Journal: Communications Biology

Article Title: Photosensitive tyrosine analogues unravel site-dependent phosphorylation in TrkA initiated MAPK/ERK signaling

doi: 10.1038/s42003-020-01396-0

Figure Lengend Snippet: a Western blot analysis of HEK293T cells expressing ONB mutants for phosphorylated ERK (Thr202 and Tyr204) in the presence of ligand NGF without (−) or with (+) UV light. b Binding of the NGF drives TrkA dimerization, autophosphorylation and initiates intracellular MAPK/ERK. In the presence of ligand NGF, ONB incorporated at the Y490, Y674, and Y675 (solid circles) sites in TrkA, respectively, after UV treatment initiate intracellular MAPK/ERK signaling (arrows). ONB incorporated in Y670 (empty circle) site does not initiate intracellular MAPK/ERK signaling (inhibited arrow) c Western blot analysis of HEK293T cells expressing AzF mutants for phosphorylated ERK (Thr202 and Tyr204) without (−) or with (+) UV light. UV photolysis was done with 365 nm UV cross-linker CL-1000L (Analytik Jena) at 40w for 5 min at 4 °C. TrkA and β-actin were used as loading control. d Binding of the NGF drives TrkA dimerization, autophosphorylation and initiates intracellular MAPK/ERK. In the presence of ligand NGF, AzF incorporated at the Y670, Y674, and Y675 (solid circles) sites in TrkA, respectively, after UV treatment initiate intracellular MAPK/ERK signaling (arrows) presumably due to the crosslinking with a putative interactive protein. AzF incorporated in Y490 (empty circle) site does not initiate intracellular MAPK/ERK signaling (inhibited arrow).

Article Snippet: UV photolysis was done with 365 nm UV cross-linker CL-1000L (Analytik Jena) at 40w for 5 min.

Techniques: Western Blot, Expressing, Binding Assay