Nuclear Aggresomes Form by Fusion of PML-associated Aggregates

Abstract

Nuclear aggregates formed by proteins containing expanded poly-glutamine (poly-Q) tracts have been linked to the pathogenesis of poly-Q neurodegenerative diseases. Here, we show that a protein (GFP170*) lacking poly-Q tracts forms nuclear aggregates that share characteristics of poly-Q aggregates. GFP170* aggregates recruit cellular chaperones and proteasomes, and alter the organization of nuclear domains containing the promyelocytic leukemia (PML) protein. These results suggest that the formation of nuclear aggregates and their effects on nuclear architecture are not specific to poly-Q proteins. Using GFP170* as a model substrate, we explored the mechanistic details of nuclear aggregate formation. Fluorescence recovery after photobleaching and fluorescence loss in photobleaching analyses show that GFP170* molecules exchange rapidly between aggregates and a soluble pool of GFP170*, indicating that the aggregates are dynamic accumulations of GFP170*. The formation of cytoplasmic and nuclear GFP170* aggregates is microtubule-dependent. We show that within the nucleus, GFP170* initially deposits in small aggregates at or adjacent to PML bodies. Time-lapse imaging of live cells shows that small aggregates move toward each other and fuse to form larger aggregates. The coalescence of the aggregates is accompanied by spatial rearrangements of the PML bodies. Significantly, we find that the larger nuclear aggregates have complex internal substructures that reposition extensively during fusion of the aggregates. These studies suggest that nuclear aggregates may be viewed as dynamic multidomain inclusions that continuously remodel their components. INTRODUCTIONNewly synthesized proteins must be properly folded and modified to function correctly. Eukaryotic cells have developed extensive folding machineries to ensure the fidelity of protein processing. Nevertheless, misfolding can occur due to mutations within a protein, outside stresses, or the overexpression of proteins. Misfolded proteins often expose their hydrophobic domains, which leads to nonproductive protein associations and results in aggregation. Aggregated proteins tend to coalesce and form large deposits termed inclusion bodies, Russell bodies, or aggresomes, depending on their composition and location. Formation of such inclusions underlies a number of aggresomal diseases, including Alzheimer's disease, Parkinson's disease, familial amyotrophic lateral sclerosis, and the poly-glutamine (poly-Q) neuropathologies (reviewed in Zoghbi and Orr, 2000;Garcia-Mata et al., 2002).The biological processes leading to protein aggregation have been actively investigated (reviewed in Kopito, 2000;Garcia-Mata et al., 2002;Goldberg, 2003;Selkoe, 2003). Aggregation of proteins most likely occurs cotranslationally, while nascent peptide chains are synthesized on polyribosomes. If the nascent peptides cannot fold correctly, they will aggregate to form aggresomal particles. Small aggresomal particles form throughout the cell and are quickly transported toward the microtubule (MT)...

Efficient Induction of Nuclear Aggresomes by Specific Single Missense Mutations in the DNA-binding Domain of a Viral AP-1 Homolog

“…Truncated ataxin-2 and GFP-170* were distributed as a solid mass within their respective aggresomes. Nuclear aggresomes induced by full-length ataxin-3 or by GFP-170* did not recruit SC35 (12,15). As has been proposed, the differences in organization and composition seen among various nuclear aggresomes may reflect variations in the cellular aggresomal response to differences in the aggregating proteins (12).…”

“…They possess complex internal substructures that undergo extensive repositioning as aggresomes increase in size (15). The larger aggresomes induced by mutant ZEBRA proteins were organized into at least two distinct layers.…”

“…For example, truncated and tagged ataxin-3 and ataxin-1 proteins localized to the aggresomal periphery in a ring-shaped pattern in each of their respective aggresomes (12). In nuclear aggresomes induced by mutant ataxin-3 or by GFP-170*, PML existed as a globular mass in the aggresomal interior (12,15). Nuclear aggresomes induced by truncated ataxin-3 recruited SC35 protein.…”

Expression profile and subcellular localization of Torque teno sus virus proteins

“…According to the size and shape, globular deposits could correspond to aggregation of misfolded protein. The formation of protein aggregates is a dynamic process that cells use to deal with misfolded protein resulting from cell stress, overexpresion of proteins or viral infection (Fu et al, 2005;Kopito, 2000). Protein aggregates have also been associated with PML bodies.…”

Herpes simplex virus type I disrupts the ATR-dependent DNA-damage response during lytic infection

“…The redistribution of Hsc70, Hsp70, Hsp40, Hsp90, ubiquitylated proteins and the core catalytic complex of the proteasome into VICE domains in infected cells suggests that HSV-1 has evolved a mechanism to deal with misfolded and unwanted proteins (Burch and Weller, 2004;Burch and Weller, 2005;Parkinson and Everett, 2001). VICE domains are reminiscent of nuclear aggresomes, which form in response to misfolded proteins and contain heat-shock proteins and components of the ubiquitin proteasome (Anton et al, 1999;Fu et al, 2005). P-RPA and ATRIP may be targeted to VICE domains because they are recognized as misfolded, perhaps because they have been separated from their normal interaction partners as described below.…”