December 2002
Volume 43, Issue 13
Free
ARVO Annual Meeting Abstract  |   December 2002
Evidence for an RPGR-interacting Protein That Is Associated With Centrosomes
Author Affiliations & Notes
  • AF Wright
    MRC Human Genetics Unit Edinburgh United Kingdom
  • B Tulloch
    MRC Human Genetics Unit Edinburgh United Kingdom
  • JW Crabb
    Cleveland Clinic Foundation Cole Eye Institute Cleveland OH
  • X Shu
    MRC Human Genetics Unit Edinburgh United Kingdom
  • A Lennon
    MRC Human Genetics Unit Edinburgh United Kingdom
  • FD C Manson
    MRC Human Genetics Unit Edinburgh United Kingdom
  • M Eckmiller
    Vogt Brain Research Institute Heinrich Heine University of Dusseldorf Dusseldorf Germany
  • R Vervoort
    MRC Human Genetics Unit Edinburgh United Kingdom
  • Footnotes
    Commercial Relationships   A.F. Wright, None; B. Tulloch, None; J.W. Crabb, None; X. Shu, None; A. Lennon, None; F.D.C. Manson, None; M. Eckmiller, None; R. Vervoort, None. Grant Identification: Foundation Fighting Blindness, British Retinitis Pigmentosa Society
Investigative Ophthalmology & Visual Science December 2002, Vol.43, 1383. doi:
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      AF Wright, B Tulloch, JW Crabb, X Shu, A Lennon, FD C Manson, M Eckmiller, R Vervoort; Evidence for an RPGR-interacting Protein That Is Associated With Centrosomes . Invest. Ophthalmol. Vis. Sci. 2002;43(13):1383.

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

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Abstract

Abstract: : Purpose: To characterise the function of the RPGR protein, which is mutated in X-linked retinitis pigmentosa, by searching for interacting proteins. Methods: Protein pull-downs were carried out using immobilised bovine RPGR ORF15 C terminal fragment (C2) and bovine retinal extracts. Bound proteins were fractionated by SDS-PAGE, gel bands excised, digested in situ with trypsin and identified by peptide mass mapping using MALDI-TOF mass spectrometry. Yeast two-hybrid analyses were used to test whether the interaction is direct or indirect. COS cell transfections were carried out and co-localisation investigated using antibodies to an epitope tag fused to the C2 fragment and to the 40 kDa protein. Results: A 40 kDa centrosome-associated protein was identified as a new potential RPGR ORF15 (C2) interacting protein by protein pull-down and peptide mass fingerprinting from bovine retinal extracts. Initial yeast two-hybrid results suggest a direct but weak interaction with RPGR. COS cell transfection of RPGR ORF15 C2 showed co-localisation with the cognate 40 kDa protein using specific antibodies. Conclusion: A 40 kDa centrosome-associated protein shows evidence of specific interaction with the RPGR ORF15 C terminal fragment.

Keywords: 562 retinal degenerations: hereditary • 555 retina: distal(photoreceptors, horizontal cells, bipolar cells) • 526 protein purification and characterization 
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