Skip to main content
PMC home page
Plant Physiology logoLink to Plant Physiology
. 1997 May;114(1):315–324. doi: 10.1104/pp.114.1.315

Characterization of AtSEC12 and AtSAR1. Proteins likely involved in endoplasmic reticulum and Golgi transport.

M Bar-Peled 1, N V Raikhel 1
PMCID: PMC158307  PMID: 9159953

Abstract

Transport of cargo proteins from the endoplasmic reticulum (ER) to the cis-Golgi network is mediated by protein-coated vesicles. The coat, called COPII coat, consists of proteins that are recruited from the cytosol and interact with integral membrane proteins of the ER. In yeast, both cytosolic proteins (Sec13/31, Sec23/24, and Sar1) and ER-associated proteins (Sec12 and others) have been purified and characterized and it has been possible to demonstrate transport vesicle formation in vitro. Arabidopsis thaliana homologs of Sar1 and Sec12 have recently been identified, but little is known about the properties of the proteins or their subcellular distribution. Here we demonstrate that AtSAR1, a 22-kD protein that binds GTP, and AtSEC12, a 43-kD GTP-exchange protein, are both associated with the ER. However, about one-half of the cellular AtSAR1 is present in the cytosol. When AtSAR1 is overexpressed in transgenic plants, the additional protein is also cytosolic. When tissue-culture cells are cold-shocked (12 h at 8 degrees C), AtSAR1 levels appeared to decline and a larger proportion of the total protein was found in the cytosol. Given the known function of AtSAR1 in yeast, we propose that the amount of ER-associated AtSAR1 is an indication of the intensity of the secretory process. Thus, we expect that such a cold shock will adversely affect ER-to-Golgi transport of proteins.

Full Text

The Full Text of this article is available as a PDF (3.7 MB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Ahmed S. U., Bar-Peled M., Raikhel N. V. Cloning and subcellular location of an Arabidopsis receptor-like protein that shares common features with protein-sorting receptors of eukaryotic cells. Plant Physiol. 1997 May;114(1):325–336. doi: 10.1104/pp.114.1.325. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Aridor M., Bannykh S. I., Rowe T., Balch W. E. Sequential coupling between COPII and COPI vesicle coats in endoplasmic reticulum to Golgi transport. J Cell Biol. 1995 Nov;131(4):875–893. doi: 10.1083/jcb.131.4.875. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bar-Peled M., Conceicao AdS., Frigerio L., Raikhel N. V. Expression and Regulation of aERD2, a Gene Encoding the KDEL Receptor Homolog in Plants, and Other Genes Encoding Proteins Involved in ER-Golgi Vesicular Trafficking. Plant Cell. 1995 Jun;7(6):667–676. doi: 10.1105/tpc.7.6.667. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Bar-Peled M., Lewinsohn E., Fluhr R., Gressel J. UDP-rhamnose:flavanone-7-O-glucoside-2''-O-rhamnosyltransferase. Purification and characterization of an enzyme catalyzing the production of bitter compounds in citrus. J Biol Chem. 1991 Nov 5;266(31):20953–20959. [PubMed] [Google Scholar]
  5. Bar-Peled M., Raikhel N. V. A method for isolation and purification of specific antibodies to a protein fused to the GST. Anal Biochem. 1996 Oct 1;241(1):140–142. doi: 10.1006/abio.1996.0390. [DOI] [PubMed] [Google Scholar]
  6. Barlowe C., Orci L., Yeung T., Hosobuchi M., Hamamoto S., Salama N., Rexach M. F., Ravazzola M., Amherdt M., Schekman R. COPII: a membrane coat formed by Sec proteins that drive vesicle budding from the endoplasmic reticulum. Cell. 1994 Jun 17;77(6):895–907. doi: 10.1016/0092-8674(94)90138-4. [DOI] [PubMed] [Google Scholar]
  7. Barlowe C., Schekman R. SEC12 encodes a guanine-nucleotide-exchange factor essential for transport vesicle budding from the ER. Nature. 1993 Sep 23;365(6444):347–349. doi: 10.1038/365347a0. [DOI] [PubMed] [Google Scholar]
  8. Barlowe C., d'Enfert C., Schekman R. Purification and characterization of SAR1p, a small GTP-binding protein required for transport vesicle formation from the endoplasmic reticulum. J Biol Chem. 1993 Jan 15;268(2):873–879. [PubMed] [Google Scholar]
  9. Bednarek S. Y., Reynolds T. L., Schroeder M., Grabowski R., Hengst L., Gallwitz D., Raikhel N. V. A small GTP-binding protein from Arabidopsis thaliana functionally complements the yeast YPT6 null mutant. Plant Physiol. 1994 Feb;104(2):591–596. doi: 10.1104/pp.104.2.591. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Bradford M. M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976 May 7;72:248–254. doi: 10.1006/abio.1976.9999. [DOI] [PubMed] [Google Scholar]
  11. Cheon C. I., Lee N. G., Siddique A. B., Bal A. K., Verma D. P. Roles of plant homologs of Rab1p and Rab7p in the biogenesis of the peribacteroid membrane, a subcellular compartment formed de novo during root nodule symbiosis. EMBO J. 1993 Nov;12(11):4125–4135. doi: 10.1002/j.1460-2075.1993.tb06096.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Daniels M. J., Mirkov T. E., Chrispeels M. J. The plasma membrane of Arabidopsis thaliana contains a mercury-insensitive aquaporin that is a homolog of the tonoplast water channel protein TIP. Plant Physiol. 1994 Dec;106(4):1325–1333. doi: 10.1104/pp.106.4.1325. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Davies C. Cloning and characterization of a tomato GTPase-like gene related to yeast and Arabidopsis genes involved in vesicular transport. Plant Mol Biol. 1994 Feb;24(3):525–531. doi: 10.1007/BF00024120. [DOI] [PubMed] [Google Scholar]
  14. Dombrowski J. E., Raikhel N. V. Isolation of a cDNA encoding a novel GTP-binding protein of Arabidopsis thaliana. Plant Mol Biol. 1995 Sep;28(6):1121–1126. doi: 10.1007/BF00032672. [DOI] [PubMed] [Google Scholar]
  15. Elazar Z., Orci L., Ostermann J., Amherdt M., Tanigawa G., Rothman J. E. ADP-ribosylation factor and coatomer couple fusion to vesicle budding. J Cell Biol. 1994 Feb;124(4):415–424. doi: 10.1083/jcb.124.4.415. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Ferro-Novick S., Novick P. The role of GTP-binding proteins in transport along the exocytic pathway. Annu Rev Cell Biol. 1993;9:575–599. doi: 10.1146/annurev.cb.09.110193.003043. [DOI] [PubMed] [Google Scholar]
  17. Fujiki Y., Hubbard A. L., Fowler S., Lazarow P. B. Isolation of intracellular membranes by means of sodium carbonate treatment: application to endoplasmic reticulum. J Cell Biol. 1982 Apr;93(1):97–102. doi: 10.1083/jcb.93.1.97. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Gu X., Verma D. P. Phragmoplastin, a dynamin-like protein associated with cell plate formation in plants. EMBO J. 1996 Feb 15;15(4):695–704. [PMC free article] [PubMed] [Google Scholar]
  19. Guan K. L., Dixon J. E. Eukaryotic proteins expressed in Escherichia coli: an improved thrombin cleavage and purification procedure of fusion proteins with glutathione S-transferase. Anal Biochem. 1991 Feb 1;192(2):262–267. doi: 10.1016/0003-2697(91)90534-z. [DOI] [PubMed] [Google Scholar]
  20. Hicks G. R., Smith H. M., Lobreaux S., Raikhel N. V. Nuclear import in permeabilized protoplasts from higher plants has unique features. Plant Cell. 1996 Aug;8(8):1337–1352. doi: 10.1105/tpc.8.8.1337. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Höfte H., Hubbard L., Reizer J., Ludevid D., Herman E. M., Chrispeels M. J. Vegetative and Seed-Specific Forms of Tonoplast Intrinsic Protein in the Vacuolar Membrane of Arabidopsis thaliana. Plant Physiol. 1992 Jun;99(2):561–570. doi: 10.1104/pp.99.2.561. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Jedd G., Richardson C., Litt R., Segev N. The Ypt1 GTPase is essential for the first two steps of the yeast secretory pathway. J Cell Biol. 1995 Nov;131(3):583–590. doi: 10.1083/jcb.131.3.583. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Kuge O., Dascher C., Orci L., Rowe T., Amherdt M., Plutner H., Ravazzola M., Tanigawa G., Rothman J. E., Balch W. E. Sar1 promotes vesicle budding from the endoplasmic reticulum but not Golgi compartments. J Cell Biol. 1994 Apr;125(1):51–65. doi: 10.1083/jcb.125.1.51. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Loraine A. E., Yalovsky S., Fabry S., Gruissem W. Tomato Rab1A homologs as molecular tools for studying Rab geranylgeranyl transferase in plant cells. Plant Physiol. 1996 Apr;110(4):1337–1347. doi: 10.1104/pp.110.4.1337. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Nakańo A., Muramatsu M. 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):2677–2691. doi: 10.1083/jcb.109.6.2677. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Nishikawa S., Nakano A. The GTP-binding Sar1 protein is localized to the early compartment of the yeast secretory pathway. Biochim Biophys Acta. 1991 Jul 10;1093(2-3):135–143. doi: 10.1016/0167-4889(91)90114-d. [DOI] [PubMed] [Google Scholar]
  27. Oka T., Nakano A. Inhibition of GTP hydrolysis by Sar1p causes accumulation of vesicles that are a functional intermediate of the ER-to-Golgi transport in yeast. J Cell Biol. 1994 Feb;124(4):425–434. doi: 10.1083/jcb.124.4.425. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Palade G. Intracellular aspects of the process of protein synthesis. Science. 1975 Aug 1;189(4200):347–358. doi: 10.1126/science.1096303. [DOI] [PubMed] [Google Scholar]
  29. Park Y. S., Song O., Kwak J. M., Hong S. W., Lee H. H., Nam H. G. Functional complementation of a yeast vesicular transport mutation ypt1-1 by a Brassica napus cDNA clone encoding a small GTP-binding protein. Plant Mol Biol. 1994 Dec;26(6):1725–1735. doi: 10.1007/BF00019487. [DOI] [PubMed] [Google Scholar]
  30. Preuss D., Mulholland J., Kaiser C. A., Orlean P., Albright C., Rose M. D., Robbins P. W., Botstein D. Structure of the yeast endoplasmic reticulum: localization of ER proteins using immunofluorescence and immunoelectron microscopy. Yeast. 1991 Dec;7(9):891–911. doi: 10.1002/yea.320070902. [DOI] [PubMed] [Google Scholar]
  31. Regad F., Bardet C., Tremousaygue D., Moisan A., Lescure B., Axelos M. cDNA cloning and expression of an Arabidopsis GTP-binding protein of the ARF family. FEBS Lett. 1993 Jan 25;316(2):133–136. doi: 10.1016/0014-5793(93)81201-a. [DOI] [PubMed] [Google Scholar]
  32. Rexach M. F., Schekman R. W. Distinct biochemical requirements for the budding, targeting, and fusion of ER-derived transport vesicles. J Cell Biol. 1991 Jul;114(2):219–229. doi: 10.1083/jcb.114.2.219. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Rothman J. E. The protein machinery of vesicle budding and fusion. Protein Sci. 1996 Feb;5(2):185–194. doi: 10.1002/pro.5560050201. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Salama N. R., Schekman R. W. The role of coat proteins in the biosynthesis of secretory proteins. Curr Opin Cell Biol. 1995 Aug;7(4):536–543. doi: 10.1016/0955-0674(95)80011-5. [DOI] [PubMed] [Google Scholar]
  35. Salama N. R., Yeung T., Schekman R. W. The Sec13p complex and reconstitution of vesicle budding from the ER with purified cytosolic proteins. EMBO J. 1993 Nov;12(11):4073–4082. doi: 10.1002/j.1460-2075.1993.tb06091.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Saraste J., Kuismanen E. Pre- and post-Golgi vacuoles operate in the transport of Semliki Forest virus membrane glycoproteins to the cell surface. Cell. 1984 Sep;38(2):535–549. doi: 10.1016/0092-8674(84)90508-7. [DOI] [PubMed] [Google Scholar]
  37. Serafini T., Orci L., Amherdt M., Brunner M., Kahn R. A., Rothman J. E. ADP-ribosylation factor is a subunit of the coat of Golgi-derived COP-coated vesicles: a novel role for a GTP-binding protein. Cell. 1991 Oct 18;67(2):239–253. doi: 10.1016/0092-8674(91)90176-y. [DOI] [PubMed] [Google Scholar]
  38. Strambio-de-Castillia C., Blobel G., Rout M. P. Isolation and characterization of nuclear envelopes from the yeast Saccharomyces. J Cell Biol. 1995 Oct;131(1):19–31. doi: 10.1083/jcb.131.1.19. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Søgaard M., Tani K., Ye R. R., Geromanos S., Tempst P., Kirchhausen T., Rothman J. E., Söllner T. A rab protein is required for the assembly of SNARE complexes in the docking of transport vesicles. Cell. 1994 Sep 23;78(6):937–948. doi: 10.1016/0092-8674(94)90270-4. [DOI] [PubMed] [Google Scholar]
  40. Yoshihisa T., Barlowe C., Schekman R. Requirement for a GTPase-activating protein in vesicle budding from the endoplasmic reticulum. Science. 1993 Mar 5;259(5100):1466–1468. doi: 10.1126/science.8451644. [DOI] [PubMed] [Google Scholar]
  41. d'Enfert C., Barlowe C., Nishikawa S., Nakano A., Schekman R. Structural and functional dissection of a membrane glycoprotein required for vesicle budding from the endoplasmic reticulum. Mol Cell Biol. 1991 Nov;11(11):5727–5734. doi: 10.1128/mcb.11.11.5727. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. d'Enfert C., Gensse M., Gaillardin C. Fission yeast and a plant have functional homologues of the Sar1 and Sec12 proteins involved in ER to Golgi traffic in budding yeast. EMBO J. 1992 Nov;11(11):4205–4211. doi: 10.1002/j.1460-2075.1992.tb05514.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. d'Enfert C., Wuestehube L. J., Lila T., Schekman R. 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):663–670. doi: 10.1083/jcb.114.4.663. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. da Silva Conceiço A., Marty-Mazars D., Bassham D. C., Sanderfoot A. A., Marty F., Raikhel N. V. The syntaxin homolog AtPEP12p resides on a late post-Golgi compartment in plants. Plant Cell. 1997 Apr;9(4):571–582. [PMC free article] [PubMed] [Google Scholar]

Articles from Plant Physiology are provided here courtesy of Oxford University Press

RESOURCES