In the last decade or so, developmental biologists have provided enormous amounts of information on the morphological events of Drosophila development from oogenesis to adult body structures, including the developing and adult nervous systems and the eyes (38,47,48). Histochemical staining techniques were developed to decipher complex regulation of gene expression by antibody staining and in situ hybridization.
Antibodies that recognize recombinant polyglutamine proteins have been used to monitor the protein expression pattern in Drosophila. When expressed in the developing Drosophila eye tissues (the eye imaginal discs), progressive nuclear translocation and aggregation of expanded polyglutamine proteins are detected (25,26). In adult photoreceptor neurons, nuclear translocation of the expanded polyglutamine was suggested to be dependent on the length of the polyglutamine expansion (26). In addition, cellular proteins such as heat-inducible molecular chaperone Hsp70 (33) and polyglutamine-con-taining developmental protein EYA (49) have been found to co-localize with expanded polyglutamine protein in nuclear inclusions in eye tissues. Interestingly, when molecular chaperones are co-expressed with expanded polyglutamine proteins in neurons, no morphological alteration of the nuclear inclusions by immunocytochemistry was noted, despite suppression of neural degeneration (28,33,34). These findings provide evidence that degeneration can be dissociated from the presence of nuclear inclusions.
Biochemical analysis has been applied to examine chaperone-mediated suppression of polyglutamine toxicity in vivo (34). Coexpression of chaperones Hsp70 and dHdj1 increases the amount of monomeric disease protein upon detergent extraction (2% sodium dodecyl sulfate [SDS]) (34). When dHdj2, a modest suppressor, is coexpressed with the polyglutamine protein, only a mild suppression is observed that parallels a limited increase in the amount of the monomeric disease protein seen by Western blot analysis (34). These findings suggest that chaperones alter the solubility properties of polyglutamine protein, concomitant with the degree of suppression of toxicity in vivo. In addition to a change in solubility properties in detergent, chaperones also appear to have some ability to alter salt extractability (250 mM sodium chloride) of the disease polyglutamine protein (34). These observations indicate that chaperones suppress polyglutamine toxicity by altering the biochemical properties of the toxic polyglutamine protein.
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