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Mingxing Wu, Xinyu Zhang, Qingning Bian, Allen Taylor, Jack J. Liang, Linlin Ding, Joseph Horwitz, Fu Shang; Oligomerization with wt αA- and αB-Crystallins Reduces Proteasome-Mediated Degradation of C-Terminally Truncated αA-Crystallin. Invest. Ophthalmol. Vis. Sci. 2012;53(6):2541-2550. doi: 10.1167/iovs.11-9147.
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We previously demonstrated that the ubiquitin-proteasome pathway (UPP) is a general protein quality control system that selectively degrades damaged or abnormal lens proteins, including C-terminally truncated αA-crystallin. The objective of this work was to determine the effects of wt αA- and αB-crystallins on the degradation of C-terminally truncated αA-crystallin (αA1–162) and vice versa.
Recombinant wt αA, αB, and αA1–162 were expressed in Escherichia coli and purified to homogeneity by chromatography. Subunit exchange and oligomerization were detected by fluorescence resonance energy transfer (FRET), multiangle-light scattering and coprecipitation assays. Protein substrates were labeled with 125I and lens epithelial cell lysates were used as the source of the UPP for degradation assays.
FRET, multiangle light scattering, and coprecipitation assays showed that αA1–162 exchanged subunits with wt αA- or wt αB- crystallin to form hetero-oligomers. αA1–162 was more susceptible than wt αA-crystallin to degradation by the UPP. When mixed with wt αA-crystallin at 1:1 or 1:4 (αA1–162 : wt) ratios to form hetero-oligomers, the degradation of αA1–162 was significantly decreased. Conversely, formation of hetero-oligomers with αA1–162 enhanced the degradation of wt αA-crystallin. The presence of αA1–162, but not wt αA-crystallin, decreased the degradation of wt αB-crystallin.
αA1–162 forms hetero-oligomers with wt αA- and αB-crystallins. Oligomerization with wt αA- or αB-crystallins reduces the susceptibility of αA1–162 to degradation by the UPP. In addition, the presence of αA1–162 in the hetero-oligomers also affects the degradation of wt αA- and αB-crystallins.
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