The coordinated migration of bilateral cardiomyocytes and the formation of the cardiac cone are essential for heart tube formation. We investigated gene regulatory mechanisms involved in myocardial migration, and regulation of the timing of cardiac cone formation in zebrafish embryos. Through screening of zebrafish treated with ethylnitrosourea, we isolated a mutant with a hypomorphic allele of mil (s1pr2)/edg5, called s1pr2as10 (as10). Mutant embryos with this allele expressed less mil/edg5 mRNA and exhibited cardia bifida prior to 28 hours post-fertilization. Although the bilateral hearts of the mutants gradually fused together, the resulting formation of two atria and one tightly-packed ventricle failed to support normal blood circulation. Interestingly, cardia bifida of s1pr2as10 embryos could be rescued and normal circulation could be restored by incubating the embryos at low temperature (22.5°C). Rescue was also observed in gata5 and bon cardia bifida morphants raised at 22.5°C. The use of DNA microarrays, digital gene expression analyses, loss-of-function, as well as mRNA and protein rescue experiments, revealed that low temperature mitigates cardia bifida by regulating the expression of genes encoding components of the extracellular matrix (fibronectin 1, tenascin-c, tenascin-w). Furthermore, the addition of N-acetyl cysteine (NAC), a reactive oxygen species (ROS) scavenger, significantly decreased the effect of low temperature on mitigating cardia bifida in s1pr2as10 embryos. Our study reveals that temperature coordinates the development of the heart tube and somitogenesis, and that extracellular matrix genes (fibronectin 1, tenascin-c and tenascin-w) are involved.
References
[1]
Tu S, Chi NC (2012) Zebrafish models in cardiac development and congenital heart birth defects. Differentiation 84: 4–16.
[2]
Miura GI, Yelon D (2011) A guide to analysis of cardiac phenotypes in the zebrafish embryo. Methods Cell Biol 101: 161–180.
[3]
Bakkers J (2011) Zebrafish as a model to study cardiac development and human cardiac disease. Cardiovasc Res 91: 279–288.
[4]
Yelon D, Ticho B, Halpern ME, Ruvinsky I, Ho RK, et al. (2000) The bHLH transcription factor hand2 plays parallel roles in zebrafish heart and pectoral fin development. Development 127: 2573–2582.
[5]
Reiter JF, Alexander J, Rodaway A, Yelon D, Patient R, et al. (1999) Gata5 is required for the development of the heart and endoderm in zebrafish. Genes Dev 13: 2983–2995.
[6]
Alexander J, Rothenberg M, Henry GL, Stainier DY (1999) casanova plays an early and essential role in endoderm formation in zebrafish. Dev Biol 215: 343–357.
[7]
Kikuchi Y, Trinh LA, Reiter JF, Alexander J, Yelon D, et al. (2000) The zebrafish bonnie and clyde gene encodes a Mix family homeodomain protein that regulates the generation of endodermal precursors. Genes Dev 14: 1279–1289.
[8]
Trinh LA, Stainier DY (2004) Fibronectin regulates epithelial organization during myocardial migration in zebrafish. Dev Cell 6: 371–382.
[9]
Kawahara A, Nishi T, Hisano Y, Fukui H, Yamaguchi A, et al. (2009) The sphingolipid transporter spns2 functions in migration of zebrafish myocardial precursors. Science 323: 524–527.
[10]
Kupperman E, An S, Osborne N, Waldron S, Stainier DY (2000) A sphingosine-1-phosphate receptor regulates cell migration during vertebrate heart development. Nature 406: 192–195.
[11]
Osborne N, Brand-Arzamendi K, Ober EA, Jin SW, Verkade H, et al. (2008) The spinster homolog, two of hearts, is required for sphingosine 1-phosphate signaling in zebrafish. Curr Biol 18: 1882–1888.
[12]
George EL, Georges-Labouesse EN, Patel-King RS, Rayburn H, Hynes RO (1993) Defects in mesoderm, neural tube and vascular development in mouse embryos lacking fibronectin. Development 119: 1079–1091.
[13]
Linask KK, Lash JW (1988) A role for fibronectin in the migration of avian precardiac cells. I. Dose-dependent effects of fibronectin antibody. Dev Biol 129: 315–323.
[14]
Garavito-Aguilar ZV, Riley HE, Yelon D (2010) Hand2 ensures an appropriate environment for cardiac fusion by limiting Fibronectin function. Development 137: 3215–3220.
[15]
Matsui T, Raya A, Callol-Massot C, Kawakami Y, Oishi I, et al. (2007) miles-apart-Mediated regulation of cell-fibronectin interaction and myocardial migration in zebrafish. Nat Clin Pract Cardiovasc Med 4 Suppl 1S77–82.
[16]
Shentu H, Wen HJ, Her GM, Huang CJ, Wu JL, et al. (2003) Proximal upstream region of zebrafish bone morphogenetic protein 4 promoter directs heart expression of green fluorescent protein. Genesis 37: 103–112.
[17]
Kimmel CB, Ballard WW, Kimmel SR, Ullmann B, Schilling TF (1995) Stages of embryonic development of the zebrafish. Dev Dyn 203: 253–310.
[18]
Solnica-Krezel L, Schier AF, Driever W (1994) Efficient recovery of ENU-induced mutations from the zebrafish germline. Genetics 136: 1401–1420.
[19]
Trinh LA, Meyer D, Stainier DY (2003) The Mix family homeodomain gene bonnie and clyde functions with other components of the Nodal signaling pathway to regulate neural patterning in zebrafish. Development 130: 4989–4998.
[20]
Serluca FC (2008) Development of the proepicardial organ in the zebrafish. Dev Biol 315: 18–27.
[21]
Trinh LA, Yelon D, Stainier DY (2005) Hand2 regulates epithelial formation during myocardial diferentiation. Curr Biol 15: 441–446.
[22]
Schweitzer J, Becker T, Lefebvre J, Granato M, Schachner M, et al. (2005) Tenascin-C is involved in motor axon outgrowth in the trunk of developing zebrafish. Dev Dyn 234: 550–566.
[23]
Koshida S, Kishimoto Y, Ustumi H, Shimizu T, Furutani-Seiki M, et al. (2005) Integrinalpha5-dependent fibronectin accumulation for maintenance of somite boundaries in zebrafish embryos. Dev Cell 8: 587–598.
[24]
Zheng Q, Wang XJ (2008) GOEAST: a web-based software toolkit for Gene Ontology enrichment analysis. Nucleic Acids Res 36: W358–363.
[25]
Huang da W, Sherman BT, Lempicki RA (2009) Bioinformatics enrichment tools: paths toward the comprehensive functional analysis of large gene lists. Nucleic Acids Res 37: 1–13.
[26]
Huang da W, Sherman BT, Zheng X, Yang J, Imamichi T, et al.. (2009) Extracting biological meaning from large gene lists with DAVID. Curr Protoc Bioinformatics Chapter 13: Unit 13 11.
[27]
Schroter C, Herrgen L, Cardona A, Brouhard GJ, Feldman B, et al. (2008) Dynamics of zebrafish somitogenesis. Dev Dyn 237: 545–553.
[28]
Reiter JF, Kikuchi Y, Stainier DY (2001) Multiple roles for Gata5 in zebrafish endoderm formation. Development 128: 125–135.
Lahiri K, Vallone D, Gondi SB, Santoriello C, Dickmeis T, et al. (2005) Temperature regulates transcription in the zebrafish circadian clock. PLoS Biol 3: e351.
[31]
Tseng YC, Chen RD, Lucassen M, Schmidt MM, Dringen R, et al. (2011) Exploring uncoupling proteins and antioxidant mechanisms under acute cold exposure in brains of fish. PLoS One 6: e18180.
[32]
Kumar S, Seqqat R, Chigurupati S, Kumar R, Baker KM, et al. (2011) Inhibition of nuclear factor kappaB regresses cardiac hypertrophy by modulating the expression of extracellular matrix and adhesion molecules. Free Radic Biol Med 50: 206–215.
[33]
Chiquet-Ehrismann R, Tucker RP (2011) Tenascins and the importance of adhesion modulation. Cold Spring Harb Perspect Biol 3.
[34]
Jones FS, Jones PL (2000) The tenascin family of ECM glycoproteins: structure, function, and regulation during embryonic development and tissue remodeling. Dev Dyn 218: 235–259.
[35]
Trebaul A, Chan EK, Midwood KS (2007) Regulation of fibroblast migration by tenascin-C. Biochem Soc Trans 35: 695–697.
[36]
Ishigaki T, Imanaka-Yoshida K, Shimojo N, Matsushima S, Taki W, et al. (2011) Tenascin-C enhances crosstalk signaling of integrin alphavbeta3/PDGFR-beta complex by SRC recruitment promoting PDGF-induced proliferation and migration in smooth muscle cells. J Cell Physiol 226: 2617–2624.
[37]
Zagzag D, Shiff B, Jallo GI, Greco MA, Blanco C, et al. (2002) Tenascin-C promotes microvascular cell migration and phosphorylation of focal adhesion kinase. Cancer Res 62: 2660–2668.
[38]
Huang JY, Cheng YJ, Lin YP, Lin HC, Su CC, et al. (2010) Extracellular matrix of glioblastoma inhibits polarization and transmigration of T cells: the role of tenascin-C in immune suppression. J Immunol 185: 1450–1459.
[39]
Brellier F, Martina E, Chiquet M, Ferralli J, van der Heyden M, et al. (2012) The adhesion modulating properties of tenascin-W. Int J Biol Sci 8: 187–194.
[40]
Imanaka-Yoshida K, Hiroe M, Yoshida T (2004) Interaction between cell and extracellular matrix in heart disease: multiple roles of tenascin-C in tissue remodeling. Histol Histopathol 19: 517–525.
[41]
Yamamoto K, Dang QN, Kennedy SP, Osathanondh R, Kelly RA, et al. (1999) Induction of tenascin-C in cardiac myocytes by mechanical deformation. Role of reactive oxygen species. J Biol Chem 274: 21840–21846.
[42]
Davis WL, Crawford LA, Cooper OJ, Farmer GR, Thomas D, et al. (1990) Generation of radical oxygen species by neural crest cells treated in vitro with isotretinoin and 4-oxo-isotretinoin. J Craniofac Genet Dev Biol 10: 295–310.
[43]
Lum H, Roebuck KA (2001) Oxidant stress and endothelial cell dysfunction. Am J Physiol Cell Physiol 280: C719–741.
[44]
Zhang X, Azhar G, Nagano K, Wei JY (2001) Differential vulnerability to oxidative stress in rat cardiac myocytes versus fibroblasts. J Am Coll Cardiol 38: 2055–2062.
Wang X, Michaelis EK (2010) Selective neuronal vulnerability to oxidative stress in the brain. Front Aging Neurosci 2: 12.
[47]
Aw TY (1999) Molecular and cellular responses to oxidative stress and changes in oxidation-reduction imbalance in the intestine. Am J Clin Nutr 70: 557–565.
[48]
Iyoda T, Fukai F (2012) Modulation of Tumor Cell Survival, Proliferation, and Differentiation by the Peptide Derived from Tenascin-C: Implication of beta1-Integrin Activation. Int J Cell Biol 2012: 647594.
[49]
Ridley AJ, Schwartz MA, Burridge K, Firtel RA, Ginsberg MH, et al. (2003) Cell migration: integrating signals from front to back. Science 302: 1704–1709.
[50]
Huttenlocher A, Horwitz AR (2011) Integrins in cell migration. Cold Spring Harb Perspect Biol 3: a005074.
[51]
Weber P, Montag D, Schachner M, Bernhardt RR (1998) Zebrafish tenascin-W, a new member of the tenascin family. J Neurobiol 35: 1–16.
