Quantification of Poly(GA) and Poly(GP) Expression

Cells were lysed in RIPA buffer (Sigma-Aldrich) supplemented with 2× complete Mini, EDTA free protease inhibitor cocktail (Merck), and 2% SDS (ThermoFisher) and homogenized by sonication. After centrifugation at 17,000 × g at 16 °C for 20 min, the protein content of the supernatants was determined as described in the Supplementary Methods. The samples were adjusted to the same concentration (0.6–1.0 mg/mL) and MSD immunoassay was performed in singleplex using 96-well SECTOR plates (MSD) to quantify Poly(GA)/(GP) expression, as described previously^{20 (link)}. In brief, plates were coated with unlabeled anti-poly(GP) or anti-poly(GA) antibodies. After blocking, samples were loaded at 45 µg of protein per well for the poly(GP) immunoassay and at 27 µg of protein for poly(GA). Biotinylated anti-poly(GP) and anti-poly(GA) antibodies were used as detectors, followed by sulfo-tagged streptavidin (Meso Scale Discovery, R32AD). Plates were read with the MSD reading buffer (Meso Scale Discovery, R92TC) using the MSD Sector Imager 2400. Signals correspond to intensity of emitted light upon electrochemical stimulation of the assay plate. Prior to analysis, the average reading from a calibrator containing no peptide was subtracted from each reading. Negative signals were set to ‘0’.