Direct Imaging of Protein-Specific Methylation in Mammalian Cells.
Franziska DollRaphael R SteimbachAndreas ZumbuschPublished in: Chembiochem : a European journal of chemical biology (2019)
Abundant post-translational modification through methylation alters the function, stability, and/or localization of a protein. Malfunctions in post-translational modification are associated with severe diseases. To unravel protein methylation sites and their biological functions, chemical methylation reporters have been developed. However, until now, their usage was limited to cell lysates. Herein, we present the first generally applicable approach for imaging methylation of individual proteins in human cells, which is based on a combination of chemical reporter strategies, bioorthogonal ligation reactions, and FRET detected by means of fluorescence lifetime imaging microscopy. Through this approach, methylation of histone 4 and the non-histone proteins tumor suppressor p53, kinase Akt1, and transcription factor Foxo1 in two human cell lines has been successfully imaged. To further demonstrate its potential, the localization-dependent methylation state of Foxo1 in the cellular context has been visualized.
Keyphrases
- dna methylation
- genome wide
- transcription factor
- high resolution
- single molecule
- signaling pathway
- endothelial cells
- amino acid
- gene expression
- binding protein
- stem cells
- pi k akt
- crispr cas
- mass spectrometry
- mesenchymal stem cells
- energy transfer
- small molecule
- photodynamic therapy
- living cells
- induced pluripotent stem cells
- quantum dots