Potentiation of amyotrophic lateral sclerosis (ALS)-associated TDP-43 aggregation by the proteasome-targeting factor, ubiquilin 1.
ABSTRACT TDP-43 (43-kDa TAR DNA-binding domain protein) is a major constituent of ubiquitin-positive cytoplasmic aggregates present in neurons of patients with fronto-temporal lobular dementia and amyotrophic lateral sclerosis (ALS). The pathologic significance of TDP-43 aggregation is not known; however, dominant mutations in TDP-43 cause a subset of ALS cases, suggesting that misfolding and/or altered trafficking of TDP-43 is relevant to the disease process. Here, we show that the presenilin-binding protein ubiquilin 1 (UBQLN) plays a role in TDP-43 aggregation. TDP-43 interacted with UBQLN both in yeast and in vitro, and the carboxyl-terminal ubiquitin-associated domain of UBQLN was both necessary and sufficient for binding to polyubiquitylated forms of TDP-43. Overexpression of UBQLN recruited TDP-43 to detergent-resistant cytoplasmic aggregates that colocalized with the autophagosomal marker, LC3. UBQLN-dependent aggregation required the UBQLN UBA domain, was mediated by non-overlapping regions of TDP-43, and was abrogated by a mutation in UBQLN previously linked to Alzheimer disease. Four ALS-associated alleles of TDP-43 also coaggregated with UBQLN, and the extent of aggregation correlated with in vitro UBQLN binding affinity. Our findings suggest that UBQLN is a polyubiquitin-TDP-43 cochaperone that mediates the autophagosomal delivery and/or proteasome targeting of TDP-43 aggregates.
- SourceAvailable from: Nam-Hyuk Cho[show abstract] [hide abstract]
ABSTRACT: Pathogens exploit host machinery to establish an environment that favors their propagation. Because of their pivotal roles in cellular physiology, protein degradation pathways are common targets for viral proteins. Protein-linking integrin-associated protein and cytoskeleton 1 (PLIC1), also called ubiquilin, contains an amino-terminal ubiquitin-like (UBL) domain and a carboxy-terminal ubiquitin-associated (UBA) domain. PLIC1 is proposed to function as a regulator of the ubiquitination complex and proteasome machinery. Kaposi's sarcoma-associated herpesvirus (KSHV) contains a small membrane protein, K7, that protects cells from apoptosis induced by various stimuli. We report here that cellular PLIC1 is a K7-interacting protein and that the central hydrophobic region of K7 and the carboxy-terminal UBA domain of PLIC1 are responsible for their interaction. Cellular PLIC1 formed a dimer and bound efficiently to polyubiquitinated proteins through its carboxy-terminal UBA domain, and this activity correlated with its ability to stabilize cellular I kappa B protein. In contrast, K7 interaction prevented PLIC1 from forming a dimer and binding to polyubiquitinated proteins, leading to the rapid degradation of I kappa B. Furthermore, K7 expression promoted efficient degradation of the p53 tumor suppressor, resulting in inhibition of p53-mediated apoptosis. These results indicate that KSHV K7 targets a regulator of the ubiquitin- and proteasome-mediated degradation machinery to deregulate cellular protein turnover, which potentially provides a favorable environment for viral reproduction.Molecular and Cellular Biology 06/2004; 24(9):3938-48. · 5.37 Impact Factor
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ABSTRACT: The Amyloid Precursor Protein (APP) undergoes sequential proteolytic cleavages through the action of beta- and gamma-secretase, which result in the generation of toxic beta-amyloid (Abeta) peptides and a C-terminal fragment consisting of the intracellular domain of APP (AICD). Mutations leading to increased APP levels or alterations in APP cleavage cause familial Alzheimer's disease (AD). Thus, identification of factors that regulate APP steady state levels and/or APP cleavage by gamma-secretase is likely to provide insight into AD pathogenesis. Here, using transgenic flies that act as reporters for endogenous gamma-secretase activity and/or APP levels (GAMAREP), and for the APP intracellular domain (AICDREP), we identified mutations in X11L and ubiquilin (ubqn) as genetic modifiers of APP. Human homologs of both X11L (X11/Mint) and Ubqn (UBQLN1) have been implicated in AD pathogenesis. In contrast to previous reports, we show that overexpression of X11L or human X11 does not alter gamma-secretase cleavage of APP or Notch, another gamma-secretase substrate. Instead, expression of either X11L or human X11 regulates APP at the level of the AICD, and this activity requires the phosphotyrosine binding (PTB) domain of X11. In contrast, Ubqn regulates the levels of APP: loss of ubqn function leads to a decrease in the steady state levels of APP, while increased ubqn expression results in an increase in APP levels. Ubqn physically binds to APP, an interaction that depends on its ubiquitin-associated (UBA) domain, suggesting that direct physical interactions may underlie Ubqn-dependent regulation of APP. Together, our studies identify X11L and Ubqn as in vivo regulators of APP. Since increased expression of X11 attenuates Abeta production and/or secretion in APP transgenic mice, but does not act on gamma-secretase directly, X11 may represent an attractive therapeutic target for AD.PLoS ONE 02/2008; 3(6):e2495. · 3.73 Impact Factor
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ABSTRACT: TAR DNA binding protein (TDP43), a highly conserved heterogeneous nuclear ribonucleoprotein, was found to down-regulate splicing of the exon 9 cystic fibrosis transmembrane conductance regulator (CFTR) through specific binding to a UG-rich polymorphic region upstream of the 3' splice site. Despite the emergence of new information regarding the protein's nuclear localization and splicing regulatory activity, TDP43's role in cells remains elusive. To investigate the function of human TDP43 and its homologues, we cloned and characterized the proteins from Drosophila melanogaster and Caenorhabditis elegans. The proteins from human, fly, and worm show striking similarities in their nucleic acid binding specificity. We found that residues at two different positions, which show a strong conservation among TDP43 family members, are linked to the tight recognition of the target sequence. Our three-dimensional model of TDP43 in complex with a (UG)(m) sequence predicts that these residues make amino acid side-chain to base contacts. Moreover, our results suggest that Drosophila TDP43 is comparable to human TDP43 in regulating exon splicing. On the other hand, C.elegans TDP43 has no effect on exon recognition. TDP43 from C.elegans lacks the glycine-rich domain found at the carboxy terminus of the other two homologues. Mutants of human and fly TDP43 devoid of the C-terminal domain are likewise unable to affect splicing. Our studies suggest that the glycine-rich domain is essential for splicing regulation by human and fly TDP43.Journal of Molecular Biology 06/2005; 348(3):575-88. · 3.91 Impact Factor