Abstract
Co-immunoprecipitation is a traditional method but still one of the most powerful tools to identify novel interaction partners by increasing its specificity and decreasing false-positive result. In the current work, we show a robust procedure where the use of two sequential approaches (separation of postsynaptic density by centrifuge separation method in combination with detergent treatment and the use of specific antibodies against target proteins for its precipitation) improved the specificity of co-immunoprecipitation methods. Mass spectrometric analysis also helped to identify novel-associated proteins with target proteins without any bias of suspected interaction partners.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Rao VS, Srinivas K, Sujini GN, Kumar GN (2014) Protein-protein interaction detection: methods and analysis. Int J Proteomics 2014:147648. https://doi.org/10.1155/2014/147648
Speth C, Toledo-Filho LA, Laubinger S (2014) Immunoprecipitation-based analysis of protein-protein interactions. Methods Mol Biol 1158:175–185. https://doi.org/10.1007/978-1-4939-0700-7_11
Hunter RG, Murakami G, Dewell S, Seligsohn M, Baker ME, Datson NA, McEwen BS, Pfaff DW (2012) Acute stress and hippocampal histone H3 lysine 9 trimethylation, a retrotransposon silencing response. Proc Natl Acad Sci U S A 109(43):17657–17662. https://doi.org/10.1073/pnas.1215810109
Mukai H, Tsurugizawa T, Murakami G, Kominami S, Ishii H, Ogiue-Ikeda M, Takata N, Tanabe N, Furukawa A, Hojo Y, Ooishi Y, Morrison JH, Janssen WGM, Rose JA, Chambon P, Kato S, Izumi S, Yamazaki T, Kimoto T, Kawato S (2007) Rapid modulation of long-term depression and spinogenesis via synaptic estrogen receptors in hippocampal principal neurons. J Neurochem 100(4):950–967. https://doi.org/10.1111/j.1471-4159.2006.04264.x
Murakami G, Hojo Y, Ogiue-Ikeda M, Mukai H, Chambon P, Nakajima K, Ooishi Y, Kimoto T, Kawato S (2014) Estrogen receptor KO mice study on rapid modulation of spines and long-term depression in the hippocampus. Brain Res 1621:133–146. https://doi.org/10.1016/j.brainres.2014.12.002
Murakami G, Tsurugizawa T, Hatanaka Y, Komatsuzaki Y, Tanabe N, Mukai H, Hojo Y, Kominami S, Yamazaki T, Kimoto T, Kawato S (2006) Comparison between basal and apical dendritic spines in estrogen-induced rapid spinogenesis of CA1 principal neurons in the adult hippocampus. Biochem Biophys Res Commun 351(2):553–558
Carlin RK, Grab DJ, Cohen RS, Siekevitz P (1980) Isolation and characterization of postsynaptic densities from various brain regions: enrichment of different types of postsynaptic densities. J Cell Biol 86(3):831–845
Cohen RS, Blomberg F, Berzins K, Siekevitz P (1977) The structure of postsynaptic densities isolated from dog cerebral cortex. I. Overall morphology and protein composition. J Cell Biol 74(1):181–203
Whittaker VP (1984) The synaptosome. In: Handbook of neurochemistryn, vol 7, 2nd edn. Plenum Press, New York
Murakami G, Hunter RG, Fontaine C, Ribeiro A, Pfaff D (2011) Relationships among estrogen receptor, oxytocin and vasopressin gene expression and social interaction in male mice. Eur J Neurosci 34(3):469–477. https://doi.org/10.1111/j.1460-9568.2011.07761.x
Toran-Allerand CD, Guan X, MacLusky NJ, Horvath TL, Diano S, Singh M, Connolly ES Jr, Nethrapalli IS, Tinnikov AA (2002) ER-X: a novel, plasma membrane-associated, putative estrogen receptor that is regulated during development and after ischemic brain injury. J Neurosci 22(19):8391–8401
Fuxe K, Agnati LF, Borroto-Escuela DO (2014) The impact of receptor-receptor interactions in heteroreceptor complexes on brain plasticity. Expert Rev Neurother 14(7):719–721. https://doi.org/10.1586/14737175.2014.922878
Corriveau RA, Huh GS, Shatz CJ (1998) Regulation of class I MHC gene expression in the developing and mature CNS by neural activity. Neuron 21(3):505–520
Edamura M, Murakami G, Meng H, Itakura M, Shigemoto R, Fukuda A, Nakahara D (2014) Functional deficiency of MHC Class I enhances LTP and abolishes LTD in the nucleus accumbens of mice. PLoS One 9(9):e107099. https://doi.org/10.1371/journal.pone.0107099
Murakami G, Edamura M, Furukawa T, Kawasaki H, Kosugi I, Fukuda A, Iwashita T, Nakahara D (2018) MHC class I in dopaminergic neurons suppresses relapse to reward seeking. Sci Adv 4(3):eaap7388. https://doi.org/10.1126/sciadv.aap7388
Grieb B, Engler G, Sharott A, von Nicolai C, Streichert T, Papageorgiou I, Schulte A, Westphal M, Lamszus K, Engel AK, Moll CK, Hamel W (2014) High-frequency stimulation of the subthalamic nucleus counteracts cortical expression of major histocompatibility complex genes in a rat model of Parkinson's disease. PLoS One 9(3):e91663. https://doi.org/10.1371/journal.pone.0091663
Miyagawa T, Toyoda H, Hirataka A, Kanbayashi T, Imanishi A, Sagawa Y, Kotorii N, Kotorii T, Hashizume Y, Ogi K, Hiejima H, Kamei Y, Hida A, Miyamoto M, Imai M, Fujimura Y, Tamura Y, Ikegami A, Wada Y, Moriya S, Furuya H, Kato M, Omata N, Kojima H, Kashiwase K, Saji H, Khor SS, Yamasaki M, Ishigooka J, Kuroda K, Kume K, Chiba S, Yamada N, Okawa M, Hirata K, Uchimura N, Shimizu T, Inoue Y, Honda Y, Mishima K, Honda M, Tokunaga K (2015) New susceptibility variants to narcolepsy identified in HLA class II region. Hum Mol Genet 24(3):891–898. https://doi.org/10.1093/hmg/ddu480
Acknowledgments
We are grateful to Dr. Shiro Kominami for preparing RC-19 antibodies. This study was supported by Japan Society for the Promotion of Science (JSPS) KAKENHI (Grant Numbers JP2389076, JP25860392, JP15K19164). The authors report no biomedical financial interests or potential conflicts of interest.
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2019 Springer Science+Business Media, LLC, part of Springer Nature
About this protocol
Cite this protocol
Murakami, G., Kawato, S. (2019). Co-immunoprecipitation Methods to Identify Associated Proteins with Estrogen Receptor α at Postsynaptic Density in Brain Tissue. In: Odagaki, Y., Borroto-Escuela, D. (eds) Co-Immunoprecipitation Methods for Brain Tissue . Neuromethods, vol 144. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-8985-0_2
Download citation
DOI: https://doi.org/10.1007/978-1-4939-8985-0_2
Published:
Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-8984-3
Online ISBN: 978-1-4939-8985-0
eBook Packages: Springer Protocols