Browsing by Author "Song, Wensi"

Now showing 1 - 3 of 3
  • Thumbnail Image
    Item
    Modulating the Lysosome-Autophagy System to Restore Homeostasis in in vitro Model Systems of Lysosomal Storage Disorders
    (2014-11-14) Song, Wensi; Segatori, Laura; Matthews, Kathleen; Gonzalez, Ramon
    The protein quality control system is a complex network that promotes the folding and trafficking of newly synthesized proteins and regulates the degradation of misfolded proteins and protein aggregates. Failure of the quality control system to maintain protein homeostasis (or proteostasis) characterizes the cellular pathogenesis of a number of human diseases. In particular, this study focuses on lysosomal storage disorders, a group of inherited metabolic diseases characterized by deficiencies in specific lysosomal hydrolytic activities that result from mutations in genes encoding for lysosomal proteins and consequent buildup of lysosomal storage material. The ultimate goal of this work is to develop cell engineering strategies to modulate cellular quality control machineries that control protein folding, processing, and degradation to restore cellular homeostasis under conditions of proteotoxic stress. Specifically, this study aims to manipulate the lysosome-autophagy system to enhance folding and processing of lysosomal enzymes as well as to enhance the cellular clearance capacity. To achieve this goal, I investigated the role of transcription factor EB (TFEB), a master regulator of lysosomal biogenesis and function, in regulating lysosomal proteostasis and autophagic clearance. Specifically, chemical and genetic modulation of TFEB was found to enhance folding, trafficking and activity of unstable, degradation-prone lysosomal enzymes in in vitro models of lysosomal storage disorders. Moreover, pharmacologic activation of autophagy achieved by treating cells with 2-hydroxypropyl-β-cyclodextrin was found to enhance autophagic clearance of storage material specifically by activating TFEB. To further investigate the molecular mechanism of autophagy induction and activation of autophagic clearance, I tested the impact of polystyrene nanoparticles of different size and surface charge on the lysosome-autophagy system with the ultimate goal to link the physicochemical properties of nanomaterials with the specific nature of the autophagic response activated upon nanomaterial uptake into cells. Efficient autophagic clearance was found to depend highly on the surface charge. Specifically, cell exposure to polystyrene nanoparticles presenting neutral or negative surface charge results in activation of autophagic clearance, whereas cell exposure to polystyrene nanoparticles presenting cationic surface charge results in impairment of lysosomal function and blockage of autophagic flux. Ceria nanoparticles (or nanoceria) are widely used in a variety of applications including as UV blockers and catalysts in industrial processes. Recent studies also revealed that ceria nanoparticles present antioxidant properties, suggesting a potential role of nanoceria in a variety of biomedical applications. In this study, I investigated the impact of ceria nanoparticles stabilized by organic surface coatings on the lysosome-autophagy system, Ceria nanoparticles were found to activate the lysosome-autophagy system and enhance autophagic clearance. In summary, this work provides proof-of-principle demonstration of chemical and biological strategies to activate the lysosome-autophagy system for restoring lysosomal proteostasis and enhancing autophagic clearance in model systems of diseases characterized by deficiencies in lysosomal enzymes activities and aberrant accumulation of undegraded lysosomal substrates. These findings lay the foundation for the development of nanotherapeutics for the treatment of diseases associated with inefficient autophagic clearance.
  • Thumbnail Image
    Item
    TFEB regulates lysosomal proteostasis
    (Oxford University Press, 2013) Song, Wensi; Wang, Fan; Savini, Marzia; Ake, Ashley; di Ronza, Alberto; Sardiello, Marco; Segatori, Laura
    Loss-of-function diseases are often caused by destabilizing mutations that lead to protein misfolding and degradation. Modulating the innate protein homeostasis (proteostasis) capacity may lead to rescue of native folding of the mutated variants, thereby ameliorating the disease phenotype. In lysosomal storage disorders (LSDs), a number of highly prevalent alleles have missense mutations that do not impair the enzyme's catalytic activity but destabilize its native structure, resulting in the degradation of the misfolded protein. Enhancing the cellular folding capacity enables rescuing the native, biologically functional structure of these unstable mutated enzymes. However, proteostasis modulators specific for the lysosomal system are currently unknown. Here, we investigate the role of the transcription factor EB (TFEB), a master regulator of lysosomal biogenesis and function, in modulating lysosomal proteostasis in LSDs. We show that TFEB activation results in enhanced folding, trafficking and lysosomal activity of a severely destabilized glucocerebrosidase (GC) variant associated with the development of Gaucher disease (GD), the most common LSD. TFEB specifically induces the expression of GC and of key genes involved in folding and lysosomal trafficking, thereby enhancing both the pool of mutated enzyme and its processing through the secretory pathway. TFEB activation also rescues the activity of a β-hexosaminidase mutant associated with the development of another LSD, Tay–Sachs disease, thus suggesting general applicability of TFEB-mediated proteostasis modulation to rescue destabilizing mutations in LSDs. In summary, our findings identify TFEB as a specific regulator of lysosomal proteostasis and suggest that TFEB may be used as a therapeutic target to rescue enzyme homeostasis in LSDs.
  • Thumbnail Image
    Item
    The autophagic response to polystyrene nanoparticles is mediated by transcription factor EB and depends on surface charge
    (BioMed Central, 2015) Song, Wensi; Popp, Lauren; Yang, Justin; Kumar, Ayushi; Gangoli, Varun S.; Segatori, Laura
    Background: A number of engineered nanoparticles induce autophagy, the main catabolic pathway that regulates bulk degradation of cytoplasmic material by the lysosomes. Depending on the specific physico-chemical properties of the nanomaterial, however, nanoparticle-induced autophagy may have different effects on cell physiology, ranging from enhanced autophagic degradation to blockage of autophagic flux. To investigate the molecular mechanisms underlying the impact of nanoparticle charge on the nature of the autophagic response, we tested polystyrene nanoparticles (50 nm) with neutral, anionic, and cationic surface charges. Results: We found all polystyrene nanoparticles investigated in this study to activate autophagy. We showed that internalization of polystyrene nanoparticles results in activation of the transcription factor EB, a master regulator of autophagy and lysosome biogenesis. Autophagic clearance, however, was observed to depend specifically on the charge of the nanoparticles. Particularly, we found that the autophagic response to polystyrene nanoparticles presenting a neutral or anionic surface involves enhanced clearance of autophagic cargo. Cell exposure to polystyrene nanoparticles presenting a cationic surface, on the other hand, results in transcriptional upregulation of the pathway, but also causes lysosomal dysfunction, ultimately resulting in blockage of autophagic flux. Conclusions: This study furthers our understanding of the molecular mechanisms that regulate the autophagic response to nanoparticles, thus contributing essential design criteria for engineering benign nanomaterials.