Home LiteratureArticle Details
PMID: 1907974 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't

Reconstitution of GTP-binding Sar1 protein function in ER to Golgi transport.

The Journal of cell biology ·Vol. 114 ·No. 4 ·1991-08-00 ·Pages 671-9

Oka T, Nishikawa S, Nakano A

Abstract

In the yeast secretory pathway, two genes SEC12 and SAR1, which encode a 70-kD integral membrane protein and a 21-kD GTP-binding protein, respectively, cooperate in protein transport from the ER to the Golgi apparatus. In vivo, the elevation of the SAR1 dosage suppresses temperature sensitivity of the sec12 mutant. In this paper, we show cell-free reconstitution of the ER-to-Golgi transport that depends on both of these gene products. First, the membranes from the sec12 mutant cells reproduce temperature sensitivity in the in vitro ER-to-Golgi transport reaction. Furthermore, the addition of the Sar1 protein completely suppresses this temperature-sensitive defect of the sec12 membranes. The analysis of Sar1p partially purified by E. coli expression suggests that GTP hydrolysis is essential for Sar1p to execute its function.

MeSH Terms
Cloning, Molecular Electrophoresis, Polyacrylamide Gel Endoplasmic Reticulum/metabolism Escherichia coli/genetics GTP-Binding Proteins/metabolism Golgi Apparatus/metabolism Guanosine 5'-O-(3-Thiotriphosphate)/pharmacology Guanosine Triphosphate/metabolism Kinetics Molecular Weight Protein Biosynthesis Recombinant Proteins/isolation & purification,metabolism Saccharomyces cerevisiae/genetics,growth & development,metabolism Temperature Transcription, Genetic
Chemicals
Recombinant Proteins Guanosine 5'-O-(3-Thiotriphosphate) Guanosine Triphosphate GTP-Binding Proteins
Authors & Affiliations
3 authors, click to expand affiliations / ORCID
Oka T
Department of Biology, Faculty of Science, University of Tokyo, Japan.
Nishikawa S
Nakano A
References (39)
39 references, click to expand
  1. Study of a temperature-sensitive mutant of the ras-related YPT1 gene product in yeast suggests a role in the regulation of intracellular calcium.
    Cell. 1988 May 20;53(4):635-47 PMID: 3286011
  2. A study of the immunochemistry of three yeast mannans.
    J Biol Chem. 1970 Mar 10;245(5):1197-203 PMID: 5461381
  3. A ras-like protein is required for a post-Golgi event in yeast secretion.
    Cell. 1987 May 22;49(4):527-38 PMID: 3552249
  4. Secretion in yeast: reconstitution of the translocation and glycosylation of alpha-factor and invertase in a homologous cell-free system.
    Cell. 1986 Feb 28;44(4):619-28 PMID: 3512097
  5. Specific binding of [alpha-32P]GTP to cytosolic and membrane-bound proteins of human platelets correlates with the activation of phospholipase C.
    Proc Natl Acad Sci U S A. 1987 Apr;84(8):2261-5 PMID: 3470789
  6. Isolation of a yeast gene, SRH1, that encodes a homologue of the 54K subunit of mammalian signal recognition particle.
    J Biochem. 1990 Mar;107(3):457-63 PMID: 2187859
  7. Molecular cloning of the microtubule-associated mechanochemical enzyme dynamin reveals homology with a new family of GTP-binding proteins.
    Nature. 1990 Sep 20;347(6290):256-61 PMID: 2144893
  8. Enzymatic coupling of cholesterol intermediates to a mating pheromone precursor and to the ras protein.
    Science. 1990 Sep 7;249(4973):1133-9 PMID: 2204115
  9. Sec12p-dependent membrane binding of the small GTP-binding protein Sar1p promotes formation of transport vesicles from the ER.
    J Cell Biol. 1991 Aug;114(4):663-70 PMID: 1907973
  10. The GTPase superfamily: conserved structure and molecular mechanism.
    Nature. 1991 Jan 10;349(6305):117-27 PMID: 1898771
  11. The GTPase superfamily: a conserved switch for diverse cell functions.
    Nature. 1990 Nov 8;348(6297):125-32 PMID: 2122258
  12. Mutants of Saccharomyces cerevisiae defective in the farnesylation of Ras proteins.
    Proc Natl Acad Sci U S A. 1990 Dec;87(24):9665-9 PMID: 2124698
  13. Distinct biochemical requirements for the budding, targeting, and fusion of ER-derived transport vesicles.
    J Cell Biol. 1991 Jul;114(2):219-29 PMID: 1649197
  14. A novel GTP-binding protein, Sar1p, is involved in transport from the endoplasmic reticulum to the Golgi apparatus.
    J Cell Biol. 1989 Dec;109(6 Pt 1):2677-91 PMID: 2512296
  15. The GTP-binding protein Ypt1 is required for transport in vitro: the Golgi apparatus is defective in ypt1 mutants.
    J Cell Biol. 1989 Sep;109(3):1015-22 PMID: 2504726
  16. Mutational analysis of SEC4 suggests a cyclical mechanism for the regulation of vesicular traffic.
    EMBO J. 1989 Jun;8(6):1685-93 PMID: 2504585
  17. GTP-binding domain: three consensus sequence elements with distinct spacing.
    Proc Natl Acad Sci U S A. 1987 Apr;84(7):1814-8 PMID: 3104905
  18. Myristoylated alpha subunits of guanine nucleotide-binding regulatory proteins.
    Proc Natl Acad Sci U S A. 1987 Nov;84(21):7493-7 PMID: 3118369
  19. The yeast GTP-binding YPT1 protein and a mammalian counterpart are associated with the secretion machinery.
    Cell. 1988 Mar 25;52(6):915-24 PMID: 3127057
  20. Sequences of the bovine and yeast ADP-ribosylation factor and comparison to other GTP-binding proteins.
    Proc Natl Acad Sci U S A. 1988 Jul;85(13):4620-4 PMID: 3133654
  21. A carboxyl-terminal cysteine residue is required for palmitic acid binding and biological activity of the ras-related yeast YPT1 protein.
    EMBO J. 1988 Apr;7(4):971-6 PMID: 3042385
  22. Saccharomyces cerevisiae and Schizosaccharomyces pombe contain a homologue to the 54-kD subunit of the signal recognition particle that in S. cerevisiae is essential for growth.
    J Cell Biol. 1989 Dec;109(6 Pt 2):3223-30 PMID: 2557350
  23. In vitro protein translocation across the yeast endoplasmic reticulum: ATP-dependent posttranslational translocation of the prepro-alpha-factor.
    Cell. 1986 May 9;45(3):397-406 PMID: 3009026
  24. A membrane glycoprotein, Sec12p, required for protein transport from the endoplasmic reticulum to the Golgi apparatus in yeast.
    J Cell Biol. 1988 Sep;107(3):851-63 PMID: 3047151
  25. Reconstitution of protein transport from the endoplasmic reticulum to the Golgi complex in yeast: the acceptor Golgi compartment is defective in the sec23 mutant.
    J Cell Biol. 1988 Oct;107(4):1465-76 PMID: 3049622
  26. Reconstitution of SEC gene product-dependent intercompartmental protein transport.
    Cell. 1988 Jul 29;54(3):335-44 PMID: 3293799
  27. Enzymes required for yeast prohormone processing.
    Annu Rev Physiol. 1988;50:345-62 PMID: 3288097
  28. Refined crystal structure of the triphosphate conformation of H-ras p21 at 1.35 A resolution: implications for the mechanism of GTP hydrolysis.
    EMBO J. 1990 Aug;9(8):2351-9 PMID: 2196171
  29. ADP-ribosylation factor is functionally and physically associated with the Golgi complex.
    Proc Natl Acad Sci U S A. 1990 Feb;87(3):1238-42 PMID: 2105501
  30. A putative GTP binding protein homologous to interferon-inducible Mx proteins performs an essential function in yeast protein sorting.
    Cell. 1990 Jun 15;61(6):1063-74 PMID: 2112425
  31. Time-resolved X-ray crystallographic study of the conformational change in Ha-Ras p21 protein on GTP hydrolysis.
    Nature. 1990 May 24;345(6273):309-15 PMID: 2111463
  32. Use of T7 RNA polymerase to direct expression of cloned genes.
    Methods Enzymol. 1990;185:60-89 PMID: 2199796
  33. Complementation of two overlapping fragments of SecA, a protein translocation ATPase of Escherichia coli, allows ATP binding to its amino-terminal region.
    J Biol Chem. 1990 May 25;265(15):8760-5 PMID: 2160468
  34. GTP-binding Ypt1 protein and Ca2+ function independently in a cell-free protein transport reaction.
    Proc Natl Acad Sci U S A. 1990 Jan;87(1):355-9 PMID: 2104983
  35. Isolation of a functional vesicular intermediate that mediates ER to Golgi transport in yeast.
    J Cell Biol. 1990 Jul;111(1):45-53 PMID: 2195039
  36. Distinct sets of SEC genes govern transport vesicle formation and fusion early in the secretory pathway.
    Cell. 1990 May 18;61(4):723-33 PMID: 2188733
  37. Identification of 23 complementation groups required for post-translational events in the yeast secretory pathway.
    Cell. 1980 Aug;21(1):205-15 PMID: 6996832
  38. The role of guanosine 5'-triphosphate in polypeptide chain elongation.
    Biochim Biophys Acta. 1978 Sep 21;505(1):95-127 PMID: 361078
  39. SEC62 encodes a putative membrane protein required for protein translocation into the yeast endoplasmic reticulum.
    J Cell Biol. 1989 Dec;109(6 Pt 1):2653-64 PMID: 2687286
Article Info
Journal
The Journal of cell biology
Abbr.
J Cell Biol
ISSN
0021-9525
DOI
10.1083/jcb.114.4.671
Published
1991-08-00
Pages
671-9
Language
English
Region
United States
NLM ID
0375356
PMCID
PMC2289892
Subset
IM
External Links
PubMed source DOI: 10.1083/jcb.114.4.671 PMC Full Text View journal details
Analysis Services
Analysis Services

Contact

No. 2 Wenbo Road, Zhangqiu District, Jinan, Shandong

Qilu Normal University · Genelibs Bioinformatics Lab

750 Shunhua Rd, Jinan

2F, Bldg F, University Science Park

Tel: 0531-88819269

WeChat Official Account

Follow our WeChat subscription account for real-time updates and the latest in medical and biological research.


Business Email

E-mail: product@genelibs.com

Copyright © 2026.Genelibs Jinan Honghui Biotechnology Co., Ltd. 鲁ICP备20007159号-2