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Open Access Publications from the University of California

Scholarly Works (8 results)

  • Thesis
  • Peer Reviewed
UC San Francisco Electronic Theses and Dissertations (2020)

Cells must routinely recycle their proteins in order to maintain protein homeostasis and health. The bulk of protein turnover is facilitated by the proteasome and locally restricted within the center of the barrel-shaped 20S core. In order to be degraded, potential substrates must enter the core through an opening at the center of this complex. The pore is gated by N-terminal extensions that limit substrate entry and can be opened by native activators, which functionally regulate protein degradation. Gate-opening occurs through HbYX motifs at the extreme C-termini of proteasome activators. To date, characterization of the HbYX mechanism has been limited to protein-protein interactions of archaeal proteasome complexes. We have uncovered unexpected differences in the residue specificity of this motif for activating the human 20S proteasome. These sequence requirements are tuned by valency, which was generally corroborated by the C-termini of natural activators, providing insight into identifying new binding partners of the eukaryotic 20S proteasome. This work has established an alternative model to the HbYX mechanism for eukaryotic proteasome complexes. Finally, the Y motif offers an updated template for the design of pharmacological activators, critical protein-protein interface will deepen our understanding of proteasome biology and its implication in disease.

    Cover page: Redefining HbYX-dependent activation of the human 20S proteasome
    • Article
    • Peer Reviewed
    UC San Francisco Previously Published Works (2018)
    Neurodegenerative diseases are a class of age-associated proteopathies characterized by the accumulation of misfolded and/or aggregation-prone proteins. This imbalance has been attributed, in part, to an age-dependent decay in the capacity of protein turnover. Most proteins are degraded by the ubiquitin-proteasome system (UPS), which is composed of ubiquitin ligases and regulatory particles, such as the 19S, that deliver cargo to the proteolytically active 20S proteasome (20S) core. However, a subset of clients, especially intrinsically disordered proteins (IDPs), are also removed by the action of the ubiquitin-independent proteasome system (UIPS). What are the specific contributions of the UPS and UIPS in the context of neurodegeneration? Here, we explore how age-associated changes in the relative contribution of the UPS and UIPS, combined with the IDP-like structure of many neurodegenerative disease-associated proteins, might contribute. Strikingly, the 20S has been shown to predominate in older neurons and to preferentially act on relevant substrates, such as synuclein and tau. Moreover, pharmacological activation of the 20S has been shown to accelerate removal of aggregation-prone proteins in some models. Together, these recent studies are turning attention to the 20S and the UIPS as potential therapeutic targets in neurodegeneration.
      Cover page: Aim for the core: suitability of the ubiquitin-independent 20S proteasome as a drug target in neurodegeneration
      • Article
      • Peer Reviewed
      UC San Francisco Previously Published Works (2020)
      An amendment to this paper has been published and can be accessed via a link at the top of the paper.
        Cover page: Author Correction: Deubiquitinase Usp12 functions noncatalytically to induce autophagy and confer neuroprotection in models of Huntington’s disease
        • Article
        • Peer Reviewed
        UC San Francisco Previously Published Works (2018)
        Huntington's disease is a progressive neurodegenerative disorder caused by polyglutamine-expanded mutant huntingtin (mHTT). Here, we show that the deubiquitinase Usp12 rescues mHTT-mediated neurodegeneration in Huntington's disease rodent and patient-derived human neurons, and in Drosophila. The neuroprotective role of Usp12 may be specific amongst related deubiquitinases, as the closely related homolog Usp46 does not suppress mHTT-mediated toxicity. Mechanistically, we identify Usp12 as a potent inducer of neuronal autophagy. Usp12 overexpression accelerates autophagic flux and induces an approximately sixfold increase in autophagic structures as determined by ultrastructural analyses, while suppression of endogenous Usp12 slows autophagy. Surprisingly, the catalytic activity of Usp12 is not required to protect against neurodegeneration or induce autophagy. These findings identify the deubiquitinase Usp12 as a regulator of neuronal proteostasis and mHTT-mediated neurodegeneration.
          Cover page: Deubiquitinase Usp12 functions noncatalytically to induce autophagy and confer neuroprotection in models of Huntington’s disease
          • Article
          • Peer Reviewed
          UC San Francisco Previously Published Works (2020)
          An amendment to this paper has been published and can be accessed via a link at the top of the paper.
            Cover page: Publisher Correction: Deubiquitinase Usp12 functions noncatalytically to induce autophagy and confer neuroprotection in models of Huntington’s disease
            • Article
            • Peer Reviewed
            UC San Francisco Previously Published Works (2022)
            The 20S proteasome (20S) facilitates turnover of most eukaryotic proteins. Substrate entry into the 20S first requires opening of gating loops through binding of HbYX motifs that are present at the C-termini of certain proteasome activators (PAs). The HbYX motif has been predominantly characterized in the archaeal 20S, whereas little is known about the sequence preferences of the human 20S (h20S). Here, we synthesize and screen ~120 HbYX-like peptides, revealing unexpected differences from the archaeal system and defining the h20S recognition sequence as the Y-F/Y (YФ) motif. To gain further insight, we create a functional chimera of the optimized sequence, NLSYYT, fused to the model activator, PA26E102A. A cryo-EM structure of PA26E102A-h20S is used to identify key interactions, including non-canonical contacts and gate-opening mechanisms. Finally, we demonstrate that the YФ sequence preferences are tuned by valency, allowing multivalent PAs to sample greater sequence space. These results expand the model for termini-mediated gating and provide a template for the design of h20S activators.
              Cover page: The YΦ motif defines the structure-activity relationships of human 20S proteasome activators
              • Article
              • Peer Reviewed
              UC San Francisco Previously Published Works (2018)
              The original version of this Article incorrectly gave a publication date of 8 October 2018; this should have been 28 September 2018. This has now been corrected in the PDF and HTML versions of the Article.
                Cover page: Publisher Correction: Deubiquitinase Usp12 functions noncatalytically to induce autophagy and confer neuroprotection in models of Huntington’s disease
                • Article
                • Peer Reviewed
                UC San Francisco Previously Published Works (2019)
                Protein-protein interactions between E3 ubiquitin ligases and protein termini help shape the proteome. These interactions are sensitive to proteolysis, which alters the ensemble of cellular N and C termini. Here we describe a mechanism wherein caspase activity reveals latent C termini that are then recognized by the E3 ubiquitin ligase CHIP. Using expanded knowledge of CHIP's binding specificity, we predicted hundreds of putative interactions arising from caspase activity. Subsequent validation experiments confirmed that CHIP binds the latent C termini at tauD421 and caspase-6D179. CHIP binding to tauD421, but not tauFL, promoted its ubiquitination, while binding to caspase-6D179 mediated ubiquitin-independent inhibition. Given that caspase activity generates tauD421 in Alzheimer's disease (AD), these results suggested a concise model for CHIP regulation of tau homeostasis. Indeed, we find that loss of CHIP expression in AD coincides with the accumulation of tauD421 and caspase-6D179. These results illustrate an unanticipated link between caspases and protein homeostasis.
                  Cover page: Specificity for latent C termini links the E3 ubiquitin ligase CHIP to caspases
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