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A.J. Zandy, S. Bassnett; In vivo Inhibition of the Proteasome by Lactacystin Perturbs Fiber Cell Differentiation . Invest. Ophthalmol. Vis. Sci. 2006;47(13):1994.
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© ARVO (1962-2015); The Authors (2016-present)
During lens fiber cell differentiation, all intracellular organelles including nuclei, mitochondria, endoplasmic reticulum and Golgi are degraded. This provides a transparent fiber cell cytoplasm through which light can pass. The proteolytic mechanism behind organelle breakdown is not understood. Here we examine whether pharmacological inhibition of the proteasome disrupts the organelle breakdown process.
An intravitreal injection of the proteasome inhibitor, lactacystin, was made into chicken embryos of day 10 (E10) of development. The efficacy of the drug treatment was measured on whole or regionally–dissected lenses 3, 24 and 48 hours post injection using a proteasome activity assay.
The trypsin–like activity of the proteasome was inhibited ∼60% in whole lenses 3 and 24 hours post injection but fully recovered by 48 hours. Inhibition of the proteasome extends through the entire fiber cell mass. At 24 hours post injection, there is a 6–fold increase in ubiquitinated proteins in the lens cortex compared to vehicle controls and a lesser increase in the lens nucleus. At this time point, the lens also contained a dense cortical opacity. A mitochondrial marker, succinate–ubiquinone reductase, was detected by western blot in regionally–dissected lenses from control and lactacystin–treated eyes. While this marker was degraded in the core of vehicle–injected lenses 24 hours post–injection, it persisted in the core of lactacystin–treated lenses.
Inhibition of the proteasome in the lens causes an accumulation of ubiquitinated proteins and a cortical opacity. Additionally, an organelle component that is normally broken down in the central fibers is retained in lactacystin injected eyes. These data suggest a role for the proteasome during organelle breakdown.
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