Nuclear Aggregation of Polyglutamine-expanded Ataxin-3

Abstract

Expansion of a polymorphic polyglutamine segment is the common denominator of neurodegenerative polyglutamine diseases. The expanded proteins typically accumulate in large intranuclear inclusions and induce neurodegeneration. However, the mechanisms that determine the subcellular site and rate of inclusion formation are largely unknown. We found that the conserved putative nuclear localization sequence Arg-LysArg-Arg, which is retained in a highly aggregation-prone fragment of ataxin-3, did not affect the site and degree of inclusion formation in a cell culture model of spinocerebellar ataxia type 3. Addition of synthetic nuclear export or import signals led to the expected localization of ataxin-3 and determined the subcellular site of aggregate formation. Triggering a cellular stress response by heat shock transcription factor ⌬HSF1 coexpression abrogated aggregation in the cytoplasm but not in the nucleus. These findings indicate that native aggregation-prone fragments derived from expanded ataxin-3 may eventually escape the cytoplasmic quality control, resulting in aggregation in the nuclear compartment.Many human diseases are caused by the continuous expression of misfolded or misfolding-sensitive proteins (for review, see Ref. 1). The related polyglutamine (polyQ) 3 family of neurodegenerative diseases is caused by expansion of a variable polyQ segment above a threshold of typically ϳ40 residues. These expansions render the otherwise unrelated proteins susceptible to adopt non-native conformations that eventually become toxic or form toxic oligomers and aggregates. Large aggregates arise as intracellular inclusion bodies that are composed mainly of the mutant protein but also contain many other proteins, including transcription factors (2), molecular chaperones, and components of the ubiquitin-proteasome system (3). Several cellular pathways, including transcriptional dysregulation, inhibition of protein degradation, impairment of energy metabolism, and activation of cell death programs, have been implicated in the molecular mechanisms of the neurodegenerative processes caused by polyQ-expanded proteins (for review, see Ref. 4). These variations are likely determined by sequences outside the polyQ tract, which are specific for the individual disease proteins (5, 6). Even though toxicity has been experimentally dissociated from detectable inclusion body formation (e.g. Refs. 7 and 8), the hallmark of polyQ diseases is the characteristic formation of intraneuronal inclusions, most often in the nuclear compartment. Hence, it seems reasonable to argue that an imbalance in the production of toxic aggregation intermediates and their clearance constitutes the fundamental commonality of these diseases (9). Notably, the aggregation products accumulate, although cells are equipped with an elaborate network of molecular chaperones and cofactors with sufficient capacity to maintain a balanced protein homeostasis essential to cellular function (9, 10). Increasing evidence suggests that the nuclear compartment is the pr...

SUMO-1 Modification on K166 of PolyQ-Expanded aTaxin-3 Strengthens Its Stability and Increases Its Cytotoxicity

“…Therefore, as a central pathological event in polyQ disorders, aggregation needs to be better understood, particularly from a therapeutic point of view. In agreement with previous studies [40], we found the amount of aggregate formation cells in mutant-type ataxin-3 as much higher than that in normal control; demonstrating polyQ expansion could induce the formation of aggregates. Although there was no significantly difference in both aggregate cell counting and density quantification between ataxin-3-68Q and ataxin-3-68Q K166R , we could found the tendency that aggregate density of ataxin-3-68Q was slightly higher than that of ataxin-3-68Q K166R , which support the results of insoluble fraction detection and indicate that SUMOylation of mutant-type ataxin-3 might partially increase its stability and probably promote aggregate formation.…”

Toward understanding Machado–Joseph disease

“…C-terminal ATXN3 fragments cleaved proximal to amino acid 190 and containing the UIMs, expanded polyQ and NLS, are more abundant in the nuclear fraction from affected brain regions of MJD patients (Colomer Gould et al , 2007; Goti et al , 2004). Nuclear accumulation of these fragments may be due less to the presence of a weak NLS than to the absence of NES signals and, consequently, escape from chaperone-mediated clearance by the UPS in the cytoplasm (Antony et al , 2009; Breuer et al , 2010). The generation of C-terminal polyQ-containing fragments not only favors aggregation but also eliminates the putative protective action of the JD which is cleaved away by proteolysis (Warrick et al , 2005).…”

Aggregation of polyglutamine-expanded ataxin-3 sequesters its specific interacting partners into inclusions: Implication in a loss-of-function pathology

“…Due to the Josephin domain also possesses Ub-interaction and deubiquitination properties41, for simplicity, we applied the C-terminal fragments of Atx3-I (Atx3-IC) comprised of two UIMs and the polyQ tract as a model for the studies. Note that the C-terminal fragments of polyQ-expanded Atx3 are also implicated in the disease pathology384243. The experimental data showed that Atx3 100Q -IC sequestered more amount of ubiquitinated proteins, both in overexpressed (Fig.…”

“…S1). The data showed that the VBM motif did not overlap with the nuclear localization signal region of Atx33738, excluding the possibility that the reduced insoluble aggregates are caused by the subcellular distribution change. We also applied filter-trap assay to detect the aggregates formed by these three Atx3 species.…”