May 2008
Volume 49, Issue 13
Free
ARVO Annual Meeting Abstract  |   May 2008
Calpain-1 Processing Differences in Normal and Diseased Ocular Tissues
Author Affiliations & Notes
  • W.-H. Lee
    Ophthalmology, Bascom Palmer Eye Institute/University of Miami, Miami, Florida
  • A. Junk
    Ophthalmology, Bascom Palmer Eye Institute/University of Miami, Miami, Florida
  • B. Govindarajan
    Chemistry, Case Western Reserve University, Cleveland, Ohio
  • J. Laird
    Chemistry, Case Western Reserve University, Cleveland, Ohio
  • R. G. Salomon
    Chemistry, Case Western Reserve University, Cleveland, Ohio
  • S. K. Bhattacharya
    Ophthalmology, Bascom Palmer Eye Institute/University of Miami, Miami, Florida
  • Footnotes
    Commercial Relationships  W. Lee, None; A. Junk, None; B. Govindarajan, None; J. Laird, None; R.G. Salomon, None; S.K. Bhattacharya, None.
  • Footnotes
    Support  Thomas R. Lee Award for National Glaucoma Research Program of American Health Assistance Foundation; NIH grants EY015266, GM021249; P30 EY014801; Research to Prevent Blindness; and RPB CDA (SKB).
Investigative Ophthalmology & Visual Science May 2008, Vol.49, 675. doi:
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    • Get Citation

      W.-H. Lee, A. Junk, B. Govindarajan, J. Laird, R. G. Salomon, S. K. Bhattacharya; Calpain-1 Processing Differences in Normal and Diseased Ocular Tissues. Invest. Ophthalmol. Vis. Sci. 2008;49(13):675.

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

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Abstract

Purpose: : To determine whether in vitro modification of cysteine proteases and calpains in particular with lipid adduct isolevuglandin E2 (Iso[4]LGE2 ) affects its ubiquitination and degradation by cellular proteasome machinery in the glaucomatous trabecular meshwork (TM) in contrast to controls. Whether there exists expression differences for calpain-1 in TM and the optic nerve.

Methods: : Enzymatic assays for cysteine proteases and calpains were made using standard substrates and inhibitors and TM tissue extracts. Western analyses were performed on human TM extracts (from 35 donors) with antibodies to Iso[4]LGE2, ubiquitin and proteasome. Calpain-1 was modified in vitro by Iso[4]LGE2. Plugged gel electrophoresis and dynamic light scattering experiments were performed to analyze protein aggregates. The translational differences were probed using Northern Blot and other published protocols.

Results: : Lipid peroxidation product modification of calpain-1 was observed in the human glaucomatous TM (but not in the optic nerve). Modified calpain-1 was conjugated and loaded onto the 26S proteosomal machinery resulting in the impairment of the later. In contrast to the TM, elevated levels of calpain-1occur in the glaucomatous optic nerve and is due to translational control.

Conclusions: : Calpains are modified by lipid peroxidation products in vivo. In vitro Iso[4]LGE2modification of calpain-1 makes it resistant to protease digestion and impairs proteasome. Calpain-1 overexpression occurs in the optic nerve (and controlled at the level of translation) but not in the TM.

Keywords: protein modifications-post translational • proteomics • outflow: trabecular meshwork 
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