LKB1 controls inflammatory potential through CRTC2-dependent histone acetylation
Deregulated inflammation is a critical feature driving progression of tumors harboring mutations in the Liver kinase B1 (LKB1), yet the mechanisms linking LKB1 mutations to deregulated inflammation remain undefined. In this research publication we identify deregulated signaling by CREB regulated transcription coactivator 2 (CRTC2) as an epigenetic driver of inflammatory potential downstream of LKB1 loss. We demonstrate that LKB1 mutations sensitize both transformed and non-transformed cells to diverse inflammatory stimuli, promoting heightened cytokine and chemokine production. LKB1 loss triggers elevated CRTC2-CREB signaling downstream of the salt inducible kinases (SIKs), increasing inflammatory gene expression in LKB1-deficient cells. The immunoblot data contained in this dataset (for Figures 4, S2, and S3) show in murine LKB1-deficient MEFs and Kras/LKB1 (KPL) mutant lung cancer cells that CRTC2 is hypophosphorylated and CREB is hyperphosphorylated.
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MEF cells were cultured in DMEM (Wisent) supplemented with 10% FBS (Wisent), L-glutamine (Gibco), and penicillin–streptomycin (Invitrogen). Kras-mutant (G12D) NSCLC cells deficient for p53 (KP) or both p53 and Stk11/LKB1 (KPL) were cultured in RPMI 1640 with L-glutamine (Gibco) supplemented with 10% FBS (Wisent) and penicillin–streptomycin (Invitrogen) where indicated. Cells were treated with rapamycin (50 nM) (Millipore Sigma) where indicated. For whole cell immunoblotting, cells were lysed in 1X RIPA buffer (Cell Signaling Technology, Danvers, MA) supplemented with protease and phosphatase inhibitors (Roche). Cytoplasmic and nuclear cell fractions were prepared with a nuclear extraction kit according to the manufacturer’s instructions (Abcam). Protein was quantified using a Pierce BCA Protein Assay Kit (Thermo Fisher Scientific, Waltham, MA, USA). Lysates were resolved by SDS-PAGE, transferred to nitrocellulose, and incubated with primary antibodies (see Key Resources Table) and HRP-conjugated secondary antibodies (Cell Signaling Technology).