University of Pittsburgh Department of Cell Biology
  • Research

    All secreted proteins, and most that ultimately reside within the cell, must traverse the secretory pathway, a network of intracellular organelles housing the "machines" that help secreted proteins mature. Critical components of these machines are a class of proteins known as molecular chaperones, some of which are associated with the endoplasmic reticulum (ER). However, if protein folding is inefficient or slow, a secreted protein may be targeted for destruction by a process we termed ER associated protein degradation (ERAD). During ERAD, proteins are selected as being defective and then degraded by the proteasome, a massive (~2 mDa) multi-catalytic protease that resides in the cytoplasm. Molecular chaperones may be required for ERAD by "deciding" whether a protein is sufficiently mature to transit through the secretory pathway; molecular chaperones may also direct these ERAD substrates to the proteasome. The importance of understanding the molecular mechanism of ERAD and molecular chaperone action is underscored by the fact that several human diseases-including cystic fibrosis, heart and liver disease, diabetes, and neurodegenerative diseases-can arise from defects in chaperone-mediated folding of secreted proteins and/or the ERAD pathway.

    For our studies, the Brodsky laboratory utilizes a model eukaryotic organism, the yeast Saccharomyces cerevisiae. Yeast possess the same intracellular membrane organization and molecular chaperones as human cells but are amenable to rapid genetic analysis. Recent research in the Brodsky laboratory is currently directed toward understanding how molecular chaperones in the ER and the cytoplasm facilitate ERAD and protein folding in the cell. Both endogenous proteins and human proteins expressed heterologously in yeast are being examined as substrates for ERAD and chaperone-mediated folding, and data derived from our genetic studies are complemented by biochemical assays that recapitulate specific aspects of these processes. More recently, because of the connection between molecular chaperone function and human disease, we have begun to identify and screen for small molecules that affect chaperone activity, and to use proteomic and genomic attacks to identify additional components that facilitate ERAD.

  • Publications

    1. Wright, C. M., R. J. Chovatiya, N. E. Jameson, D. M. Turner, G. Zhu, S. Werner, D. M. Huryn, J. M. Pipas, B. W. Day, P. Wipf, and J. L. Brodsky. Pyrimidinone-Peptoid Hybrid Molecules with Distinct Effects on Molecular Chaperone Function and Cell Proliferation. Bioorg. Med. Chem. (2008) 16: 3291-3301
    2. Tonsor, S. J., C. Scott, I. Boumaza, T. R. Liss, J. L. Brodsky, and E. Vierling. Heat shock protein 101 effects in A. thaliana: genetic variation, fitness, and phenotype under controlled temperate growth conditions. Molec. Ecol. (2008) 17: 1614-1626
    3. Nakatsukasa, K., G. Huyer, S. Michaelis, and J. L. Brodsky. Dissecting the ER- associated degradation of a misfolded polytopic membrane protein. Cell (2008) 132: 101-112
    4. Nakatsukasa, K. and J. L. Brodsky. Recognition and retro-translocation of misfolded proteins from the endoplasmic reticulum. Traffic (2008) 9: 861-870
    5. Kang, Y., T. Talsone, C. C. Clement, S. W. Fewell, J. Aguirre, J. L. Brodsky, and G. Chiosis. Design of a fluorescent polarization assay platform for the study of human Hsp70. Bioorg. Med. Chem. Lett. (2008) 18: 3749-3751
    6. Goeckeler, J. L., A, P. Petruso, J. Aguirre, C. C. Clement, G. Chiosis, and J. L. Brodsky. The yeast Hsp110, Sse1p, exhibits high affinity peptide binding. FEBS Lett. (2008) 582: 2393-2396
    7. Hrizo, S. L., V. Gusarova, D. M. Habiel, J. L. Goeckeler, E. A. Fisher, and J. L. Brodsky. The Hsp110 Molecular Chaperone Stabilizes Apolipoprotein B from Endoplasmic Reticulum Associated Degradation (ERAD). J. Biol. Chem. (2007) 282: 32665-32675
    8. Feng, D., Z. Xueqiang, C. Soromani, J. Toikkanen, K. Römisch, S. S. Vembar, J. L. Brodsky, S. Keränen, and J. Jäntti. The Trans-Membrane Domain is Sufficient for Sbh1p Function, its Association with the Sec61 Complex and Interaction with Rtn1p. J. Biol. Chem. (2007) 282: 30618-30628
    9. Kashlan, O.B., G. M. Mueller, M. Z. Qamar, P. A. Poland, A. Ahner, R. C. Rubenstein, R. P. Hughey, J. L. Brodsky, and T. R. Kleyman. Small heat shock protein aA-crystallin regulates ENaC expression. J. Biol. Chem. (2007) 282: 28149-28156
    10. Gusarova, V., J. Seo, M. L. Sullivan, S. C. Watkins, J. L. Brodsky, and E. A. Fisher. Golgi-associated maturation of very low density lipoproteins involves conformational changes of apolipoprotein B, but is not dependent on apolipoprotein E. J. Biol. Chem. (2007) 282:19453-19462
    11. Scott, C. M., K. B. Kruse, B. Z. Schmidt, D. H. Perlmutter, A. A. McCracken, and J. L. Brodsky. ADD66, A Gene Required for the Endoplasmic Reticulum Associated Degradation (ERAD) of Alpha-1-Antitrypsin-Z in Yeast, Facilitates Proteasome Activity and Assembly. Mol. Biol. Cell (2007) 18: 3774-3787
    12. Wahlman, J., G. N. DeMartino, W. R. Skach, N. J. Bulleid, J. L. Brodsky, and A. E. Johnson. Real-time fluorescence detection of ERAD substrate retro-translocation in a mammalian in vitro system. Cell (2007) 129: 943-955
    13. Buck, T., C. M. Wright, and J. L. Brodsky. The activities and functions of molecular chaperones in the endoplasmic reticulum. Seminars Cell. Dev. Biol. (2007) 18: 751-761
    14. Mandal, A., P. Lee, J. Chen, A. Heller, S. DiStasio, H. Oen, J. Victor, D. M. Nair, J. L. Brodsky and A. J. Caplan. Cdc37 has distinct roles in kinase biogenesis that affect synthesis and post-translational maturation. J. Cell Biol. (2007) 176: 319-328
    15. Wright, C. M., S. W. Fewell, M. L. Sullivan, J. M. Pipas, S. C. Watkins, and J. L. Brodsky. The Hsp40 molecular chaperone, Ydj1p, along with the protein kinase C pathway, impact cell wall integrity in the yeast Saccharomyces cerevisiae. Genetics (2007) 175: 1649-1664
    16. Rodina, A., M. Vilenchik, K. Moulick, J. Aguirre, J. Kim, A. Chiang, J. Litz, C. C. Clement, Y. Kang, Y. She, N. Wu, S. Felts, P. Wipf, J. Massague, X. Jiang, J. L. Brodsky, G. W. Krystal, and G. Chiosis. Selective chemical tools define Hsp90 as a major inhibitor of apoptosis in small cell lung cancer. Nature Chem. Biol. (2007) 3: 498-507
    17. Brodsky, J. L. The protective and destructive roles played by molecular chaperones during endoplasmic reticulum associated degradation (ERAD). Biochem. J., (2007) 404: 353-363
    18. Sparvero, L. J., S. Patz, J. L. Brodsky, and C. M. Coughlan. Proteomic analysis of the Amyloid Precursor Protein fragment C99: Expression in yeast. Analytic. Biochem. (2007) 370: 162-170
    19. Jelenska, J., N. Yao, B. A. Vinatzer, C. M. Wright, J. L. Brodsky, J. T. Greenberg. A J- domain Virulence Effector of Pseudomonas syringae Remodels Host Chloroplasts and Suppresses Defenses. Curr. Biol. (2007) 17: 499-508
    20. Ahner, A., K. Nakatsukasa, H. Zhang, R. A. Frizzell, and J. L. Brodsky. Small Heat Shock Proteins Select îF508-CFTR for ER Associated Degradation (ERAD). Mol. Biol. Cell (2007) 18: 806-814


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