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Spinal and bulbar muscular atrophy (SBMA) is a neurodegenerative disorder caused by the expansion of a polyglutamine tract in the androgen receptor (AR). The N-terminal fragment of AR containing the expanded polyglutamine tract aggregates in cytoplasm and/or in nucleus and induces cell death. Some chaperones such as Hsp40 and Hsp70 have been identified as important regulators of polyglutamine aggregation and/or cell death in neuronal cells. Recently, Hsp105␣, expressed at especially high levels in mammalian brain, has been shown to suppress apoptosis in neuronal cells and prevent the aggregation of protein caused by heat shock in vitro. However, its role in polyglutamine-mediated cell death and toxicity has not been studied. In the present study, we examined the effects of Hsp105␣ on the aggregation and cell toxicity caused by expansion of the polyglutamine tract using a cellular model of SBMA. The transient expression of truncated ARs (tARs) containing an expanded polyglutamine tract caused aggregates to form in COS-7 and SK-N-SH cells and concomitantly apoptosis in the cells with the nuclear aggregates. When Hsp105␣ was overexpressed with tAR97 in the cells, Hsp105␣ was colocalized to aggregates of tAR97, and the aggregation and cell toxicity caused by expansion of the polyglutamine tract were markedly reduced. Both ␤-sheet and ␣-helix domains, but not the ATPase domain, of Hsp105␣ were necessary to suppress the formation of aggregates in vivo and in vitro. Furthermore, Hsp105␣ was found to localize in nuclear inclusions formed by ARs containing an expanded polyglutamine tract in tissues of patients and transgenic mice with SBMA. These findings suggest that overexpression of Hsp105␣ suppresses cell death caused by expansion of the polyglutamine tract without chaperone activity, and the enhanced expression of the essential domains of Hsp105␣ in brain may provide an effective therapeutic approach for CAG repeat diseases.

We previously reported that heat shock protein 105 (HSP105), identified by serological analysis of a recombinant cDNA expression library (SEREX) using serum from a pancreatic cancer patient, was overexpressed in various human tumors and in the testis of adult men by immunohistochemical analysis. In the present study, to elucidate the biological function of the HSP105 protein in cancer cells, we first established NIH3T3 cells overexpressing murine HSP105 (NIH3T3-HSP105). The NIH3T3-HSP105 cells acquired resistance to apoptosis induced by heat shock or doxorubicin. (1) is a stress protein belonging to the HSP105/110 family that is expressed constitutively in most tissues at low levels, whereas HSP105 mRNA is expressed at high levels in mouse and rat brain. At the protein level, high expression levels have been reported only in the brain of mice.(1-4) Like other heat shock proteins, HSP105 plays an important role as a chaperone under physiological conditions. HSP105 is induced by various stressors and plays an important role in protecting cells from the cytotoxic effects mediated by such stressors. HSP105 is composed of an ATP-binding domain, a β-sheet, a loop and α-helical domains similar to those observed in the HSP70 family of proteins, and it binds to non-native protein substrates, thereby preventing the aggregation of denatured protein through an interaction with the β-sheet domain of HSP105. (2,(5)(6)(7) The rat neuronal cell line PC12 transfected stably with murine HSP105 exhibited resistance to caspase-mediated apoptosis induced by stressors.(8) In a spinal and bulbar muscular atrophy model, the transient expression of tAR containing an expanded polyglutamine tract caused aggregates of polyglutamine in COS-7 and SK-N-SH cells and then induced the cells to undergo apoptosis. In contrast, in cells cotransfected with tAR and HSP105, both the aggregation of polyglutamine and the degree of it's cell toxicity decreased. (9) In contrast, HSP105 has demonstrated apoptosis-enhancing activity during murine embryogenesis.(10 -13) These observations suggest that HSP105 is involved in the regulation of apoptosis.We previously reported that HSP105 is overexpressed in various human tumors, including colon cancer cells but not colorectal adenomas, thus suggesting that the overexpression of HSP105 is a late event in the colorectal adenoma-carcinoma sequence. (14,15) Recent studies have also demonstrated that the expression level of HSP105 is elevated in highly metastatic colon cancer cell lines, and is correlated with advanced clinical stages and positive lymph node involvement. (16) RNA interference is used widely for manipulating biological systems, and has also been utilized successfully as a therapeutic material in experimental animals. (17)(18)(19) Synthetic siRNA strongly inhibits the expression of target proteins when they are transfected with cationic liposomes, which are thought to be safer than viral vectors for human therapy. The local injection of synthetic siRNA against VEGF (20) or sphingosine 1-phosphate rece...

Hsp105␣ is a mammalian member of the HSP105/110 family, a diverged subgroup of the HSP70 family. Hsp105␣ associates with Hsp70/Hsc70 as complexes in vivo and regulates the chaperone activity of Hsp70/ Hsc70 negatively in vitro and in vivo. In this study, we examined the mechanisms by which Hsp105␣ regulates Hsc70 chaperone activity. Using a series of deletion mutants of Hsp105␣ and Hsc70, we found that the interaction between Hsp105␣ and Hsc70 was necessary for the suppression of Hsc70 chaperone activity by Hsp105␣. Furthermore, Hsp105␣ and deletion mutants of Hsp105␣ that interacted with Hsc70 suppressed the ATPase activity of Hsc70, with the concomitant appearance of ATPase activity of Hsp105␣. As the ATPase activity of Hsp70/Hsc70 is essential for the efficient folding of nonnative protein substrates, Hsp105␣ is suggested to regulate the substrate binding cycle of Hsp70/Hsc70 by inhibiting the ATPase activity of Hsp70/Hsc70, thereby functioning as a negative regulator of the Hsp70/Hsc70 chaperone system.