Focal distortion of the nuclear envelope by huntingtin aggregates revealed by lamin immunostaining

Abstract

Huntington's disease is an autosomal dominant neurodegenerative disorder caused by the expansion of a polyglutamine repeat tract in the huntingtin protein. Polyglutamine-expanded huntingtin forms intranuclear as well as perinuclear inclusion bodies. Perinuclear aggregates formed by polyglutamine-expanded proteins are associated with a characteristic indentation of the nuclear envelope. We examined the nuclear envelope in cells containing huntingtin aggregates using immunostaining for lamin B1, a major component of the nuclear lamina. Laser confocal microscopy analysis revealed that huntingtin aggregates in a juxtanuclear position were associated with a clear focal distortion in the nuclear envelope in cells transfected with polyglutamine-expanded huntingtin. Lamin B1 distribution was not altered by aggregates of polyglutamine-expanded ataxin-1, that are exclusively intranuclear. Thus lamin immunocytochemistry demonstrates clearly the depression of the nuclear envelope resulting from the formation of perinuclear aggregates by polyglutamine-expanded huntingtin. Lamin immunocytochemistry would be of value to monitor the state of the nuclear envelope in experimental paradigms aimed at establishing the significance of perinuclear aggregates of pathogenic proteins.

Fig. 1

Confocal microscopy analysis of the nuclear lamina in polyQ-expanded huntingtin-expressing cells. CHO cells were transfected with huntingtin exon 1 with 103 glutamines tagged with EGFP (Htt103Q). Cells were stained with an antibody to lamin B1 and imaged by laser scanning confocal microscopy. Perinuclear Htt103Q aggregates are associated with a distortion of the nuclear envelope, clearly evident with lamin B1 staining. By contrast, cells with cytoplasmic aggregates (arrows) have an intact nuclear envelope. (A) EGFP fluorescence, (B) lamin B1 staining, and (C) merged signals. Scale bar = 10 μm.

Fig. 2

Intranuclear aggregates of polyQ-expanded huntingtin do not alter lamin B1 distribution in transfected cells. CHO cells were transfected with huntingtin exon 1 with 103 glutamines tagged with EGFP (Htt103Q) and stained for lamin B1. (A) EGFP fluorescence, (B) lamin B1 staining, and (C) merged signals. Scale bar = 10 μm.

Fig. 3

Nuclear aggregates of ataxin-1 do not alter lamin B1 distribution in transfected cells. CHO cells were transfected with EGFP-tagged ataxin-1 with 82 glutamines (Atxn82Q) and stained for lamin B1. Atxn82Q forms characteristic intranuclear aggregates but the nuclear envelope or the distribution of lamin B1 are not affected. (A) EGFP fluorescence, (B) lamin B1 staining, and (C) merged signals. Scale bar = 10 μm.