Skip to main content

Advertisement

Log in

Immunolocalization of myosin Va in the developing nervous system of embryonic chicks

  • Original Article
  • Published:
Anatomy and Embryology Aims and scope Submit manuscript

Abstract

Myosins are molecular motors associated with the actin cytoskeleton that participate in the mechanisms of cellular motility. During the development of the nervous system, migration of nerve cells to specific sites, extension of growth cones, and axonal transport are dramatic manifestations of cellular motility. We demonstrate, via immunoblots, the expression of myosin Va during early stages of embryonic development in chicks, extending from the blastocyst period to the beginning of the fetal period. The expression of myosin Va in specific regions and cellular structures of the nervous system during these early stages was determined by immunocytochemistry using a polyclonal antibody. Whole mounts of chick embryos at 24–30-h stages showed intense immunoreactivity of the neural tube in formation along its full extent. Cross-sections at these stages of development showed strong labeling in neuroepithelial cells at the basal and apical regions of the neural tube wall. Embryos at more advanced periods of development (48h and 72 h) showed distinctive immunolabeling of neuroepithelial cells, neuroblasts and their cytoplasmic extensions in the mantle layer of the stratified neural tube wall, and neuroblasts and their cytoplasmic extensions in the internal wall of the optic cup, as well as a striking labeling of cells in the apparent nuclei of cranial nerves and budding fibers. These immunolocalization studies indicate temporal and site-specific expression of myosin Va during chick embryo development, suggesting that myosin Va expression is related to recruitment for specific cellular tasks.

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

Access this article

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

Instant access to the full article PDF.

Fig. 1
Fig. 2
Fig. 3A, B
Fig. 4A–D
Fig. 5A–C
Fig. 6

Similar content being viewed by others

References

  • Alvarez IS, Schoenwolf GC (1992) Expansion of surface epithelium provides the major extrinsic force for bending of the neural plate. J Exp Zool 261:340–348

    CAS  PubMed  Google Scholar 

  • Baas PW, Ahmad FJ (2001) Force generation by cytoskeletal motor proteins as a regulator of axonal elongation and retraction. Trends Cell Biol 11:244–249

    Article  CAS  PubMed  Google Scholar 

  • Berg JS, Powell BC, Cheney RE (2001) A millennial myosin census. Mol Biol Cell 12:780–794

    CAS  PubMed  Google Scholar 

  • Bloom G.S, Endow SA. 1995. Motor proteins 1: kinesins. Protein Profile 2:1109–1138

    Google Scholar 

  • Casaletti L., Tauhata SBF, Moreira JE, Larson RE (2003) Myosin-Va proteolysis by Ca2+/calpain in depolarized nerve endings from rat brain. Biochem Biophys Res Commun 308:159–164

    Article  CAS  PubMed  Google Scholar 

  • Colas JF, Schoenwolf GC (2001) Towards a cellular and molecular understanding of neurulation. Dev Dyn 221:117–145

    Article  CAS  PubMed  Google Scholar 

  • Costa MC, Mani F, Santoro WJr, Espreafico EM, Larson RE (1999) Brain myosin-V, a calmodulin-carrying myosin, binds to calmodulin-dependent protein kinase II and activates its kinase activity. J Biol Chem 274:15811–15819

    Article  CAS  PubMed  Google Scholar 

  • Dekker-Ohno K, Hayasaka S, Takagishi Y, Oda S, Wakasugi N, Mikoshiba K, Inouye M, Yamamura H (1996) Endoplasmic reticulum is missing in dendritic spines of Purkinje cells of the ataxic mutant rat. Brain Res 714:226–230

    Article  CAS  PubMed  Google Scholar 

  • Espindola FS, Espreafico EM, Coelho MV, Martins AR, Costa FRC, Mooseker MS, Larson RE (1992) Biochemical and immunological characterization of P190-calmodulin complex from vertebrate brain—a novel calmodulin-binding myosin. J Cell Biol 118:359–368

    Article  CAS  PubMed  Google Scholar 

  • Espreafico EM, Cheney RE, Matteoli M, Nascimento AA, De Camilli PV, Larson RE, Mooseker MS (1992) Primary structure and cellular localization of chicken brain myosin-V (p190), an unconventional myosin with calmodulin light chains. J Cell Biol 119:1541–1557

    Article  CAS  PubMed  Google Scholar 

  • Evans LL, Lee AJ, Bridgman PC, Mooseker MS (1998) Vesicle-associated brain myosin-V can be activated to catalyze actin-based transport. J Cell Sci 111:2055–2066

    CAS  PubMed  Google Scholar 

  • Futaki S, Takagishi Y, Hayashi Y, Ohmori S, Kanou Y, Inouye M, Oda S, Seo H, Iwaikawa Y, Murata Y (2000) Identification of a novel myosin-Va mutation in an ataxic mutant rat, dilute-opisthotonus. Mamm Genome 11:649–655

    Article  CAS  PubMed  Google Scholar 

  • Griscelli C, Durandy A, Guygrand D, DaGuillard F, Herzog C, Pruniers M (1978) Syndrome associating partial albinism and immunodeficiency. Am J Med 65:691–702

    Article  CAS  PubMed  Google Scholar 

  • Hamburger V, Hamilton HL (1951) A series of normal stages in the development of the chick embryo. J Morphol 88:49–92

    Google Scholar 

  • Hodge T, Cope MJTV (2000) A myosin family tree. J Cell Sci 113:3353–3354

    CAS  PubMed  Google Scholar 

  • Laemmli UK, Favre M (1973) Maturation of the head of bacteriophage T4. I. DNA packaging events. J Mol Biol 80:575–599

    CAS  PubMed  Google Scholar 

  • Langford GM, Molyneaux BJ (1998) Myosin V in the brain: mutations lead to neurological defects. Brain Res Rev 28:1–8

    Article  CAS  Google Scholar 

  • Larson RE, Ferro JA, Queiroz EA (1986) Isolation and purification of actomyosin ATPase from mammalian brain. J Neurosci Methods 16:47–58

    Article  CAS  PubMed  Google Scholar 

  • Lee HY, Kosciuk MC, Nagele RG, Roisen FJ (1983) Studies on the mechanisms of neurulation in the chick: possible involvement of myosin in elevation of neural folds. J Exp Zool 225:449–457

    CAS  PubMed  Google Scholar 

  • Lin CH, Espreafico EM, Mooseker MS, Forscher P (1996) Myosin drives retrograde F-actin flow in neuronal growth cones. Neurology 16:769–782

    Article  CAS  Google Scholar 

  • Mani F, Espreafico EM, Larson RE (1994) Myosin-V is present in synaptosomes from rat cerebral cortex. Braz J Med Biol Res 27:2639–2643

    CAS  PubMed  Google Scholar 

  • Martins AR, Dias MM, Vasconcelos TM, Caldo H, Costa MCR, Chimelli L, Larson RE (1999) Microwave-stimulated recovery of myosin-V immunoreactivity from formalin-fixed, paraffin-embedded human CNS. J Neurosci Methods 92:25–29

    Article  CAS  PubMed  Google Scholar 

  • McLean IW, Nakane PD (1974) Periodate-lysine-paraformaldehyde fixative: a new fixative for immunoelectron microscopy. J Histochem Cytochem 22:1077–1083

    CAS  PubMed  Google Scholar 

  • Mercer JA, Seperack PK, Strobel MC, Copeland NG, Jenkins NA (1991) Novel myosin heavy chain encoded by murine dilute coat colour locus [published erratum appears in Nature 1991, 352: 547]. Nature 349:709–713

    Article  CAS  PubMed  Google Scholar 

  • Miller KE, Sheetz MP (2000) Characterization of myosin V binding to brain vesicles. J Biol Chem 275:2598–606

    Article  CAS  PubMed  Google Scholar 

  • Ohyama A, Komiya Y, Igarashi M (2001) Globular tail of myosin-V is bound to VAMP/synaptobrevin. Biochem Biophys Res Commun 280:988–991

    Article  CAS  PubMed  Google Scholar 

  • Olmsted JB (1986) Analysis of cytoskeletal structures using blot-purified monospecific antibodies. Methods Enzymol 134:467–472

    Article  CAS  PubMed  Google Scholar 

  • Pasteral E, Ersoy F, Yalman N, Wulffraat N, Grillo E, Ozkinay F, Tezcan I, Gedikoglu G, Philippe N, Fisher A, Saint Basile G (2000) Two genes are responsible for Griscelli syndrome at the same 15q21 locus. Genomics 63:299–306

    Article  PubMed  Google Scholar 

  • Prekeris R, Terrian DM (1997) Brain myosin V is a synaptic vesicle-associated motor protein: evidence for a Ca2+-dependent interaction with the synaptobrevin-synaptophysin complex. J Cell Biol 137:1589–1601

    Article  CAS  PubMed  Google Scholar 

  • Provance DW, Mercer JA (1999) Myosin-V: head to tail. Cell Mol Life Sci 56:233–242

    Article  CAS  PubMed  Google Scholar 

  • Reck-Peterson SL, Provance DW, Mooseker MS, Mercer JA (2000) Class V myosins. Biochim Biophys Acta 1496:36–51

    Article  CAS  PubMed  Google Scholar 

  • Rosé SD, Lejen T, Casaletti L, Larson RE, Pene TD, Trifaró JM (2003) Myosin Va, a chromaffin vesicle molecular motor involved in secretion. J Neurochem 85:287–298

    PubMed  Google Scholar 

  • Sadler TW, Greenberg D, Coughlin P, Lessard JL (1982) Actin distribution patterns in the mouse neural tube during neurulation. Science 215:172–174

    CAS  PubMed  Google Scholar 

  • Schoenwolf GC, Powers ML (1987) Shaping of the chick neuroepithelium during primary and secondary neurulation: role of cell elongation. Anat Rec 218:182–195

    CAS  PubMed  Google Scholar 

  • Silvers WK (1979) Dilute and Leaden, the p-locus, ruby-eye, and ruby-eye-2. In: Silvers WK (ed) Coat colors of mice: a model for mammalian gene action and interaction. Springer, Berlin, Heidelberg New York, pp 83–89

  • Smith PK, Krohn RI, Hermanson GT, Mallia AK, Gartner FH, Provenzano MD, Fujimoto EK, Goeke NM, Olson BJ, Klenk DC (1985) Measurement of protein using bicinchoninic acid. Anal Biochem 150:76–85

    CAS  PubMed  Google Scholar 

  • Supp DM, Potter SS, Brueckner M (2000) Molecular motors: the driving force behind mammalian left-right development. Trends Cell Biol 10:41–45

    Article  CAS  PubMed  Google Scholar 

  • Suter DM, Espindola FS, Lin CH, Forscher P, Mooseker MS (2000) Localization of unconventional myosins V and VI in neuronal growth cones. J Neurobiol 42:370–382

    Article  CAS  PubMed  Google Scholar 

  • Tabb JS, Molyneaux BJ, Cohen DL, Kuznetsov SA, Langford GM (1998) Transport of ER vesicles on actin filaments in neurons by myosin V. J Cell Sci 111:3221–3234

    CAS  PubMed  Google Scholar 

  • TakagishiY, Oda S, Hayasaka S, Dekker Ohno K, Shikata T, Inouye M, Yamamura H (1996) The dilute-lethal (dl) gene attacks a Ca2+ store in the dendritic spine of Purkinje cells in mice. Neurosci Lett 215:169–172

    PubMed  Google Scholar 

  • Tilelli CQ, Martins AR, Larson RE, Garcia-Cairasco N (2003) Immunohistochemical localization of myosin Va in adult rat brain. J Neurosci 121:573–586

    Article  CAS  Google Scholar 

  • Towbin H, Staehelin T, Gordon J (1979) Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets—procedure and some applications. Proc Natl Acad Sci USA 76:4350–4354

    CAS  PubMed  Google Scholar 

  • Yin H, Pruyne D, Huffaker TC, Bretscher A (2000) Myosin V orientates the mitotic spindle in yeast. Nature 406:1013–1015

    Article  CAS  PubMed  Google Scholar 

  • Young PE, Pesacreta TC, Kiehart DP (1991) Dynamic changes in the distribution of cytoplasmic myosin during Drosophila embryogenesis. Development 111:1–14

    CAS  PubMed  Google Scholar 

Download references

Acknowledgements

We thank Domingus E. Pitta, Silvas Regina Andrade, José Eduardo Bozano, and Luis Carlos de Oliveira for expert technical assistance. Alexandre de Azevedo was a predoctoral student with financial support from the Coordenação de Aperfeiçoamento de Pessoal de Nivel Superior (CAPES).

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Alexandre Azevedo.

Additional information

Grant sponsor: Fundação de Amparo à Pesquisa do Estado de São Paulo; grant number 99/3951-7 to R.E. Larson. Grant sponsor: Fundação para o Desenvolvimento da Universidade Estadual Paulista; grant number DFP-014 to A. de Azevedo

Rights and permissions

Reprints and permissions

About this article

Cite this article

Azevedo, A., Lunardi, L.O. & Larson, R.E. Immunolocalization of myosin Va in the developing nervous system of embryonic chicks. Anat Embryol 208, 395–402 (2004). https://doi.org/10.1007/s00429-004-0405-2

Download citation

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00429-004-0405-2

Keywords

Navigation