April 2010
Volume 51, Issue 13
Free
ARVO Annual Meeting Abstract  |   April 2010
Recombinant Myocilin Interacts With the Matricellular Protein SPARC
Author Affiliations & Notes
  • D. Aroca-Aguilar
    Laboratorio de Genética Molecular Humana, Facultad de Medicina, Univ. de Castilla-La Mancha, Albacete, Spain
    Cooperative Research Network on Age-Related Ocular Pathology, Visual and Life Quality, Instituto de Salud Carlos III, Madrid, Spain
  • F. Sánchez-Sánchez
    Laboratorio de Genética Molecular Humana, Facultad de Medicina, Univ. de Castilla-La Mancha, Albacete, Spain
    Cooperative Research Network on Age-Related Ocular Pathology, Visual and Life Quality, Instituto de Salud Carlos III, Madrid, Spain
  • S. Ghosh
    Department of Ophthalmology and Visual Science, Yale University School of Medicine, New Haven, Connecticut
  • M. Coca-Prados
    Department of Ophthalmology and Visual Science, Yale University School of Medicine, New Haven, Connecticut
    Fundación de Investigación Oftalmológica (FIO), Oviedo, Spain
  • J. Escribano
    Laboratorio de Genética Molecular Humana, Facultad de Medicina, Univ. de Castilla-La Mancha, Albacete, Spain
    Cooperative Research Network on Age-Related Ocular Pathology, Visual and Life Quality, Instituto de Salud Carlos III, Madrid, Spain
  • Footnotes
    Commercial Relationships  D. Aroca-Aguilar, None; F. Sánchez-Sánchez, None; S. Ghosh, None; M. Coca-Prados, None; J. Escribano, None.
  • Footnotes
    Support  (Spanish Ministry of Science SAF2008-02228), (Regional Ministry of Health GCS-2006_C/12 and Science PAI-05-002 and PCI08-0036), (I. S. Carlos III RD07/0062/0014), (NIH Grant EY00785),
Investigative Ophthalmology & Visual Science April 2010, Vol.51, 6093. doi:
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    • Get Citation

      D. Aroca-Aguilar, F. Sánchez-Sánchez, S. Ghosh, M. Coca-Prados, J. Escribano; Recombinant Myocilin Interacts With the Matricellular Protein SPARC. Invest. Ophthalmol. Vis. Sci. 2010;51(13):6093.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Purpose: : Myocilin is an extracellular glycoprotein with unknown function which is associated with different types of glaucoma. We have recently reported a proteolytical processing of myocilin by Calpain II within the linker region of the protein releasing the C-terminal olfactomedin domain. We have described also that myocilin interacts with the C-terminal region of hevin, a secretory glycoprotein belonging to the SPARC family of matricellular proteins. The main objective of this study is to investigate the interaction of recombinant myocilin with SPARC.

Methods: : Analysis of protein-protein interactions were performed using the yeast two-hybrid system and solid-phase binding assays with Ni-chelating HPLC purified recombinant proteins. Coexpression of myocilin, SPARC and hevin in ocular tissues was confirmed by immunoflorescence microscopy, western blot and array-based gene profiling.

Results: : Yeast two-hybrid analyses showed that myocilin interacted with the highly conserved C-terminal extracellular calcium binding (EC) domain within SPARC and hevin. Solid-phase binding assays showed that full-length myocilin interacted with higher affinity with SPARC and its EC domain than the myocilin C-terminal fragment.

Conclusions: : Recombinant myocilin and SPARC interact through their C-terminal domains. Our data suggest that the proteolytic processing of myocilin modulates this interaction supporting a functional role for this proteolytic clevage.

Keywords: proteins encoded by disease genes • extracellular matrix 
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