Figure 4.
The MT network in COS-7 cells expressing Cdk5, p35, and/or mhtt. A, MTs in COS-7 cells expressing Cdk5 and p35 were observed by immunofluorescence staining with anti-β-tubulin antibody 24 h after transfection (top). Bottom, Immunoblots showing the amount of β-tubulin in the pellet (p) and supernatant (s) in control cells (−), cells treated with nocodazole for 18 h (Noc), transfected with Cdk5/p35 (Cdk5/p35), or treated with Taxol for 18 h (Tax). Polymeric and protomeric tubulin are expressed as the percentage ratio to total β-tubulin under the each lane. B, MT distribution in COS-7 cells expressing tNhtt-16Q, 50Q, or 90Q-EGFP. MTs were stained with anti-β-tubulin antibody (red), and tNhtt-poly(Q)-EGFP was detected by EGFP fluorescence (green). The merged image is shown (bottom). The arrows indicate the discrete tubulin staining observed in the cytoplasm and at the cell periphery. C, Confocal microscopic images of tNhtt-90Q-EGFP (green; top), anti-β-tubulin staining (red; middle), and merged images (bottom) in COS-7 cells expressing Cdk5/p35 or not (vector). D, tNhtt-90Q-EGFP was transfected into COS-7 cells alone or with Cdk5 and p35. Cells were treated with 20 μm Taxol for 18 h starting 6 h after transfection. At the end of treatment, cells were observed for inclusions (left). Cells containing EGFP inclusions were counted and expressed as the percentage ratio to total EGFP-positive cells (right; mean ± SEM; **p < 0.001). Scale bars, 10 μm.