Faculty Research Page
Professor of Biochemistry, Biophysics and Structural Biology*
*And William V. Power Chair in Biology (1991-2011), And Affiliate, Division of Cell and Developmental Biology
Transmembrane and intracellular signal transduction mechanisms are the focus of our group, especially understanding how extracellular stimuli control cell growth and division, cell morphology, and gene expression at the biochemical level.
Structural analysis of a peptide hormone receptor. To understand the molecular biology of peptide hormone action, we study response of budding yeast (Saccharomyces cerevisiae) to its peptide mating pheromones (a-factor and α-factor). The pheromone receptors have seven hydrophobic segments and are coupled to a heterotrimeric G protein. Receptors of this type are ubiquitous and transduce binding of a wide variety of extracellular ligands (peptide hormones, neurotransmitters and other bioactive compounds) into a physiological signal. Attempts to determine the three-dimensional structure of the purified α-factor receptor by NMR and X-ray crystallography are underway in collaborative studies. We are also interested in proteins involved in adaptation and recovery after the pheromone-induced signal, especially: structure and function of Sst2, the prototype RGS (Regulator of G-protein Signaling) protein; α-arrestins involved in selective ubiquitinylation and endocytosis of integral membrane proteins; and, various classes of phosphotyrosine-directed, phosphoserine- / phosphothreonine-directed, and dual-specificity phosphoprotein phosphatases that can act to dephosphorylate activated MAPKs.
Molecular genetics and biochemistry of MAPK cascades. Activation of the receptor-coupled G protein initiates a four-tiered cascade of protein kinases, ultimately resulting in stimulation of a messenger-activated protein kinase (MAPK) that translocates into the nucleus. A different developmental pathway (invasive or filamentous growth) triggered by nutrient limitation stimulates another MAPK. Subjecting cells to hyperosmotic conditions activates yet another MAPK. Thus, MAPK cascades are universally employed in eukaryotic cells for mounting an appropriate response to an extracellular stimulus and, like yeast, every eukaryotic cell contains multiple MAPK pathways. We are investigating the mechanisms that impose specificity and fidelity at each tier in these signaling networks. We showed that one device used by the cell for discrimination between parallel MAPK pathways is a specific docking interaction between a MAPK and the N-terminus of its cognate upstream protein kinase (MEK). We also showed that the scaffold protein Ste5 helps ensure signaling fidelity in pheromone response by binding the appropriate MAPK, MEK, and upstream activating kinase (MEKK) and by shuttling from the nucleus to the plasma membrane, thereby delivering the MAPK module to its most proximal activator (a fourth membrane-associated protein kinase). A different scaffold protein, Ste50, is required for signal propagation in the invasive growth and hyperosmotic stress response pathways and is also under study. The roles of other factors that prevent inappropriate MAPK activation in response to a given stimulus are also under study using genetic and biochemical methods.
Septin-associated protein kinases involved in cell proliferation, differentiation, and cell cycle control. The Wee1 class of protein-tyrosine kinase has an important role in cell cycle control. We are investigating pathways that regulate the activity, localization, and stability of this enzyme, and other bud-neck associated protein kinases (Gin4, Kcc4 and Hsl1), including their recruitment to septin filaments, which assemble at the presumptive site of cell division. We have shown that septin filaments are assembled from hetero-octameric complexes containing two each of four different septin subunits. A considerable effort is underway to understand the roles of other classes of protein kinases and additional post-translational modifications in septin complex assembly, formation of different septin-based supramolecular ensembles, disassembly of septin-containing structures, and the function of septin organization in the events required for cell division and membrane septation during cytokinesis. Our studies on septin ultrastructure are conducted in close collaboration with our colleague Prof. Eva Nogales in our Division (BBS) of MCB.
Protein kinases that control plasma membrane lipid homeostasis. We have shown that a cascade of protein kinases conserved from yeast to humans (including the TORC2 complex) is involved in maintenance of sphingolipid and glycerolipid homeostasis and bilayer lipid organization in the plasma membrane. This pathway and the mechanisms of its regulation are under intensive study.
Molecular biology of phosphoinositide-dependent signaling. In previous work, we have shown that phosphatidylinositol 4-phosphate generated by a specific phosphatidylinositol 4-kinase isoform has a specific role in the Golgi-to-plasma membrane stage of the secretory pathway. This enzyme also has an essential role in supplying the PtdIns(4)P that is converted to PtdIns(4,5)P2 in the nucleus, where it is hydrolyzed by a specific phospholipase C to generate inositol polyphosphates that regulate transcription, chromatin remodeling and mRNA export. We found that the phosphatidylinositol 4-kinase is also regulated by a small calcium-binding protein whose ortholog in animal cells (frequenin or neuronal calcium sensor-1) is found mainly in neuronal and neuroendocrine cells. We continue to explore the roles of these lipids in cell polarization, cell division and cell signaling.
Novel mechanisms for translocation across membranes. We discovered that export of a-factor pheromone requires an integral plasma membrane protein that is a dedicated ATP-dependent transporter, rather than the classical secretory pathway. The mechanism of transmembrane translocation of pheromone by the transporter (Ste6) is being examined. The functions of other members of the same transporter class, namely ABC transporters, have also been under study.
Muir A, Roelants FM, Timmons G, Thorner J (2015) Down-regulation of TORC2-Ypk1 signaling promotes MAPK-independent survival under hyperosmotic stress. Elife [E-published, 14 Aug 2015].
Finnigan GC, Thorner J (2015) Complex in vivo ligation using homologous recombination and high-efficiency plasmid rescue from Saccharomyces cerevisiae. Bio-protocol 5: e1521.1-e1521.14.
Finnigan GC, Booth EA, Duvalyan A, Liao EN, Thorner J (2015) The carboxy-terminal tails of septins Cdc11 and Shs1 recruit myosin-II binding factor Bni5 to the bud neck in Saccharomyces cerevisiae. Genetics 200: 843–862.
Finnigan GC, Takagi J, Cho C, Thorner J (2015) Comprehensive genetic analysis of paralogous terminal septin subunits Shs1 and Cdc11 in Saccharomyces cerevisiae. Genetics 200: 821–841.
Johnson CR, Weems AD, Brewer JM, Thorner J, McMurray MA (2015) Cytosolic chaperones mediate quality control of higher-order septin assembly in budding yeast. Mol. Biol. Cell 26: 1323-1344.
Roelants FM, Su BM, von Wulffen J, Ramachandran S, Sartorel E, Trott AE, Thorner J (2015) Protein kinase Gin4 negatively regulates flippase function and controls plasma membrane asymmetry. J. Cell Biol. 208: 299-311.
O’Donnell AF, McCartney RR, Chandrashekarappa DG, Zhang BB, Thorner J, Schmidt MC (2015) 2-Deoxyglucose impairs Saccharomyces cerevisiae growth by stimulating Snf1-regulated and alpha-arrestin-mediated trafficking of hexose transporters 1 and 3. Mol. Cell. Biol. 35: 939-955.
Sartorel E, Barrey E, Lau RK, Thorner J (2015) Plasma membrane aminoglycerolipid flippase function is required for signaling competence in the yeast mating pheromone response pathway. Mol. Biol. Cell 26: 134-150.
Thorner J, Hunter T, Cantley LC, Sever R (2014) Signal transduction: from the atomic age to the post-genomic era. Cold Spring Harb. Perspect. Biol. 6: a022913.1-a022913.17.
Muir A, Ramachandran S, Roelants FM, Timmons G, Thorner J (2014) TORC2-dependent protein kinase Ypk1 phosphorylates ceramide synthase to stimulate synthesis of complex sphingolipids. eLIFE 3: e03779.1-e03779.34.
Cantley LC, Hunter T, Sever R, Thorner J (eds.) Signal Transduction: Principles, Pathways, and Processes. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, 452pp., 2014.
Alvaro CG, O'Donnell AF, Prosser DC, Augustine AA, Goldman AR, Brodsky JL, Cyert MS, Wendland B, Thorner J (2014) Specific alpha-arrestins negatively regulate Saccharomyces cerevisiae pheromone response by down-modulating the G-protein coupled receptor Ste2. Mol. Cell. Biol. 34: 2660-2681.
O'Donnell AF, Huang L, Thorner J, Cyert MS (2013) A calcineurin-dependent switch controls the trafficking function of alpha-arrestin Aly1/Art6. J. Biol. Chem. 288: 24063-24080.
de Val N, McMurray MA, Lam LH, Hsiung C, Bertin A, Nogales E, Thorner J (2013) Native cysteine residues are dispensable for the structure and function of all five yeast mitotic septins. Proteins 81: 1964-1979.
Konopka JB, Thorner JW (2013) Pheromone Receptors (Yeast). In: Lennarz WJ, Lane MD (eds.) The Encyclopedia of Biological Chemistry, Vol. 3, pp. 441-446. Waltham, MA: Academic Press, Inc.
Lee YJ, Jeschke G, Roelants F, Thorner J, Turk B (2012) Reciprocal phosphorylation of yeast glycerol-3-phosphate dehydrogenases in adaptation to distinct types of stress. Mol. Cell. Biol. 32: 4705-4717.
Wu H-J, Henzie J, Lin W-C, Rhodes C, Li Z, Sartorel E, Thorner J, Yang P, Groves JT (2012) Membrane-protein binding measured with solution-phase plasmonic nanocube sensors. Nature Methods 9: 1189-1191.
Bertin A, McMurray MA, Pierson J, Thai L, McDonald KL, Zehr EA, Garcia G 3rd, Peters P, Thorner J, Nogales E (2012) Three-dimensional ultrastructure of the septin filament network in Saccharomyces cerevisiae. Mol. Biol. Cell 23: 423-432.
Roelants FM, Breslow DK, Muir A, Weissman JS, Thorner J (2011) Protein kinase Ypk1 phosphorylates Orm1 and Orm2 to control sphingolipid homeostasis in Saccharomyces cerevisiae. Proc. Natl. Acad. Sci. USA 108: 19222-19227.
Garcia G III, Bertin A, Li Z, Song Y, McMurray MA, Thorner J, Nogales E (2011) Subunit-dependent modulation of septin assembly: Budding yeast septin Shs1 promotes ring and gauze formation. J. Cell Biol. 195: 993-1004.
McMurray MA, Stefan CJ, Wemmer M, Odorizzi G, Emr SD, Thorner J (2011) Genetic interactions with mutations affecting septin assembly reveal ESCRT functions in budding yeast cytokinesis. Biol. Chem. 392: 699-712.
McMurray MA, Bertin A, Garcia G 3rd, Lam L, Nogales E, Thorner J (2011) Septin filament formation is essential in budding yeast. Dev. Cell 20: 540-549.
Lim S, Strahl T, Thorner J, Ames JB (2011) Structure of a Ca2+-myristoyl switch protein that controls activation of a phosphatidylinositol 4-kinase in fission yeast. J. Biol. Chem. 286: 12565-12577.
Patterson JC, Klimenko ES, Thorner J (2010) Single-cell analysis reveals that insulation maintains signaling specificity between two yeast MAPK pathways with common components. Sci. Sig. 3: ra75.1-ra75.11.
Bertin A, McMurray MA, Thai L, Garcia G III, Votin V, Grob P, Allyn T, Thorner J, Nogales E (2010) Phosphatidylinositol-4,5-bisphosphate promotes budding yeast septin filament assembly and organization. J. Mol. Biol. 404: 711-731.
Chen RE, Patterson JC, Goupil LS, Thorner J (2010) Dynamic localization of Fus3 MAPK is necessary to evoke appropriate responses and avoid cytotoxic effects. Mol. Cell. Biol. 30: 4293-4307.
Chen RE, Thorner J (2010) Systematic epistasis analysis of the contributions of PKA- and MAPK-dependent signaling to nutrient limitation-evoked responses in the yeast Saccharomyces cerevisiae. Genetics 185: 855-870.
Garrenton LS, Stefan CJ, McMurray MA, Emr SD, Thorner J (2010) Pheromone-induced anisotropy in yeast plasma membrane phosphatidylinositol-4,5-bisphosphate distribution is required for MAPK signaling. Proc. Natl. Acad. Sci. USA 107: 11805-11810.
Roelants FM, Baltz AG, Trott AE, Fereres S, Thorner J (2010) A protein kinase network regulates the function of aminophospholipid flippases. Proc. Natl. Acad. Sci. USA 107: 34-39.
Garrenton LS, Braunwarth A, Irniger S, Hurt E, Künzler M, Thorner J (2009) Nucleus-specific and cell cycle-regulated degradation of mitogen-activated protein kinase scaffold protein Ste5 contributes to the control of signaling competence. Mol. Cell. Biol. 29: 582-601.
McMurray MA, Thorner J (2009) Septins: molecular partitioning and the generation of cellular asymmetry. Cell Div. 4: 18.1-18.40.
McMurray MA, Thorner J (2009) Reuse, replace, recycle. Specificity in subunit inheritance and assembly of higher-order septin structures during mitotic and meiotic division in budding yeast. Cell Cycle 8: 195-203.
Rockwell NC, Wolfger H, Kuchler K, Thorner J (2009) ABC transporter Pdr10 regulates the membrane microenvironment of Pdr12 in Saccharomyces cerevisiae. J. Membr. Biol. 229: 27-52.
Westfall PJ, Patterson JC, Chen RE, Thorner J (2008) Stress resistance and signal fidelity independent of nuclear MAPK function. Proc. Natl. Acad. Sci. USA 105: 12212-12217.
McMurray MA, Thorner J (2008) Septin stability and recycling during dynamic structural transitions in cell division and development. Curr. Biol. 18: 1203-1208.
McMurray MA, Thorner J (2008) Biochemical properties and supramolecular architecture of septin hetero-oligomers and septin filaments, In "The Septins" (Hall PA, Russell SEG, Pringle JR, Eds.), John Wiley & Sons, Ltd., Chicester, West Sussex, UK, pp. 49-100.
Bertin A, McMurray MA, Grob P, Park SS, Garcia G 3rd, Patanwala I, Ng HL, Alber T, Thorner J, Nogales E (2008) Saccharomyces cerevisiae septins: supramolecular organization of heterooligomers and the mechanism of filament assembly. Proc. Natl. Acad. Sci. USA 105: 8274-8279.
Photo credit: Mark Hanson at Mark Joseph Studios.
Last Updated 2015-08-27