Enrichment of ER tubule-derived microsomes by differential centrifugation and immunoprecipitation

Yiduo Liu; Junjie Hu; Bing Yan

doi:10.52601/bpr.2023.230031

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Yiduo Liu, Junjie Hu, Bing Yan. Enrichment of ER tubule-derived microsomes by differential centrifugation and immunoprecipitation[J]. Biophysics Reports. doi: 10.52601/bpr.2023.230031

Citation:

Yiduo Liu, Junjie Hu, Bing Yan. Enrichment of ER tubule-derived microsomes by differential centrifugation and immunoprecipitation[J]. Biophysics Reports. doi: 10.52601/bpr.2023.230031

Yiduo Liu, Junjie Hu, Bing Yan. Enrichment of ER tubule-derived microsomes by differential centrifugation and immunoprecipitation[J]. Biophysics Reports. doi: 10.52601/bpr.2023.230031

Citation:

Yiduo Liu, Junjie Hu, Bing Yan. Enrichment of ER tubule-derived microsomes by differential centrifugation and immunoprecipitation[J]. Biophysics Reports. doi: 10.52601/bpr.2023.230031

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Enrichment of ER tubule-derived microsomes by differential centrifugation and immunoprecipitation

doi: 10.52601/bpr.2023.230031

Yiduo Liu^{1, 2},
Junjie Hu^{1, 2
,
,},
Bing Yan^{1
,
,}

1.
National Laboratory of Biomacromolecules, CAS Center for Excellence in Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China
2.
University of Chinese Academy of Sciences, Beijing 101408, China

More Information

Corresponding author: huj@ibp.ac.cn (J. Hu); bing_yan@ibp.ac.cn (B. Yan)
Received Date: 30 October 2023
Accepted Date: 15 December 2023

Available Online: 28 March 2024

Abstract

Abstract

The endoplasmic reticulum (ER) is an essential component of the endomembrane system in eukaryotes and plays a crucial role in protein and lipid synthesis, as well as the maintenance of calcium homeostasis. Morphologically, the ER is composed primarily of sheets and tubules. The tubular ER is composed of a network of tubular membrane structures, each with diameters ranging from 30 to 50 nanometers. In recent years, there has been in-depth research on the molecular mechanisms of membrane shaping and membrane fusion of the tubular ER. However, there is still limited understanding of the specific physiological functions of the tubular ER. Here, we report a protocol that combines differential centrifugation and immunoprecipitation to specifically enrich microsomes originating from the tubular ER in yeast. The ER tubule-derived microsomes can be further used for proteomic and lipidomic studies or other biochemical analyses.
- Endoplasmic reticulum,
- Microsome,
- Differential centrifugation,
- Immunoprecipitation

Yiduo Liu, Junjie Hu and Bing Yan declare that they have no conflict of interest.
This article does not contain any studies with human or animal subjects performed by any of the authors.

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References(17)

References

Anwar K, Klemm RW, Condon A, Severin KN, Zhang M, Ghirlando R, Hu J, Rapoport TA, Prinz WA (2012) The dynamin-like GTPase Sey1p mediates homotypic ER fusion in S. cerevisiae. J Cell Biol 197(2): 209−217 doi: 10.1083/jcb.201111115

Bian X, Klemm RW, Liu TY, Zhang M, Sun S, Sui X, Liu X, Rapoport TA, Hu J (2011) Structures of the atlastin GTPase provide insight into homotypic fusion of endoplasmic reticulum membranes. Proc Natl Acad Sci USA 108(10): 3976−3981 doi: 10.1073/pnas.1101643108

Griffing LR (2010) Networking in the endoplasmic reticulum. Biochem Soc Trans 38(3): 747−753 doi: 10.1042/BST0380747

Guillen-Samander A, De Camilli P (2023) Endoplasmic reticulum membrane contact sites, lipid transport, and neurodegeneration. Cold Spring Harb Perspect Biol 15(4): a041257. https://doi.org/10.1101/cshperspect.a041257

Hu J, Prinz WA, Rapoport TA (2011) Weaving the web of ER tubules. Cell 147(6): 1226−1231 doi: 10.1016/j.cell.2011.11.022

Hu J, Shibata Y, Voss C, Shemesh T, Li Z, Coughlin M, Kozlov MM, Rapoport TA, Prinz WA (2008) Membrane proteins of the endoplasmic reticulum induce high-curvature tubules. Science 319(5867): 1247−1250 doi: 10.1126/science.1153634

Hu J, Shibata Y, Zhu PP, Voss C, Rismanchi N, Prinz WA, Rapoport TA, Blackstone C (2009) A class of dynamin-like GTPases involved in the generation of the tubular ER network. Cell 138(3): 549−561 doi: 10.1016/j.cell.2009.05.025

Kriechbaumer V (2018) ER microsome preparation in Arabidopsis thaliana. Methods Mol Biol 1691: 117−123

Li J, Yan B, Si H, Peng X, Zhang SL, Hu J (2017) Atlastin regulates store-operated calcium entry for nerve growth factor-induced neurite outgrowth. Sci Rep 7: 43490. https://doi.org/10.1038/srep43490

Phillips MJ, Voeltz GK (2016) Structure and function of ER membrane contact sites with other organelles. Nat Rev Mol Cell Biol 17(2): 69−82 doi: 10.1038/nrm.2015.8

Salinas S, Proukakis C, Crosby A, Warner TT (2008) Hereditary spastic paraplegia: clinical features and pathogenetic mechanisms. Lancet Neurol 7(12): 1127−1138 doi: 10.1016/S1474-4422(08)70258-8

Shibata Y, Voss C, Rist JM, Hu J, Rapoport TA, Prinz WA, Voeltz GK (2008) The reticulon and DP1/Yop1p proteins form immobile oligomers in the tubular endoplasmic reticulum. J Biol Chem 283(27): 18892−18904 doi: 10.1074/jbc.M800986200

Voeltz GK, Prinz WA, Shibata Y, Rist JM, Rapoport TA (2006) A class of membrane proteins shaping the tubular endoplasmic reticulum. Cell 124(3): 573−586 doi: 10.1016/j.cell.2005.11.047

Wang X, Li S, Wang H, Shui W, Hu J (2017) Quantitative proteomics reveal proteins enriched in tubular endoplasmic reticulum of Saccharomyces cerevisiae. Elife 6: e23816. https://doi.org/10.7554/eLife.23816

Xiang Y, Lyu R, Hu J (2023) Oligomeric scaffolding for curvature generation by ER tubule-forming proteins. Nat Commun 14(1): 2617. https://doi.org/10.1038/s41467-023-38294-y

Yan L, Sun S, Wang W, Shi J, Hu X, Wang S, Su D, Rao Z, Hu J, Lou Z (2015) Structures of the yeast dynamin-like GTPase Sey1p provide insight into homotypic ER fusion. J Cell Biol 210(6): 961−972 doi: 10.1083/jcb.201502078

Zhang M, Wu F, Shi J, Zhu Y, Zhu Z, Gong Q, Hu J (2013) ROOT HAIR DEFECTIVE3 family of dynamin-like GTPases mediates homotypic endoplasmic reticulum fusion and is essential for Arabidopsis development. Plant Physiol 163(2): 713−720 doi: 10.1104/pp.113.224501

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