Benjamin Glick

Research Summary
Our goal is to understand the processes that generate compartments of the secretory pathway, including ER exit sites (ERES; also known as transitional ER or tER sites) and the cisternae of the Golgi apparatus. Self-organization models provide the conceptual framework. Specifically, we postulate that ERES are generated together with early Golgi cisternae by an integrated self-organization pathway, and that early cisternae progressively mature into late cisternae. For exploring these ideas, our main experimental system is a pair of budding yeasts. In Saccharomyces cerevisiae, Golgi cisternae are dispersed throughout the cytoplasm and the ER contains multiple small ERES, whereas in Pichia pastoris, ordered Golgi stacks are located next to large, stable ERES. These two yeasts have complementary advantages for testing specific hypotheses about the secretory pathway. We use a combination of genetics, molecular biology, 4D confocal microscopy, and electron tomography. This work is revealing evolutionarily conserved principles of cellular organization. A second project in the lab involves optimizing fluorescent proteins, including the red fluorescent protein DsRed. Wild-type DsRed matures very slowly. We overcame this problem by using directed evolution to create the first rapidly maturing DsRed variants, one of which is marketed commercially as DsRed-Express. More recent work yielded a noncytotoxic variant called DsRed-Express2, as well as a far-red variant called E2-Crimson. These engineering efforts inspired a basic research project in which we clarified the pathway of DsRed chromophore formation. Current efforts are focused on creating improved monomeric green and red fluorescent proteins.
Organelles, Membranes, Cell, Microscopy, Fluorescence, Microscopy, Electron
  • Amherst College, Amherst, MA, BA Neuroscience / Mathematics 05/1983
  • Stanford University, Stanford, CA, PhD Biochemistry 09/1988
  • Biozentrum, University of Basel, Basel, Switzerland, (postdoc) Cell Biology 12/1994
Biosciences Graduate Program Association
  1. Editorial overview: Membranes in perpetual motion. Curr Opin Cell Biol. 2022 Oct; 78:102128. View in: PubMed

  2. Clathrin adaptors mediate two sequential pathways of intra-Golgi recycling. J Cell Biol. 2022 01 03; 221(1). View in: PubMed

  3. Activity-dependent Golgi satellite formation in dendrites reshapes the neuronal surface glycoproteome. Elife. 2021 09 21; 10. View in: PubMed

  4. A General Method to Improve Fluorophores Using Deuterated Auxochromes. JACS Au. 2021 May 24; 1(5):690-696. View in: PubMed

  5. TRAPP structures reveal the big picture. EMBO J. 2021 06 15; 40(12):e108537. View in: PubMed

  6. Acetyl-CoA flux from the cytosol to the ER regulates engagement and quality of the secretory pathway. Sci Rep. 2021 01 21; 11(1):2013. View in: PubMed

  7. ESCargo: a regulatable fluorescent secretory cargo for diverse model organisms. Mol Biol Cell. 2020 12 15; 31(26):2892-2903. View in: PubMed

  8. Bioreactor-scale cell performance and protein production can be substantially increased by using a secretion signal that drives co-translational translocation in Pichia pastoris. N Biotechnol. 2021 Jan 25; 60:85-95. View in: PubMed

  9. A microscopy-based kinetic analysis of yeast vacuolar protein sorting. Elife. 2020 06 25; 9. View in: PubMed

  10. A photostable monomeric superfolder green fluorescent protein. Traffic. 2020 08; 21(8):534-544. View in: PubMed

  11. ER arrival sites associate with ER exit sites to create bidirectional transport portals. J Cell Biol. 2020 04 06; 219(4). View in: PubMed

  12. A Kinetic View of Membrane Traffic Pathways Can Transcend the Classical View of Golgi Compartments. Front Cell Dev Biol. 2019; 7:153. View in: PubMed

  13. 4D Microscopy of Yeast. J Vis Exp. 2019 04 28; (146). View in: PubMed

  14. Maturation-driven transport and AP-1-dependent recycling of a secretory cargo in the Golgi. J Cell Biol. 2019 05 06; 218(5):1582-1601. View in: PubMed

  15. Visualizing Secretory Cargo Transport in Budding Yeast. Curr Protoc Cell Biol. 2019 06; 83(1):e80. View in: PubMed

  16. An improved secretion signal enhances the secretion of model proteins from Pichia pastoris. Microb Cell Fact. 2018 Oct 12; 17(1):161. View in: PubMed

  17. Budding Yeast Has a Minimal Endomembrane System. Dev Cell. 2018 01 08; 44(1):56-72.e4. View in: PubMed

  18. Improved deconvolution of very weak confocal signals. F1000Res. 2017; 6:787. View in: PubMed

  19. New insights into protein secretion: TANGO1 runs rings around the COPII coat. J Cell Biol. 2017 04 03; 216(4):859-861. View in: PubMed

  20. An improved reversibly dimerizing mutant of the FK506-binding protein FKBP. Cell Logist. 2016 Jul-Sep; 6(3):e1204848. View in: PubMed

  21. 4D Confocal Imaging of Yeast Organelles. Methods Mol Biol. 2016; 1496:1-11. View in: PubMed

  22. Refined Pichia pastoris reference genome sequence. J Biotechnol. 2016 Oct 10; 235:121-31. View in: PubMed

  23. The Atg17-Atg31-Atg29 Complex Coordinates with Atg11 to Recruit the Vam7 SNARE and Mediate Autophagosome-Vacuole Fusion. Curr Biol. 2016 Jan 25; 26(2):150-160. View in: PubMed

  24. COPI selectively drives maturation of the early Golgi. Elife. 2015 Dec 28; 4. View in: PubMed

  25. Gottfried Schatz (1936-2015)-mitochondrial pioneer and ambassador for science. EMBO J. 2015 Nov 12; 34(22):2725-6. View in: PubMed

  26. GenoLIB: a database of biological parts derived from a library of common plasmid features. Nucleic Acids Res. 2015 May 26; 43(10):4823-32. View in: PubMed

  27. Secretion of a foreign protein from budding yeasts is enhanced by cotranslational translocation and by suppression of vacuolar targeting. Microb Cell Fact. 2014 Aug 28; 13(1):125. View in: PubMed

  28. Golgi compartmentation and identity. Curr Opin Cell Biol. 2014 Aug; 29:74-81. View in: PubMed

  29. Integrated self-organization of transitional ER and early Golgi compartments. Bioessays. 2014 Feb; 36(2):129-33. View in: PubMed

  30. Golgi enlargement in Arf-depleted yeast cells is due to altered dynamics of cisternal maturation. J Cell Sci. 2014 Jan 01; 127(Pt 1):250-7. View in: PubMed

  31. Sec16 influences transitional ER sites by regulating rather than organizing COPII. Mol Biol Cell. 2013 Nov; 24(21):3406-19. View in: PubMed

  32. A three-stage model of Golgi structure and function. Histochem Cell Biol. 2013 Sep; 140(3):239-49. View in: PubMed

  33. Sec12 binds to Sec16 at transitional ER sites. PLoS One. 2012; 7(2):e31156. View in: PubMed

  34. Models for Golgi traffic: a critical assessment. Cold Spring Harb Perspect Biol. 2011 Nov 01; 3(11):a005215. View in: PubMed

  35. Organelle structure and biogenesis. Mol Biol Cell. 2011 Mar 15; 22(6):723. View in: PubMed

  36. Noncytotoxic DsRed derivatives for whole-cell labeling. Methods Mol Biol. 2011; 699:355-70. View in: PubMed

  37. The yeast GRASP Grh1 colocalizes with COPII and is dispensable for organizing the secretory pathway. Traffic. 2010 Sep; 11(9):1168-79. View in: PubMed

  38. Chromophore formation in DsRed occurs by a branched pathway. J Am Chem Soc. 2010 Jun 23; 132(24):8496-505. View in: PubMed

  39. High-quality immunofluorescence of cultured cells. Methods Mol Biol. 2010; 619:403-10. View in: PubMed

  40. Journeys through the Golgi--taking stock in a new era. J Cell Biol. 2009 Nov 16; 187(4):449-53. View in: PubMed

  41. The yeast Golgi apparatus: insights and mysteries. FEBS Lett. 2009 Dec 03; 583(23):3746-51. View in: PubMed

  42. A rapidly maturing far-red derivative of DsRed-Express2 for whole-cell labeling. Biochemistry. 2009 Sep 08; 48(35):8279-81. View in: PubMed

  43. Membrane traffic within the Golgi apparatus. Annu Rev Cell Dev Biol. 2009; 25:113-32. View in: PubMed

  44. Mutation of SYNE-1, encoding an essential component of the nuclear lamina, is responsible for autosomal recessive arthrogryposis. Hum Mol Genet. 2009 Sep 15; 18(18):3462-9. View in: PubMed

  45. Noncytotoxic orange and red/green derivatives of DsRed-Express2 for whole-cell labeling. BMC Biotechnol. 2009 Apr 03; 9:32. View in: PubMed

  46. A noncytotoxic DsRed variant for whole-cell labeling. Nat Methods. 2008 Nov; 5(11):955-7. View in: PubMed

  47. Cdc1p is an endoplasmic reticulum-localized putative lipid phosphatase that affects Golgi inheritance and actin polarization by activating Ca2+ signaling. Mol Cell Biol. 2008 May; 28(10):3336-43. View in: PubMed

  48. Structural rearrangements near the chromophore influence the maturation speed and brightness of DsRed variants. Protein Eng Des Sel. 2007 Nov; 20(11):525-34. View in: PubMed

  49. Fluorescence microscopy and thin-section electron microscopy. Methods Mol Biol. 2007; 389:251-60. View in: PubMed

  50. Identification of pexophagy genes by restriction enzyme-mediated integration. Methods Mol Biol. 2007; 389:203-18. View in: PubMed

  51. GRASPing unconventional secretion. Cell. 2007 Aug 10; 130(3):407-9. View in: PubMed

  52. Let there be order. Nat Cell Biol. 2007 Feb; 9(2):130-2. View in: PubMed

  53. Two mammalian Sec16 homologues have nonredundant functions in endoplasmic reticulum (ER) export and transitional ER organization. Mol Biol Cell. 2007 Mar; 18(3):839-49. View in: PubMed

  54. MICA: desktop software for comprehensive searching of DNA databases. BMC Bioinformatics. 2006 Oct 03; 7:427. View in: PubMed

  55. The budding yeast Pichia pastoris has a novel Sec23p homolog. FEBS Lett. 2006 Oct 02; 580(22):5215-21. View in: PubMed

  56. Golgi maturation visualized in living yeast. Nature. 2006 Jun 22; 441(7096):1002-6. View in: PubMed

  57. Brighter reporter genes from multimerized fluorescent proteins. Biotechniques. 2005 Dec; 39(6):814, 816, 818 passim. View in: PubMed

  58. Imaging pancreatic beta-cells in the intact pancreas. Am J Physiol Endocrinol Metab. 2006 May; 290(5):E1041-7. View in: PubMed

  59. Sec16 is a determinant of transitional ER organization. Curr Biol. 2005 Aug 23; 15(16):1439-47. View in: PubMed

  60. Golgi inheritance in small buds of Saccharomyces cerevisiae is linked to endoplasmic reticulum inheritance. Proc Natl Acad Sci U S A. 2004 Dec 28; 101(52):18018-23. View in: PubMed

  61. The transitional ER localization mechanism of Pichia pastoris Sec12. Dev Cell. 2004 May; 6(5):649-59. View in: PubMed

  62. Monitoring changes in the subcellular location of proteins in S. cerevisiae. Methods Mol Biol. 2004; 241:299-311. View in: PubMed

  63. The mechanisms of vesicle budding and fusion. Cell. 2004 Jan 23; 116(2):153-66. View in: PubMed

  64. Protein import into mitochondria: two systems acting in tandem? Trends Cell Biol. 1991 Oct; 1(4):99-103. View in: PubMed

  65. Tomographic evidence for continuous turnover of Golgi cisternae in Pichia pastoris. Mol Biol Cell. 2003 Jun; 14(6):2277-91. View in: PubMed

  66. De novo formation of transitional ER sites and Golgi structures in Pichia pastoris. Nat Cell Biol. 2002 Oct; 4(10):750-6. View in: PubMed

  67. Can the Golgi form de novo? Nat Rev Mol Cell Biol. 2002 08; 3(8):615-9. View in: PubMed

  68. Paz2 and 13 other PAZ gene products regulate vacuolar engulfment of peroxisomes during micropexophagy. Genes Cells. 2002 Jan; 7(1):75-90. View in: PubMed

  69. Tagging Hansenula polymorpha genes by random integration of linear DNA fragments (RALF). Mol Genet Genomics. 2001 Dec; 266(4):646-56. View in: PubMed

  70. Rapidly maturing variants of the Discosoma red fluorescent protein (DsRed). Nat Biotechnol. 2002 Jan; 20(1):83-7. View in: PubMed

  71. Deconstructing Golgi inheritance. Traffic. 2001 Sep; 2(9):589-96. View in: PubMed

  72. Vector for pop-in/pop-out gene replacement in Pichia pastoris. Biotechniques. 2001 Aug; 31(2):306-10, 312. View in: PubMed

  73. ER export: more than one way out. Curr Biol. 2001 May 01; 11(9):R361-3. View in: PubMed

  74. A role for actin, Cdc1p, and Myo2p in the inheritance of late Golgi elements in Saccharomyces cerevisiae. J Cell Biol. 2001 Apr 02; 153(1):47-62. View in: PubMed

  75. Barreling through the outer membrane. Nat Struct Biol. 2001 Apr; 8(4):284-6. View in: PubMed

  76. Raising the speed limits for 4D fluorescence microscopy. Traffic. 2000 Dec; 1(12):935-40. View in: PubMed

  77. Dynamics of transitional endoplasmic reticulum sites in vertebrate cells. Mol Biol Cell. 2000 Sep; 11(9):3013-30. View in: PubMed

  78. Isolation of Pichia pastoris genes involved in ER-to-Golgi transport. Yeast. 2000 Aug; 16(11):979-93. View in: PubMed

  79. Organization of the Golgi apparatus. Curr Opin Cell Biol. 2000 Aug; 12(4):450-6. View in: PubMed

  80. Golgi structure correlates with transitional endoplasmic reticulum organization in Pichia pastoris and Saccharomyces cerevisiae. J Cell Biol. 1999 Apr 05; 145(1):69-81. View in: PubMed

  81. The curious status of the Golgi apparatus. Cell. 1998 Dec 23; 95(7):883-9. View in: PubMed

  82. A yeast t-SNARE involved in endocytosis. Mol Biol Cell. 1998 Oct; 9(10):2873-89. View in: PubMed

  83. A versatile set of vectors for constitutive and regulated gene expression in Pichia pastoris. Yeast. 1998 Jun 15; 14(8):783-90. View in: PubMed

  84. Strong precursor-pore interactions constrain models for mitochondrial protein import. Biophys J. 1998 Apr; 74(4):1732-43. View in: PubMed

  85. Active unfolding of precursor proteins during mitochondrial protein import. EMBO J. 1997 Nov 17; 16(22):6727-36. View in: PubMed

  86. A cisternal maturation mechanism can explain the asymmetry of the Golgi stack. FEBS Lett. 1997 Sep 08; 414(2):177-81. View in: PubMed

  87. What is the driving force for protein import into mitochondria? Biochim Biophys Acta. 1997 Jan 16; 1318(1-2):71-8. View in: PubMed

  88. Cell biology: alternatives to baker's yeast. Curr Biol. 1996 Dec 01; 6(12):1570-2. View in: PubMed

  89. Saccharomyces cerevisiae mitochondria lack a bacterial-type sec machinery. Protein Sci. 1996 Dec; 5(12):2651-2. View in: PubMed

  90. Translocation arrest of an intramitochondrial sorting signal next to Tim11 at the inner-membrane import site. Nature. 1996 Dec 12; 384(6609):585-8. View in: PubMed

  91. The mitochondrial protein import motor: dissociation of mitochondrial hsp70 from its membrane anchor requires ATP binding rather than ATP hydrolysis. Protein Sci. 1996 Apr; 5(4):759-67. View in: PubMed

  92. Hsp60-independent protein folding in the matrix of yeast mitochondria. EMBO J. 1996 Feb 15; 15(4):764-74. View in: PubMed

  93. Pathways and energetics of mitochondrial protein import in Saccharomyces cerevisiae. Methods Enzymol. 1995; 260:224-31. View in: PubMed

  94. Isolation of highly purified mitochondria from Saccharomyces cerevisiae. Methods Enzymol. 1995; 260:213-23. View in: PubMed

  95. Import of cytochrome b2 to the mitochondrial intermembrane space: the tightly folded heme-binding domain makes import dependent upon matrix ATP. Protein Sci. 1993 Nov; 2(11):1901-17. View in: PubMed

  96. A mitochondrial homolog of bacterial GrpE interacts with mitochondrial hsp70 and is essential for viability. EMBO J. 1994 Apr 15; 13(8):1998-2006. View in: PubMed

  97. Protein import into mitochondria: the requirement for external ATP is precursor-specific whereas intramitochondrial ATP is universally needed for translocation into the matrix. Mol Biol Cell. 1994 Apr; 5(4):465-74. View in: PubMed

  98. Fusion proteins containing the cytochrome b2 presequence are sorted to the mitochondrial intermembrane space independently of hsp60. J Biol Chem. 1994 Jun 24; 269(25):17279-88. View in: PubMed

  99. Cloning and disruption of the gene encoding yeast mitochondrial chaperonin 10, the homolog of E. coli groES. FEBS Lett. 1993 Dec 13; 335(3):358-60. View in: PubMed

  100. Identification and functional analysis of chaperonin 10, the groES homolog from yeast mitochondria. Proc Natl Acad Sci U S A. 1993 Dec 01; 90(23):10967-71. View in: PubMed

  101. Can Hsp70 proteins act as force-generating motors? Cell. 1995 Jan 13; 80(1):11-4. View in: PubMed

  102. Dynamic interaction between Isp45 and mitochondrial hsp70 in the protein import system of the yeast mitochondrial inner membrane. Proc Natl Acad Sci U S A. 1994 Dec 20; 91(26):12818-22. View in: PubMed

  103. A C-terminally-anchored Golgi protein is inserted into the endoplasmic reticulum and then transported to the Golgi apparatus. Proc Natl Acad Sci U S A. 1995 May 23; 92(11):5102-5. View in: PubMed

  104. The protein import receptor of mitochondria. Trends Biochem Sci. 1995 Mar; 20(3):98-101. View in: PubMed

  105. Cyclophilin catalyzes protein folding in yeast mitochondria. Proc Natl Acad Sci U S A. 1995 Jul 03; 92(14):6319-23. View in: PubMed

  106. Sequential intermediates in the pathway of intercompartmental transport in a cell-free system. Cell. 1984 Dec; 39(3 Pt 2):525-36. View in: PubMed

  107. Structure and expression of amplified cKi-ras gene sequences in Y1 mouse adrenal tumor cells. EMBO J. 1985 May; 4(5):1199-203. View in: PubMed

  108. Possible role for fatty acyl-coenzyme A in intracellular protein transport. Nature. 1987 Mar 19-25; 326(6110):309-12. View in: PubMed

  109. Yeast and mammals utilize similar cytosolic components to drive protein transport through the Golgi complex. Proc Natl Acad Sci U S A. 1986 Mar; 83(6):1622-6. View in: PubMed

  110. Role of an N-ethylmaleimide-sensitive transport component in promoting fusion of transport vesicles with cisternae of the Golgi stack. Cell. 1988 Jul 15; 54(2):221-7. View in: PubMed

  111. Purification of an N-ethylmaleimide-sensitive protein catalyzing vesicular transport. Proc Natl Acad Sci U S A. 1988 Nov; 85(21):7852-6. View in: PubMed

  112. Components responsible for transport between successive Golgi cisternae are highly conserved in evolution. J Biol Chem. 1986 Apr 05; 261(10):4367-70. View in: PubMed

  113. A new type of coated vesicular carrier that appears not to contain clathrin: its possible role in protein transport within the Golgi stack. Cell. 1986 Jul 18; 46(2):171-84. View in: PubMed

  114. Involvement of GTP-binding "G" proteins in transport through the Golgi stack. Cell. 1987 Dec 24; 51(6):1053-62. View in: PubMed

  115. Fatty acyl-coenzyme A is required for budding of transport vesicles from Golgi cisternae. Cell. 1989 Oct 06; 59(1):95-102. View in: PubMed

  116. Vesicular transport between the endoplasmic reticulum and the Golgi stack requires the NEM-sensitive fusion protein. Nature. 1989 Jun 01; 339(6223):397-8. View in: PubMed

  117. A role for GTP-binding proteins in vesicular transport through the Golgi complex. Soc Gen Physiol Ser. 1989; 44:175-88. View in: PubMed

  118. Fatty acylation promotes fusion of transport vesicles with Golgi cisternae. J Cell Biol. 1990 Apr; 110(4):955-61. View in: PubMed

  119. Mitochondrial protein import. J Bioenerg Biomembr. 1990 Dec; 22(6):725-51. View in: PubMed

  120. Protein import into isolated yeast mitochondria. Methods Cell Biol. 1991; 34:389-99. View in: PubMed

  121. Import of proteins into mitochondria. Annu Rev Genet. 1991; 25:21-44. View in: PubMed

  122. The energetics of protein import into mitochondria. Biochim Biophys Acta. 1992 Jul 17; 1101(2):249-51. View in: PubMed

  123. Protein sorting in mitochondria. Trends Biochem Sci. 1992 Nov; 17(11):453-9. View in: PubMed

  124. Cytochromes c1 and b2 are sorted to the intermembrane space of yeast mitochondria by a stop-transfer mechanism. Cell. 1992 May 29; 69(5):809-22. View in: PubMed

  125. Role of ATP in the intramitochondrial sorting of cytochrome c1 and the adenine nucleotide translocator. EMBO J. 1992 Dec; 11(13):4787-94. View in: PubMed