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Fig. 8. MyrFAK signals through multiple, independent complexes to suppress apoptosis. (a) Mock-transfected HEK293T or HEK293T cells transiently expressing myrFAK constructs were detached for 15 minutes and treated with and without the membrane permeable crosslinker DSS. Cell lysates were immunoprecipitated with anti-V5. Immunoprecipitates were immunoblotted with anti-FAK Ab and the same immunoblot re-probed with anti-paxillin Ab. ^*, Position of the 200 and 300 kDa complexes seen following crosslinking. (b) Suppression of anoikis in MECs expressing myrFAK, myrFAKY925F or myrFAKI936/998E alone, or both myrFAKY925F and myrFAKI936/998E. Results are the mean of three independent experiments. Error bars indicate standard error. ^*, Significant suppression of apoptosis compared with either myrFAKY925F or myrFAKI936/998E alone. (c) HEK293T cells transiently expressing myrFAK constructs were detached for 15 minutes as indicated. Whole-cell lysates were immunoblotted as indicated. (d) MECs and MEFs were left adherent or detached for the indicated times. MECs were left in complete growth medium. MEFs were deprived of serum growth factors, except for the indicated adherent cells. Whole-cell lysates (WCL) were prepared and immunoblotted for total Akt or phospho-Akt serine 473. (e) MECs or MEFs expressing myrFAK (+), or non-expressing controls (–), were left adherent or detached as indicated. WCL were prepared and immunoblotted for phospho-Akt serine 473, total Akt, or myrFAK (anti-V5).

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