Extended Data Fig. 3: Overexpression of mutant KRAS increases nuclear export of EZH2 and Survivin, which is independent of PI3K and MEK signaling; KRAS inhibition does not prevent XPO1:Survivin complex formation.
a-d Overexpression of KRAS-G12C in H1703 (a,b) or KRAS-G12D in HBEC (c,d) lines increased the level of cytoplasmic EZH2 and cytoplasmic Survivin and decreased the cytoplasmic DLC1 protein. α-tubulin and lamin B1 were used, respectively, as cytoplasmic and nuclear marker proteins. C, Cytoplasmic; N, Nuclear. e In A549 cells, MEK inhibitor U0126-ethanol and PI3K inhibitor wortmannin did not affect the level of DLC1 protein, while the XPO1 inhibitor selinexor did increase DLC1 protein. U0126-ethanol inhibited MEK activity (measured by pERK-T202/Y204) and wortmannin inhibited PI3K activity (measured by pAKT-S473) in all treated samples. f,g In the KRAS-G12C NCI-H23 line, selinexor prevented the complex formation between XPO1 and EZH2 in the nucleus, as it is confirmed from the purified nuclear extract. h,i KRAS inhibition by sotorasib (h) or siRNA knockdown of KRAS (i) did not prevent XPO1:Survivin complex formation. Lysates from NCI-H23 cells treated without or with sotorasib or KRAS siRNA were IP with antibodies to XPO1, Survivin, or mock IgG, followed by IB with antibodies to Survivin or XPO1. WCE, whole cell extract. j Complex formation between KRAS and NTF2 in A549 cells. Lysates from A549 cells were IP with antibodies to KRAS or mock IgG, followed by IB with antibodies to NTF2 or KRAS. k-n Overexpressed KRAS-G12C (k,l) or KRAS-G12D (m,n) formed a complex with full-length RanGAP1 (k,m) and the RanGAP1 catalytic domain (l,n). Two independent experiments were performed for each image, with similar results.
