Fig. 4: The intracellular domain (ICD) of TrkB transduces signals to FAK, while a membrane-targeted ICD of TrkB is linked to MAPK phosphorylation.

A Model depicting intracellular kinase-active domain constructs of TrkB. In Myr-ICD, an N-terminal myristoylation motif with a glycine-serine (GGSGG)-linker was used for plasma membrane targeting. The ICD (intracellular domain) construct lacks ligand-binding and transmembrane domains. The model includes the results of the experiment. Constitutively active TrkB-wt signals to MAPK and FAK, whereas Myr-ICD activates MAPK, but not FAK. ICD activates FAK but not MAPK. B TrkB-ICD and TrkB-Myr-ICD undergo self-activation but differ in their cellular localization patterns. Immunofluorescence of pTrk-kin (magenta), DAPI (blue) and Acti-stain-670 phalloidin (phall, green). HEK293 cells were immunostained 30 h after transfection. TrkB-ICD is preferentially observed at intracellular sites. The Myr-ICD construct showed typical plasma membrane targeting as expected. Note morphological changes in DAPI staining of ICD-expressing cells, indicating changes in chromatin compaction (cyan arrows) and pTrk-kin clusters (magenta arrows). Scale bar: 20 µm. C TrkB-ICD, but not TrkB-Myr-ICD, induced FAK phosphorylation. Western blotting of whole-cell lysates generated from HEK293 cells expressing TrkB-ICD, TrkB-Myr-ICD, TrkB-wt, or TrkB-YFF. K252a was used to inhibit the Trk kinase activity. TrkB-wt expression increased MAPK and FAK phosphorylation. ICD signaled to FAK, but not to MAPK. Myr-ICD induced MAPK phosphorylation but failed to activate FAK. D–F Quantification of Western blots for pTrkB-kin normalized to total TrkB levels (in D), pFAK to total FAK (in E), and pMAPK to total MAPK (in F). The relative integrated densities are also presented. Bar graph: Mean ± SEM, overlaid with single data points; n = 4. UT untransfected control.