molecule. Science 254(5033):821–825

    Article  CAS  PubMed  Google Scholar 

  • Dominici FP, Diaz GA, Bartke A, Kopchick JJ, Turyn D (2000) Compensatory alterations of insulin signal transduction in liver of growth hormone receptor knockout mice. J Endocrinol 166(3):579–590

    Article  CAS  PubMed  Google Scholar 

  • Dorer DR, Henikoff S (1994) Expansions of transgene repeats cause heterochromatin formation and gene silencing in Drosophila. Cell 77(7):993–1002

    Article  CAS  PubMed  Google Scholar 

  • Estrada J, Sommer J, Collins B, Mir B, Martin A, York A, Petters RM, Piedrahita JA (2007) Swine generated by somatic cell nuclear transfer have increased incidence of intrauterine growth restriction (IUGR). Cloning Stem Cells 9(2):229–236

    Article  CAS  PubMed  Google Scholar 

  • Fain JN, Ihle JH, Bahouth SW (1999) Stimulation of lipolysis but not of leptin release by growth hormone is abolished in adipose tissue from Stat5a and b knockout mice. Biochem Biophys Res Commun 263(1):201–205

    Article  CAS  PubMed  Google Scholar 

  • Fan Y, Menon RK, Cohen P, Hwang D, Clemens T, DiGirolamo DJ, Kopchick JJ, Le Roith D, Trucco M, Sperling MA (2009) Liver-specific deletion of the growth hormone receptor reveals essential role of growth hormone signaling in hepatic lipid metabolism. J Biol Chem 284(30):19937–19944

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Fang P, Riedl S, Amselem S, Pratt KL, Little BM, Haeusler G, Hwa V, Frisch H, Rosenfeld RG (2007) Primary growth hormone (GH) insensitivity and insulin-like growth factor deficiency caused by novel compound heterozygous mutations of the GH receptor gene: genetic and functional studies of simple and compound heterozygous states. J Clin Endocrinol Metab 92(6):2223–2231

    Article  CAS  PubMed  Google Scholar 

  • Foxcroft GR, Dixon WT, Novak S, Putman CT, Town SC, Vinsky MD (2006) The biological basis for prenatal programming of postnatal performance in pigs. J Anim Sci 84(Suppl):E105–E112

    PubMed  Google Scholar 

  • Freemark M, Avril I, Fleenor D, Driscoll P, Petro A, Opara E, Kendall W, Oden J, Bridges S, Binart N et al (2002) Targeted deletion of the PRL receptor: effects on islet development, insulin production, and glucose tolerance. Endocrinology 143(4):1378–1385

    Article  CAS  PubMed  Google Scholar 

  • Fukunaga R, Ishizaka-Ikeda E, Pan CX, Seto Y, Nagata S (1991) Functional domains of the granulocyte colony-stimulating factor receptor. EMBO J 10(10):2855–2865

    PubMed Central  CAS  PubMed  Google Scholar 

  • Godowski PJ, Leung DW, Meacham LR, Galgani JP, Hellmiss R, Keret R, Rotwein PS, Parks JS, Laron Z, Wood WI (1989) Characterization of the human growth hormone receptor gene and demonstration of a partial gene deletion in two patients with Laron-type dwarfism. Proc Natl Acad Sci USA 86(20):8083–8087

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Goujon L, Allevato G, Simonin G, Paquereau L, Le Cam A, Clark J, Nielsen JH, Djiane J, Postel-Vinay MC, Edery M et al (1994) Cytoplasmic sequences of the growth hormone receptor necessary for signal transduction. Proc Natl Acad Sci USA 91(3):957–961

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Guevara-Aguirre J, Balasubramanian P, Guevara-Aguirre M, Wei M, Madia F, Cheng CW, Hwang D, Martin-Montalvo A, Saavedra J, Ingles S et al (2011) Growth hormone receptor deficiency is associated with a major reduction in pro-aging signaling, cancer, and diabetes in humans. Sci Transl Med 3(70):70ra13

    Article  PubMed Central  PubMed  Google Scholar 

  • Handwerger S, Freemark M (2000) The roles of placental growth hormone and placental lactogen in the regulation of human fetal growth and development. J Pediatr Endocrinol Metab JPEM 13(4):343–356

    Article  CAS  PubMed  Google Scholar 

  • Hansen LH, Wang X, Kopchick JJ, Bouchelouche P, Nielsen JH, Galsgaard ED, Billestrup N (1996) Identification of tyrosine residues in the intracellular domain of the growth hormone receptor required for transcriptional signaling and Stat5 activation. J Biol Chem 271(21):12669–12673

    Article  CAS  PubMed  Google Scholar 

  • Hiragami-Hamada K, Xie SQ, Saveliev A, Uribe-Lewis S, Pombo A, Festenstein R (2009) The molecular basis for stability of heterochromatin-mediated silencing in mammals. Epigenet Chromatin 2(1):14

    Article  Google Scholar 

  • Iida K, Takahashi Y, Kaji H, Takahashi MO, Okimura Y, Nose O, Abe H, Chihara K (1999) Functional characterization of truncated growth hormone (GH) receptor-(1-277) causing partial GH insensitivity syndrome with high GH-binding protein. J Clin Endocrinol Metab 84(3):1011–1016

    CAS  PubMed  Google Scholar 

  • Kenth G, Mergelas JA, Goodyer CG (2008) Developmental changes in the human GH receptor and its signal transduction pathways. J Endocrinol 198(1):71–82

    Article  CAS  PubMed  Google Scholar 

  • Kitamura T, Ido Kitamura Y: (2007) Role of FoxO proteins in pancreatic beta cells. Endocr J 54(4):507–515

    Article  CAS  PubMed  Google Scholar 

  • Laron Z (2004) Laron syndrome (primary growth hormone resistance or insensitivity): the personal experience 1958–2003. J Clin Endocrinol Metab 89(3):1031–1044

    Article  CAS  PubMed  Google Scholar 

  • Laron Z, Pertzelan A, Mannheimer S (1966) Genetic pituitary dwarfism with high serum concentation of growth hormone—a new inborn error of metabolism? Isr J Med Sci 2(2):152–155

    CAS  PubMed  Google Scholar 

  • Leung DW, Spencer SA, Cachianes G, Hammonds RG, Collins C, Henzel WJ, Barnard R, Waters MJ, Wood WI (1987) Growth hormone receptor and serum binding protein: purification, cloning and expression. Nature 330(6148):537–543

    Article  CAS  PubMed  Google Scholar 

  • List EO, Sackmann-Sala L, Berryman DE, Funk K, Kelder B, Gosney ES, Okada S, Ding J, Cruz-Topete D, Kopchick JJ (2011) Endocrine parameters and phenotypes of the growth hormone receptor gene disrupted (GHR−/−) mouse. Endocr Rev 32(3):356–386

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Liu JL, Coschigano KT, Robertson K, Lipsett M, Guo Y, Kopchick JJ, Kumar U, Liu YL (2004) Disruption of growth hormone receptor gene causes diminished pancreatic islet size and increased insulin sensitivity in mice. Am J Physiol Endocrinol Metab 287(3):E405–E413

    Article  CAS  PubMed  Google Scholar 

  • Liu H, Li Y, Wei Q, Liu C, Bolund L, Vajta G, Dou H, Yang W, Xu Y, Luan J et al (2013) Development of transgenic minipigs with expression of antimorphic human cryptochrome 1. PLoS One 8(10):e76098

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Melnik BC, John SM, Schmitz G (2011) Over-stimulation of insulin/IGF-1 signaling by western diet may promote diseases of civilization: lessons learnt from laron syndrome. Nutr Metab 8:41

    Article  CAS  Google Scholar 

  • Mercado M, DaVila N, McLeod JF, Baumann G (1994) Distribution of growth hormone receptor messenger ribonucleic acid containing and lacking exon 3 in human tissues. J Clin Endocrinol Metab 78(3):731–735

    CAS  PubMed  Google Scholar 

  • Milman S, Atzmon G, Huffman DM, Wan J, Crandall JP, Cohen P, Barzilai N (2014) Low insulin-like growth factor-1 level predicts survival in humans with exceptional longevity. Aging Cell 13(4):769–771

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Murakami M, Narazaki M, Hibi M, Yawata H, Yasukawa K, Hamaguchi M, Taga T, Kishimoto T (1991) Critical cytoplasmic region of the interleukin 6 signal transducer gp130 is conserved in the cytokine receptor family. Proc Natl Acad Sci USA 88(24):11349–11353

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Piwien-Pilipuk G, Huo JS, Schwartz J (2002) Growth hormone signal transduction. J Pediatr Endocrinol Metab JPEM 15(6):771–786

    Article  CAS  PubMed  Google Scholar 

  • Rhodes CJ (2005) Type 2 diabetes-a matter of beta-cell life and death? Science 307(5708):380–384

    Article  CAS  PubMed  Google Scholar 

  • Rosenfeld RG, Rosenbloom AL, Guevara-Aguirre J (1994) Growth hormone (GH) insensitivity due to primary GH receptor deficiency. Endocr Rev 15(3):369–390

    Article  CAS  PubMed  Google Scholar 

  • Ross RJ, Esposito N, Shen XY, Von Laue S, Chew SL, Dobson PR, Postel-Vinay MC, Finidori J (1997) A short isoform of the human growth hormone receptor functions as a dominant negative inhibitor of the full-length receptor and generates large amounts of binding protein. Mol Endocrinol 11(3):265–273

    Article  CAS  PubMed  Google Scholar 

  • Rowland JE, Lichanska AM, Kerr LM, White M, d’Aniello EM, Maher SL, Brown R, Teasdale RD, Noakes PG, Waters MJ (2005) In vivo analysis of growth hormone receptor signaling domains and their associated transcripts. Mol Cell Biol 25(1):66–77

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Saveliev A, Everett C, Sharpe T, Webster Z, Festenstein R (2003) DNA triplet repeats mediate heterochromatin-protein-1-sensitive variegated gene silencing. Nature 422(6934):909–913

    Article  CAS  PubMed  Google Scholar 

  • Silbergeld A, Dastot F, Klinger B, Kanety H, Eshet R, Amselem S, Laron Z (1997) Intronic mutation in the growth hormone (GH) receptor gene from a girl with Laron syndrome and extremely high serum GH binding protein: extended phenotypic study in a very large pedigree. J Clin Endocrinol Metab JPEM 10(3):265–274

    CAS  Google Scholar 

  • Stenson PD, Ball EV, Mort M, Phillips AD, Shiel JA, Thomas NS, Abeysinghe S, Krawczak M, Cooper DN (2003) Human gene mutation database (HGMD): 2003 update. Hum Mutat 21(6):577–581

    Article  CAS  PubMed  Google Scholar 

  • Strous GJ, van Kerkhof P, Govers R, Ciechanover A, Schwartz AL (1996) The ubiquitin conjugation system is required for ligand-induced endocytosis and degradation of the growth hormone receptor. EMBO J 15(15):3806–3812

    PubMed Central  CAS  PubMed  Google Scholar 

  • Tiulpakov A, Rubtsov P, Dedov I, Peterkova V, Bezlepkina O, Chrousos GP, Hochberg Z (2005) A novel C-terminal growth hormone receptor (GHR) mutation results in impaired GHR-STAT5 but normal STAT-3 signaling. J Clin Endocrinol Metab 90(1):542–547

    Article  CAS  PubMed  Google Scholar 

  • van Bunderen CC, Oosterwerff MM, van Schoor NM, Deeg DJ, Lips P, Drent ML (2013) Serum IGF1, metabolic syndrome, and incident cardiovascular disease in older people: a population-based study. Eur J Endocrinol 168(3):393–401

    Article  PubMed  Google Scholar 

  • Vijayakumar A, Wu Y, Sun H, Li X, Jeddy Z, Liu C, Schwartz GJ, Yakar S, LeRoith D (2012) Targeted loss of GHR signaling in mouse skeletal muscle protects against high-fat diet-induced metabolic deterioration. Diabetes 61(1):94–103

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Wang X, Souza SC, Kelder B, Cioffi JA, Kopchick JJ (1995) A 40-amino acid segment of the growth hormone receptor cytoplasmic domain is essential for GH-induced tyrosine-phosphorylated cytosolic proteins. J Biol Chem 270(11):6261–6266

    Article  CAS  PubMed  Google Scholar 

  • Williams A, Harker N, Ktistaki E, Veiga-Fernandes H, Roderick K, Tolaini M, Norton T, Williams K, Kioussis D (2008) Position effect variegation and imprinting of transgenes in lymphocytes. Nucleic Acids Res 36(7):2320–2329

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Woods KA, Fraser NC, Postel-Vinay MC, Savage MO, Clark AJ (1996) A homozygous splice site mutation affecting the intracellular domain of the growth hormone (GH) receptor resulting in Laron syndrome with elevated GH-binding protein. J Clin Endocrinol Metab 81(5):1686–1690

    CAS  PubMed  Google Scholar 

  • Zhou Y, Xu BC, Maheshwari HG, He L, Reed M, Lozykowski M, Okada S, Cataldo L, Coschigamo K, Wagner TE et al (1997) A mammalian model for Laron syndrome produced by targeted disruption of the mouse growth hormone receptor/binding protein gene (the Laron mouse). Proc Natl Acad Sci USA 94(24):13215–13220

    Article  PubMed Central  CAS  PubMed  Google Scholar 

Download references

Acknowledgment

We are grateful to all participants of the transgenic group at BGI and the cloning group at BGI Ark Biotechnology (BAB). This work was supported by research grants from China and the National Basic Research Program of China (973 Program 2011CB944201), National Transgenic Project of China (2014ZX0801007B), Special Foundation of Shenzhen Municipal Government of China for the development of Bio-industry (ZYC201105170390A), Shenzhen overseas high level talents innovation Program (KQC201109050086A), Shenzhen Municipal Basic Research Program (JCYJ20120830105157538).

Author information

Author notes

    Authors and Affiliations

    1. BGI-Shenzhen, Shenzhen, 518083, China

      Feida Li, Yong Li, Huan Liu, Xingju Zhang, Chuxin Liu, Kai Tian, Lars Bolund, Huanming Yang & Yutao Du

    2. BGI Ark Biotechnology (BAB), Shenzhen, 518083, China

      Yong Li, Hongwei Dou, Wenxian Yang & Yutao Du

    3. ShenZhen Engineering Laboratory for Genomics-Assisted Animal Breeding, BGI-Shenzhen, 8F Main Building, Beishan Industrial Zone, Yantian District, Shenzhen, 518083, China

      Feida Li, Yong Li, Huan Liu, Xingju Zhang, Chuxin Liu, Kai Tian, Hongwei Dou, Wenxian Yang & Yutao Du

    4. Department of Biomedicine, Aarhus University, Wilhelm Meyers allé 1240, 8000, Aarhus C, Denmark

      Lars Bolund & Nicklas Heine Staunstrup

    Authors
    1. Feida Li
      View author publications

      You can also search for this author in PubMed Google Scholar

    2. Yong Li
      View author publications

      You can also search for this author in PubMed Google Scholar

    3. Huan Liu
      View author publications

      You can also search for this author in PubMed Google Scholar

    4. Xingju Zhang
      View author publications

      You can also search for this author in PubMed Google Scholar

    5. Chuxin Liu
      View author publications

      You can also search for this author in PubMed Google Scholar

    6. Kai Tian
      View author publications

      You can also search for this author in PubMed Google Scholar

    7. Lars Bolund
      View author publications

      You can also search for this author in PubMed Google Scholar

    8. Hongwei Dou
      View author publications

      You can also search for this author in PubMed Google Scholar

    9. Wenxian Yang
      View author publications

      You can also search for this author in PubMed Google Scholar

    10. Huanming Yang
      View author publications

      You can also search for this author in PubMed Google Scholar

    11. Nicklas Heine Staunstrup
      View author publications

      You can also search for this author in PubMed Google Scholar

    12. Yutao Du
      View author publications

      You can also search for this author in PubMed Google Scholar

    Corresponding authors

    Correspondence to Nicklas Heine Staunstrup or Yutao Du.

    Ethics declarations

    Conflict of interest

    There is no conflict of interest and financial disclosure statements are clearly listed.

    Additional information

    Feida Li, Yong Li and Huan Liu have contributed equally to this work .

    Electronic supplementary material

    Below is the link to the electronic supplementary material.

    Supplementary Table 1

    List of primers. The color-code refers to PCR (black) and RT-qPCR (red) primers. The underlined sequences mark the recognition sites for the restriction enzyme NheI and XhoI, respectively (PPTX 44 kb)

    Supplementary Table 2

    Summary of cloning efficiencies obtained with the GHRdm transgenic Wuzhishan fetal donor fibroblasts (PPTX 42 kb)

    Supplementary Table 3

    Skeleton size of a GHRdm transgenic versus a non-transgenic Wuzhishan minipig (PPTX 40 kb)

    Supplementary material 4 (DOCX 114 kb)

    Rights and permissions

    Reprints and permissions

    About this article

    Check for updates. Verify currency and authenticity via CrossMark

    Cite this article

    Li, F., Li, Y., Liu, H. et al. Transgenic Wuzhishan minipigs designed to express a dominant-negative porcine growth hormone receptor display small stature and a perturbed insulin/IGF-1 pathway. Transgenic Res 24, 1029–1042 (2015). https://doi.org/10.1007/s11248-015-9912-6

    Download citation

    • Received:

    • Accepted:

    • Published:

    • Issue Date:

    • DOI: https://doi.org/10.1007/s11248-015-9912-6

    Keywords

    Search

    Navigation