Nuclear and cytoplasmic extracts were made according to previously published protocols (Smith et al., 2004 (link); Rozenfeld and Devi, 2007 (link)). Briefly, PANC-1 cells were treated with 10 nM dermorphin or 10 nM (1R,1′S,3′R/1R,1′R,3′S)-L-054,264 (Tocris, Bristol, UK) individually and in combination for the indicated times. After the treatment, cells were washed in ice-cold PBS three times and scraped into lysis buffer (10 mM Tris, pH 7.4, 10 mM NaCl, 3 mM MgCl2, 1 mM ethylene glycol tetraacetic acid, 1 mM sodium orthovanadate, and a protease phosphatase inhibitor tablet [ThermoFisher Pierce]) and incubated on ice for 10 min. The lysate was then homogenized and centrifuged at 375 × g for 5 min. The pellet consisting of the nuclear fraction was washed five times with lysis buffer containing 0.1% NP-40 to remove any nonnuclear contamination and resuspended in lysis buffer containing NP-40. The soluble fraction was centrifuged twice at 375 × g to remove nuclear contamination and used as the cytosolic fraction. Fractions were then used for immunoblotting for anti–phospho-ERK1/2, anti–phospho-p90RSK (Thr-573; rabbit polyclonal; Cell Signaling, Danvers, MA), and anti–β-actin as a loading control for the cytoplasmic fraction, and Histone H3 antibody (rabbit monoclonal; Cell Signaling) was used as a loading control for the nuclear fraction.