Abstract
Among posttranslational modifications, the phosphorylation of tyrosine residues is a key modification in cell signaling. Because of its biological importance, characterization of the cellular state of tyrosine phosphorylation is of great interest. Based on the unique properties of endogenously expressed SH2 domains recognizing tyrosine phosphorylated signaling proteins with high specificity we have developed an alternative approach, coined SH2 profiling, enabling us to decipher complex patterns of tyrosine phosphorylation in various normal and cancerous tissues. So far, SH2 profiling has largely been applied for the analysis of protein extracts with the limitation that information on spatial distribution and intensity of tyrosine phosphorylation within a tissue is lost. Here, we describe a novel SH2 domain based strategy for differential characterization of the state of tyrosine phosphorylation in formaldehyde-fixed and paraffin-embedded tissues. This approach demonstrates that SH2 domains may serve as very valuable tools for the analysis of the differential state of tyrosine phosphorylation in primary tissues fixed and processed under conditions frequently applied by routine pathology laboratories.
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References
Machida K, Mayer BJ, Nollau P (2003) Profiling the global tyrosine phosphorylation state. Mol Cell Proteomics 2(4):215–233. doi:10.1074/mcp.R300002-MCP200
Hunter T (2009) Tyrosine phosphorylation: thirty years and counting. Curr Opin Cell Biol 21(2):140–146. doi:10.1016/j.ceb.2009.01.028
Machida K, Khenkhar M, Nollau P (2012) Deciphering phosphotyrosine-dependent signaling networks in cancer by SH2 profiling. Genes Cancer 3(5–6):353–361. doi:10.1177/1947601912459048
Liu BA, Jablonowski K, Raina M, Arce M, Pawson T, Nash PD (2006) The human and mouse complement of SH2 domain proteins-establishing the boundaries of phosphotyrosine signaling. Mol Cell 22(6):851–868. doi:10.1016/j.molcel.2006.06.001
Tinti M, Kiemer L, Costa S, Miller ML, Sacco F, Olsen JV, Carducci M, Paoluzi S, Langone F, Workman CT, Blom N, Machida K, Thompson CM, Schutkowski M, Brunak S, Mann M, Mayer BJ, Castagnoli L, Cesareni G (2013) The SH2 domain interaction landscape. Cell Rep 3(4):1293–1305. doi:10.1016/j.celrep.2013.03.001
Dierck K, Machida K, Voigt A, Thimm J, Horstmann M, Fiedler W, Mayer BJ, Nollau P (2006) Quantitative multiplexed profiling of cellular signaling networks using phosphotyrosine-specific DNA-tagged SH2 domains. Nat Methods 3(9):737–744. doi:10.1038/nmeth917
Machida K, Thompson CM, Dierck K, Jablonowski K, Karkkainen S, Liu B, Zhang H, Nash PD, Newman DK, Nollau P, Pawson T, Renkema GH, Saksela K, Schiller MR, Shin DG, Mayer BJ (2007) High-throughput phosphotyrosine profiling using SH2 domains. Mol Cell 26(6):899–915. doi:10.1016/j.molcel.2007.05.031
Jadwin JA, Mayer BJ, Machida K (2015) Detection and quantification of protein-protein interactions by far-western blotting. Methods Mol Biol 1312:379–398. doi:10.1007/978-1-4939-2694-7_38
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Buhs, S., Nollau, P. (2017). SH2 Domain Histochemistry. In: Machida, K., Liu, B. (eds) SH2 Domains. Methods in Molecular Biology, vol 1555. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-6762-9_31
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DOI: https://doi.org/10.1007/978-1-4939-6762-9_31
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Publisher Name: Humana Press, New York, NY
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