pmc logo image
Logo of jcellbiolHomeThe Rockefeller University PressEditorsContactInstructions for AuthorsThis issue
Journal List > J Cell Biol > v.118(6); Sep 2, 1992

Formats:
J Cell Biol. 1992 September 2; 118(6): 1321–1332.
PMCID: PMC2289609
Copyright notice
The activity of Golgi transport vesicles depends on the presence of the N-ethylmaleimide-sensitive factor (NSF) and a soluble NSF attachment protein (alpha SNAP) during vesicle formation
Small right arrow pointing to: This article has been cited by other articles in PMC.
Abstract
An assay designed to measure the formation of functional transport vesicles was constructed by modifying a cell-free assay for protein transport between compartments of the Golgi (Balch, W. E., W. G. Dunphy, W. A. Braell, and J. E. Rothman. 1984. Cell. 39:405-416). A 35- kD cytosolic protein that is immunologically and functionally indistinguishable from alpha SNAP (soluble NSF attachment protein) was found to be required during vesicle formation. SNAP, together with the N-ethylmaleimide-sensitive factor (NSF) have previously been implicated in the attachment and/or fusion of vesicles with their target membrane. We show that NSF is also required during the formation of functional vesicles. Strikingly, we found that after vesicle formation, the NEM- sensitive function of NSF was no longer required for transport to proceed through the ensuing steps of vesicle attachment and fusion. In contrast to these functional tests of vesicle formation, SNAP was not required for the morphological appearance of vesicular structures on the Golgi membranes. If SNAP and NSF have a direct role in transport vesicle attachment and/or fusion, as previously suggested, these results indicate that these proteins become incorporated into the vesicle membranes during vesicle formation and are brought to the fusion site on the transport vesicles.
Full Text
The Full Text of this article is available as a PDF (2.1M).
Selected References
These references are in PubMed. This may not be the complete list of references from this article.
  • Balch WE. Biochemistry of interorganelle transport. A new frontier in enzymology emerges from versatile in vitro model systems. J Biol Chem. 1989 Oct 15;264(29):16965–16968. [PubMed]
  • Balch WE, Dunphy WG, Braell WA, Rothman JE. Reconstitution of the transport of protein between successive compartments of the Golgi measured by the coupled incorporation of N-acetylglucosamine. Cell. 1984 Dec;39(2 Pt 1):405–416. [PubMed]
  • Balch WE, Glick BS, Rothman JE. Sequential intermediates in the pathway of intercompartmental transport in a cell-free system. Cell. 1984 Dec;39(3 Pt 2):525–536. [PubMed]
  • Block MR, Glick BS, Wilcox CA, Wieland FT, Rothman JE. Purification of an N-ethylmaleimide-sensitive protein catalyzing vesicular transport. Proc Natl Acad Sci U S A. 1988 Nov;85(21):7852–7856. [PubMed]
  • Clary DO, Rothman JE. Purification of three related peripheral membrane proteins needed for vesicular transport. J Biol Chem. 1990 Jun 15;265(17):10109–10117. [PubMed]
  • Clary DO, Griff IC, Rothman JE. SNAPs, a family of NSF attachment proteins involved in intracellular membrane fusion in animals and yeast. Cell. 1990 May 18;61(4):709–721. [PubMed]
  • Diaz R, Mayorga LS, Weidman PJ, Rothman JE, Stahl PD. Vesicle fusion following receptor-mediated endocytosis requires a protein active in Golgi transport. Nature. 1989 Jun 1;339(6223):398–400. [PubMed]
  • Fries E, Rothman JE. Transport of vesicular stomatitis virus glycoprotein in a cell-free extract. Proc Natl Acad Sci U S A. 1980 Jul;77(7):3870–3874. [PubMed]
  • Goda Y, Pfeffer SR. Cell-free systems to study vesicular transport along the secretory and endocytic pathways. FASEB J. 1989 Nov;3(13):2488–2495. [PubMed]
  • Goud B, Salminen A, Walworth NC, Novick PJ. A GTP-binding protein required for secretion rapidly associates with secretory vesicles and the plasma membrane in yeast. Cell. 1988 Jun 3;53(5):753–768. [PubMed]
  • Hicke L, Schekman R. Molecular machinery required for protein transport from the endoplasmic reticulum to the Golgi complex. Bioessays. 1990 Jun;12(6):253–258. [PubMed]
  • Hiebsch RR, Raub TJ, Wattenberg BW. Primaquine blocks transport by inhibiting the formation of functional transport vesicles. Studies in a cell-free assay of protein transport through the Golgi apparatus. J Biol Chem. 1991 Oct 25;266(30):20323–20328. [PubMed]
  • Kaiser CA, Schekman R. Distinct sets of SEC genes govern transport vesicle formation and fusion early in the secretory pathway. Cell. 1990 May 18;61(4):723–733. [PubMed]
  • Laemmli UK. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. [PubMed]
  • Malhotra V, Orci L, Glick BS, Block MR, Rothman JE. 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):221–227. [PubMed]
  • Melançon P, Glick BS, Malhotra V, Weidman PJ, Serafini T, Gleason ML, Orci L, Rothman JE. Involvement of GTP-binding "G" proteins in transport through the Golgi stack. Cell. 1987 Dec 24;51(6):1053–1062. [PubMed]
  • Mellman I, Simons K. The Golgi complex: in vitro veritas? Cell. 1992 Mar 6;68(5):829–840. [PubMed]
  • Orci L, Glick BS, Rothman JE. 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):171–184. [PubMed]
  • Orci L, Malhotra V, Amherdt M, Serafini T, Rothman JE. Dissection of a single round of vesicular transport: sequential intermediates for intercisternal movement in the Golgi stack. Cell. 1989 Feb 10;56(3):357–368. [PubMed]
  • Rexach MF, Schekman RW. Distinct biochemical requirements for the budding, targeting, and fusion of ER-derived transport vesicles. J Cell Biol. 1991 Jul;114(2):219–229. [PubMed]
  • Rothman JE, Orci L. Molecular dissection of the secretory pathway. Nature. 1992 Jan 30;355(6359):409–415. [PubMed]
  • Rothman JE, Urbani LJ, Brands R. Transport of protein between cytoplasmic membranes of fused cells: correspondence to processes reconstituted in a cell-free system. J Cell Biol. 1984 Jul;99(1 Pt 1):248–259. [PubMed]
  • Schekman R. Protein localization and membrane traffic in yeast. Annu Rev Cell Biol. 1985;1:115–143. [PubMed]
  • Stanley P, Narasimhan S, Siminovitch L, Schachter H. Chinese hamster ovary cells selected for resistance to the cytotoxicity of phytohemagglutinin are deficient in a UDP-N-acetylglucosamine--glycoprotein N-acetylglucosaminyltransferase activity. Proc Natl Acad Sci U S A. 1975 Sep;72(9):3323–3327. [PubMed]
  • Wattenberg BW. The molecular control of transport vesicle fusion. New Biol. 1990 Jun;2(6):505–511. [PubMed]
  • Wattenberg BW, Balch WE, Rothman JE. A novel prefusion complex formed during protein transport between Golgi cisternae in a cell-free system. J Biol Chem. 1986 Feb 15;261(5):2202–2207. [PubMed]
  • Wattenberg BW, Hiebsch RR, LeCureux LW, White MP. Identification of a 25-kD protein from yeast cytosol that operates in a prefusion step of vesicular transport between compartments of the Golgi. J Cell Biol. 1990 Apr;110(4):947–954. [PubMed]
  • Weidman PJ, Melançon P, Block MR, Rothman JE. Binding of an N-ethylmaleimide-sensitive fusion protein to Golgi membranes requires both a soluble protein(s) and an integral membrane receptor. J Cell Biol. 1989 May;108(5):1589–1596. [PubMed]
  • Wilson DW, Whiteheart SW, Wiedmann M, Brunner M, Rothman JE. A multisubunit particle implicated in membrane fusion. J Cell Biol. 1992 May;117(3):531–538. [PubMed]
Articles from The Journal of Cell Biology are provided here courtesy of
The Rockefeller University Press