Reconstitution of constitutive secretion using semi-intact cells: regulation by GTP but not calcium - PubMed (original) (raw)
Reconstitution of constitutive secretion using semi-intact cells: regulation by GTP but not calcium
S G Miller et al. J Cell Biol. 1991 Jan.
Abstract
Regulated exocytosis in many permeabilized cells can be triggered by calcium and nonhydrolyzable GTP analogues. Here we examine the role of these effectors in exocytosis of constitutive vesicles using a system that reconstitutes transport between the trans-Golgi region and the plasma membrane. Transport is assayed by two independent methods: the movement of a transmembrane glycoprotein (vesicular stomatitis virus glycoprotein [VSV G protein]) to the cell surface; and the release of a soluble marker, sulfated glycosaminoglycan (GAG) chains, that have been synthesized and radiolabeled in the trans-Golgi. The plasma membrane of CHO cells was selectively perforated with the bacterial cytolysin streptolysin-O. These perforated cells allow exchange of ions and cytosolic proteins but retain intracellular organelles and transport vesicles. Incubation of the semi-intact cells with ATP and a cytosolic fraction results in transport of VSV G protein and GAG chains to the cell surface. The transport reaction is temperature dependent, requires hydrolyzable ATP, and is inhibited by N-ethylmaleimide. Nonhydrolyzable GTP analogs such as GTP gamma S, which stimulate the fusion of regulated secretory granules, completely abolish constitutive secretion. The rate and extent of constitutive transport between the trans-Golgi and the plasma membrane is independent of free Ca2+ concentrations. This is in marked contrast to fusion of regulated secretory granules with the plasma membrane, and transport between the ER and the cis-Golgi (Beckers, C. J. M., and W. E. Balch. 1989. J. Cell Biol. 108:1245-1256; Baker, D., L. Wuestehube, R. Schekman, and D. Botstein. 1990. Proc. Natl. Acad. Sci. USA. 87:355-359).
Similar articles
- Sequential intermediates in the transport of protein between the endoplasmic reticulum and the Golgi.
Beckers CJ, Plutner H, Davidson HW, Balch WE. Beckers CJ, et al. J Biol Chem. 1990 Oct 25;265(30):18298-310. J Biol Chem. 1990. PMID: 2120230 - Analysis of protein transport through the Golgi in a reconstituted cell-free system.
Wattenberg BW. Wattenberg BW. J Electron Microsc Tech. 1991 Feb;17(2):150-64. doi: 10.1002/jemt.1060170204. J Electron Microsc Tech. 1991. PMID: 1901603 Review. - The protein machinery of vesicle budding and fusion.
Rothman JE. Rothman JE. Protein Sci. 1996 Feb;5(2):185-94. doi: 10.1002/pro.5560050201. Protein Sci. 1996. PMID: 8745395 Free PMC article. Review.
Cited by
- TRIM21 and PHLDA3 negatively regulate the crosstalk between the PI3K/AKT pathway and PPP metabolism.
Cheng J, Huang Y, Zhang X, Yu Y, Wu S, Jiao J, Tran L, Zhang W, Liu R, Zhang L, Wang M, Wang M, Yan W, Wu Y, Chi F, Jiang P, Zhang X, Wu H. Cheng J, et al. Nat Commun. 2020 Apr 20;11(1):1880. doi: 10.1038/s41467-020-15819-3. Nat Commun. 2020. PMID: 32312982 Free PMC article. - Intracellular Delivery by Membrane Disruption: Mechanisms, Strategies, and Concepts.
Stewart MP, Langer R, Jensen KF. Stewart MP, et al. Chem Rev. 2018 Aug 22;118(16):7409-7531. doi: 10.1021/acs.chemrev.7b00678. Epub 2018 Jul 27. Chem Rev. 2018. PMID: 30052023 Free PMC article. Review. - Astrocyte VAMP3 vesicles undergo Ca2+ -independent cycling and modulate glutamate transporter trafficking.
Li D, Hérault K, Zylbersztejn K, Lauterbach MA, Guillon M, Oheim M, Ropert N. Li D, et al. J Physiol. 2015 Jul 1;593(13):2807-32. doi: 10.1113/JP270362. Epub 2015 May 18. J Physiol. 2015. PMID: 25864578 Free PMC article. - NAADP/TPC2/Ca(2+) Signaling Inhibits Autophagy.
Lu Y, Hao B, Graeff R, Yue J. Lu Y, et al. Commun Integr Biol. 2013 Nov 1;6(6):e27595. doi: 10.4161/cib.27595. Epub 2014 Jan 8. Commun Integr Biol. 2013. PMID: 24753792 Free PMC article. - Trafficking of the NMDAR2B receptor subunit distal cytoplasmic tail from endoplasmic reticulum to the synapse.
Standley S, Petralia RS, Gravell M, Hamilton R, Wang YX, Schubert M, Wenthold RJ. Standley S, et al. PLoS One. 2012;7(6):e39585. doi: 10.1371/journal.pone.0039585. Epub 2012 Jun 27. PLoS One. 2012. PMID: 22761831 Free PMC article.
References
- J Cell Biol. 1989 Apr;108(4):1245-56 - PubMed
- Cell. 1980 Sep;21(2):417-27 - PubMed
- Eur J Cell Biol. 1988 Dec;47(2):241-50 - PubMed
- Nature. 1989 Jun 1;339(6223):397-8 - PubMed
- Science. 1989 Aug 18;245(4919):718-25 - PubMed
Publication types
MeSH terms
Substances
LinkOut - more resources
Full Text Sources
Miscellaneous