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Mouse monoclonal 12E8 antibody, which recognises conserved serine phosphorylated KXGS motifs in the microtubule binding domains of tau/tau-like microtubule associated proteins (MAPs), shows elevated binding in brain during normal embryonic development (mammals and birds) and at the early stages of human Alzheimer's disease (AD). It also labels ADF/cofilin-actin rods that form in neurites during exposure to stressors. We aimed to identify direct and indirect 12E8 binding proteins in postnatal mouse brain and embryonic chick brain by immunoprecipitation (IP), mass spectrometry and immunofluorescence. Tau and/or MAP2 were major direct 12E8-binding proteins detected in all IPs, and actin and/or tubulin were co-immunoprecipitated in most samples. Additional proteins were different in mouse versus chick brain IP. In mouse brain IPs, FSD1l and intermediate filament proteins - vimentin, α-internexin, neurofilament polypeptides - were prominent. Immunofluorescence and immunoblot using recombinant intermediate filament subunits, suggests an indirect interaction of these proteins with the 12E8 antibody. In chick brain IPs, subunits of eukaryotic translation initiation factor 3 (EIF3) were found, but no direct interaction between 12E8 and recombinant Eif3e protein was detected. Fluorescence microscopy in primary cultured chick neurons showed evidence of co-localisation of Eif3e and tubulin labelling, consistent with previous data demonstrating cytoskeletal organisation of the translation apparatus. Neither total tau or MAP2 immunolabelling accumulated at ADF/cofilin-actin rods generated in primary cultured chick neurons, and we were unable to narrow down the major antigen recognised by 12E8 antibody on ADF/cofilin-actin rods.

Competing Interests: Competing interests The authors declare no competing or financial interests.

Titel:	Protein complexes from mouse and chick brain that interact with phospho-KXGS motif tau/microtubule associated protein antibody.
Autor/in / Beteiligte Person:	Davies, DS ; Arthur, AT ; Aitken, HL ; Crossett, B ; Goldsbury, CS
Link:	Full Text Finder (Volltext)
Zeitschrift:	Biology open, Jg. 13 (2024-02-15), Heft 2
Veröffentlichung:	Cambridge : Company of Biologists Ltd., 2024
Medientyp:	academicJournal
ISSN:	2046-6390 (electronic)
DOI:	10.1242/bio.060067
Schlagwort:	Mice Animals Humans Actin Depolymerizing Factors metabolism Tubulin metabolism Brain metabolism Carrier Proteins metabolism Mammals metabolism Microtubule-Associated Proteins metabolism Actins metabolism
Sonstiges:	Nachgewiesen in: MEDLINE Sprachen: English Publication Type: Journal Article Language: English [Biol Open] 2024 Feb 15; Vol. 13 (2). <i>Date of Electronic Publication: </i>2024 Feb 27. MeSH Terms: Microtubule-Associated Proteins* / metabolism ; Actins* / metabolism ; Mice ; Animals ; Humans ; Actin Depolymerizing Factors / metabolism ; Tubulin / metabolism ; Brain / metabolism ; Carrier Proteins / metabolism ; Mammals / metabolism References: Cell. 2004 Mar 5;116(5):671-82. (PMID: 15006350) ; J Cell Sci. 2002 Sep 1;115(Pt 17):3389-402. (PMID: 12154070) ; Nat Cell Biol. 2007 Feb;9(2):139-48. (PMID: 17187063) ; Mol Cell. 2009 Oct 9;36(1):141-52. (PMID: 19818717) ; Amino Acids. 2011 Jul;41(2):363-85. (PMID: 20567863) ; Methods Mol Biol. 2017;1523:263-272. (PMID: 27975255) ; J Biol Chem. 1995 Aug 11;270(32):18917-22. (PMID: 7642549) ; PLoS One. 2018 Oct 16;13(10):e0198709. (PMID: 30325927) ; Mol Biol Cell. 2002 Nov;13(11):4013-28. (PMID: 12429843) ; Hum Mol Genet. 2014 Jan 1;23(1):104-16. (PMID: 23962722) ; J Neurosci. 2012 Jun 13;32(24):8270-83. (PMID: 22699908) ; J Biol Chem. 1995 Mar 31;270(13):7679-88. (PMID: 7706316) ; Mol Biol Cell. 2000 Oct;11(10):3573-87. (PMID: 11029056) ; Curr Biol. 2002 Nov 19;12(22):1946-51. (PMID: 12445389) ; Proc Natl Acad Sci U S A. 2022 Dec 13;119(50):e2211536119. (PMID: 36469763) ; J Neurosci. 2005 Dec 7;25(49):11313-21. (PMID: 16339026) ; Nat Cell Biol. 2000 Sep;2(9):628-36. (PMID: 10980704) ; Brain. 2020 Sep 1;143(9):2803-2817. (PMID: 32812023) ; Biochim Biophys Acta. 2001 Mar 19;1518(1-2):200-3. (PMID: 11267680) ; Trends Biochem Sci. 2013 Oct;38(10):494-506. (PMID: 24029419) ; Curr Alzheimer Res. 2010 May;7(3):241-50. (PMID: 20088812) ; FEBS Lett. 1996 Dec 16;399(3):344-9. (PMID: 8985176) ; Nat Rev Neurosci. 2016 Jan;17(1):5-21. (PMID: 26631930) ; PLoS One. 2011;6(6):e20878. (PMID: 21738590) ; Elife. 2017 Aug 02;6:. (PMID: 28767038) ; J Proteome Res. 2020 Jun 5;19(6):2429-2442. (PMID: 32357304) ; J Biol Chem. 1993 Oct 25;268(30):22695-702. (PMID: 8226779) ; J Neurosci. 2009 Oct 14;29(41):12994-3005. (PMID: 19828813) ; Int J Mol Sci. 2021 Aug 26;22(17):. (PMID: 34502116) ; Front Neurol. 2020 Oct 19;11:590059. (PMID: 33193056) ; Neural Dev. 2015 Feb 25;10:3. (PMID: 25886013) ; EMBO J. 2013 Nov 13;32(22):2920-37. (PMID: 24065130) ; Nat Rev Mol Cell Biol. 2010 Jan;11(1):75-81. (PMID: 20027187) ; Acta Neuropathol. 2002 Jan;103(1):26-35. (PMID: 11837744) ; Cell Motil Cytoskeleton. 2000 Mar;45(3):235-46. (PMID: 10706778) ; J Alzheimers Dis. 2014;42(4):1443-60. (PMID: 25024349) ; Biochemistry. 2002 Dec 24;41(51):15203-11. (PMID: 12484758) Grant Information: 2009/GNT0633269 Australian National Health and Medical Research Council; The University of Sydney Contributed Indexing: Keywords: ADF; Actin depolymerizing factor; Cofilin; Eif3; Eukaryotic translation initiation factor 3; FSD1l; KXGS motif; MAP2; Microtubule associated protein 2; Tau Substance Nomenclature: 0 (Microtubule-Associated Proteins) ; 0 (Actins) ; 0 (Actin Depolymerizing Factors) ; 0 (Tubulin) ; 0 (Carrier Proteins) Entry Date(s): Date Created: 20240201 Date Completed: 20240228 Latest Revision: 20240312 Update Code: 20240312 PubMed Central ID: PMC10924212

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Protein complexes from mouse and chick brain that interact with phospho-KXGS motif tau/microtubule associated protein antibody.