Abstract
Desmin-related myopathy and cataract are both caused by the R120G mutation in alphaB-crystallin. Desmin-related myopathy is one of several diseases characterized by the coaggregation of intermediate filaments with alphaB-crystallin, and it identifies intermediate filaments as important physiological substrates for alphaB-crystallin. Using recombinant human alphaB-crystallin, the effects of the disease-causing mutation R120G upon the structure and the chaperone activities of alphaB-crystallin are reported. The secondary, tertiary, and quaternary structural features of alphaB-crystallin are all altered by the mutation as deduced by near- and far-UV circular dichroism spectroscopy, size exclusion chromatography, and chymotryptic digestion assays. The R120G alphaB-crystallin is also less stable than wild type alphaB-crystallin to heat-induced denaturation. These structural changes coincide with a significant reduction in the in vitro chaperone activity of the mutant alphaB-crystallin protein, as assessed by temperature-induced protein aggregation assays. The mutation also significantly altered the interaction of alphaB-crystallin with intermediate filaments. It abolished the ability of alphaB-crystallin to prevent those filament-filament interactions required to induce gel formation while increasing alphaB-crystallin binding to assembled intermediate filaments. These activities are closely correlated to the observed disease pathologies characterized by filament aggregation accompanied by alphaB-crystallin binding. These studies provide important insight into the mechanism of alphaB-crystallin-induced aggregation of intermediate filaments that causes disease.
Highlights
Desmin-related myopathy (DRM)1 can be caused by mutations either in the intermediate filament protein desmin, or in the small heat shock protein ␣B-crystallin
The R120G mutation in ␣B-crystallin affects a highly conserved residue among the whole sHSP family [44, 45]. It is important for the function of ␣A- and ␣B-crystallin as mutating this residue causes the human diseases, cataract [46] and DRM [3], respectively
Other mutational studies using the Mycobacterium tuberculosis sHSP, HSP16.3, and the mammalian sHSPs HSP27 and ␣A-crystallin [21] show this highly conserved arginine residue is structurally very important as it is part of a -strand involved in subunit interactions [21]
Summary
Desmin-related myopathy (DRM) can be caused by mutations either in the intermediate filament protein desmin, or in the small heat shock protein ␣B-crystallin. A characteristic disease pathology links the different causes of DRM, which consists of aggregates of intermediate filaments containing ␣B-crystallin present in the muscle cells of affected individuals [3, 4]. Other diseases, such as Alexander’s disease [5] and druginduced hepatitis [6], are characterized by intermediate filament aggregates. Similar structures can be generated in vitro under appropriate coassembly conditions [29] These studies show that the association of sHSPs with intermediate filament networks is not just a stress-induced event [30], but is a feature of normal cells, and suggest a general role for sHSPs in intermediate filament biology. It was subsequently demonstrated that sHSPs could inhibit GFAP as
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