Abstract
Formation of fibrillar intranuclear inclusions and related neuropathologies of the CAG-repeat disorders are linked to the expansion of a polyglutamine tract. Despite considerable effort, the etiology of these devastating diseases remains unclear. Although polypeptides with glutamine tracts recapitulate many of the observed characteristics of the gene products with CAG repeats, such as in vitro and in vivo aggregation and toxicity in model organisms, extended polyglutamine segments have also been reported to structurally perturb proteins into which they are inserted. Additionally, the sequence context of a polyglutamine tract has recently been shown to modulate its propensity to aggregate. These findings raise the possibility that indirect influences of the repeat tract on adjacent protein domains are contributory to pathologies. Destabilization of an adjacent domain may lead to loss of function, as well as favoring non-native structures in the neighboring domain causing them to be prone to intermolecular association and consequent aggregation. To explore these phenomena, we have used chimeras of a well studied globular protein and exon 1 of huntingtin. We find that expansion of the polyglutamine segment beyond the pathological threshold (>35 glutamines) results in structural perturbation of the neighboring protein whether the huntingtin exon is N- or C-terminal. Elongation of the polyglutamine region also substantially increases the propensity of the chimera to aggregate, both in vitro and in vivo, and in vitro aggregation kinetics of a chimera with a 53-glutamine repeat follow a nucleation polymerization mechanism with a monomeric nucleus.
Highlights
Onset and disease severity [5]
In ataxin-3, linked to type 3 spinocerebellar ataxia (Machado-Joseph disease), the poly(Q) repeat is located in the C-terminal portion of the protein, and there is a similar correlation of repeat length with disease severity and onset [6]
In contrast to the recent study of ataxin-3 [30], we find that poly(Q) expansion within Htt exon 1 in the pathological range causes structural perturbation and decreased stability of the neighboring cellular retinoic acid binding protein I (CRABP I) protein domain as well as increased propensity for aggregation both in vivo and in vitro, whereas a fusion of CRABP I with Htt exon 1 containing a shorter poly(Q) repeat (n ϭ 20) remains soluble with no evidence of structural disruption
Summary
Onset and disease severity [5]. In the specific case of HD, the elongated CAG repeat encoding a stretch of glutamines is located within the N-terminal exon 1 of the gene encoding the 350-kDa protein ‘huntingtin’ (htt). Poly(Q) Tracts Perturb Adjacent Domains linked to a solubility-enhancing fusion partner, maltose-binding protein Their findings appeared to be supported by another study of a non-pathological variant of the protein ataxin-3, which was observed to form fibrillar aggregates upon partial denaturation using guanidine HCl [29]. In the latter study the authors were forced to use pH denaturation and kinetics of unfolding/folding to obtain stability data, because aggregation processes confounded the use of chaotropes such as guanidinium hydrochloride They concluded that the length-dependent influence of the poly(Q) region was not on the native state of the flanking domain but rather on a fibrillogenic state. While not uniform in their conclusions, point to the importance of understanding how poly(Q) tracts influence the sequences around them, and in turn how adjacent sequences may influence the behavior of poly(Q) repeats, to better understand CAG-repeat diseases
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