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L. W. Reneker, H. Chen; Calpain and Caspase-12 Activation Mediates Apoptosis in Transgenic Mouse Lens Expressing a Dominant-Negative Mutant of FGFR. Invest. Ophthalmol. Vis. Sci. 2010;51(13):2617.
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© ARVO (1962-2015); The Authors (2016-present)
Expressing a dominant negative (DN) mutant of FGFR, which is a truncated form of FGFR1, in the transgenic mouse lens can induce cell death in the lens fiber cells. Previously we demonstrated that DN-FGFR-induced cell death is independent of the classical mitochondrial apoptosis mechanism. The alternative cell death mechanism is investigated in this study.
Water-soluble proteins were prepared from newborn wild type (WT) and DN-FGFR mouse lenses. Calpain activation was evaluated by the presence of cleaved crystalline fragments. Cleaved (active) caspase-12 in the lens homogenate was detected by western blot. Caspase-12 localization in mouse lens was demonstrated by immunofluorescence.
alpha- and beta-crystallin breakdown products were detected by western blot in the DN-FGFR lenses. The same crystallin proteolytic patterns were seen in the WT lens proteins after incubation with 1 mM calcium at 37°C for one hour. These results suggested that abnormal activation of calpain occurred in the DN-FGFR lens. A low level of cleaved caspase-12 was present in the WT lens. In comparison, the level was significantly increased in the DN-FGFR transgenic lens. Immunostaining showed that, in the WT lens, caspase-12 was found in the lens epithelial cells. During fiber cell differentiation, caspase-12 was localized to the nuclei of the fiber cells. In contrast to the WT lens, caspase-12 immunofluorescence in the DN-FGFR lens did not co-localize with the cell nuclear staining, suggesting that caspase-12 released from the cell nuclei can be cleaved (activated) by the calpain in the cytoplasm in the presence of calcium.
We have identified a new calpain activated caspase-12 mediated apoptosis pathway in the lens fiber cells. We propose that accumulation of folding-incompetent DN-FGFR proteins in the lens fiber cells can induce ER stress, disrupt calcium homeostasis, and result in the activation of calpain and caspase-12 which contributes to the mechanism of apoptosis.
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