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Figure 5


Fig. 5. The region between L545 and E587 in Gab1 blocks membrane recruitment of full-length Gab1. (A) Schematic representation of the analysed Gab1-GFP fusion proteins used in this study. Tyrosine motifs involved in protein binding are indicated by Y. Proline-rich regions are highlighted in red. The Met-binding domain is underlined. The numbers represent the first and last amino acids in the mutant proteins corresponding to the numbering in Gab1 wild-type protein. GFP is fused to the C-terminus of Gab1. (B) HEK293T cells were transfected with expression vectors for Gab1-GFP fusion proteins depicted in A, together with vectors encoding EpoR-gp130 chimeric receptors. 24 hours after transfection, cells were transferred to poly-L-lysine-coated coverslips. The next day cells were starved for 3 hours in KRH and transferred into the perfusion chamber of a laser-scanning microscope. After 20 minutes, the cells were stimulated with Epo (7 U/ml). Gab1 localisation is shown before stimulation and after 20 minutes of Epo stimulation.





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