L* inhibits mouse RNase L dimerization and oligomerization.

<p>C-terminal Flag-tagged RNase L (RNase L-Flag) and L* were overexpressed in 293T cells by transfection. Cells lysates were divided into samples where RNase L dimerization was induced by adding increasing concentrations of 2-5A. Dimers/oligomers were crosslinked using DSS. A. RNA was extracted from lysates of (B) and analyzed on agarose gel electrophoresis as a control for RNase L activation. 2-5A concentrations: 0.25, 0.5, 1 and 2μM ATP equivalent. B. The upper panel shows the detection by immunoblot of WT mouse RNase L-Flag monomers, dimers and oligomers using an anti-Flag antibody. Lower panels show detection of L* and GAPDH used as a loading control. 2-5A concentrations: 0.25, 0.5, 1 and 2μM ATP equivalent. C. The upper panel shows the detection of WT human RNase L-Flag monomers, dimers and oligomers using an anti-Flag antibody. Lower panels show detection of L* and GAPDH. 2-5A concentrations: 0, 2, 4 and 8μM ATP equivalent. D. Dimerization-defective (K391R) mouse RNase L was used as a dimerization control. The upper panel shows the detection of RNase L-Flag monomers, dimers and oligomers using an anti-Flag antibody. Lower panels show detection of L* and GAPDH. 2-5A concentrations: 0.5 and 2μM ATP equivalent. Reproducible results were obtained in 2 independent experiments. E. Mass spectrometry analysis of high molecular weight RNase L complexes. Proteins for which 2 or more unique peptides were detected, with a false discovery rate below 5%, are listed in the table. # PSMs: number of peptide spectrum matches. /: no peptide detected above threshold.</p>