FIGURE 3.
In the absence of LAMTOR2, LAMTOR3 is predominantly found as a monomeric protein. A, size exclusion chromatography from LAMTOR2−/−MEF cytosol. Cytosolic fraction was prepared as described under “Experimental Procedures” and run on an Äkta HIPrep1660 Sephacryl S200HR column. The obtained 5-ml fractions were precipitated using trichloroacetic acid and deoxycholate and resuspended in Laemmli sample buffer. The samples were separated by SDS-PAGE, analyzed by Western blotting, and probed with the indicated antibodies. B, size exclusion chromatography from LAMTOR2−/−, LAMTOR2f/−, LAMTOR2−/−GFPLAMTOR2, and LAMTOR2−/−LAMTOR2Δb3.GFP MEF lysates. The same experimental procedure as described in A was performed with total cell lysates from the mentioned MEF cell lines. Images are representative examples from at least two independent experiments.