Biochemically distinct complexes composed by Baf200 and Baf180 coexist in U2OS cells.
A, Western blot of chromatin fractions showing Baf200, Baf180, Brg1, and Lamin B levels after treatment of U2OS cells with the indicated siRNA and etoposide (10 μm for 20 min). B, Western blot showing Baf200 and Baf180 levels in etoposide-treated Brg1 knock-out U2OS cells generated using CRISPR/Cas. Total (T), soluble (S), and chromatin-bound (C) cell lysate fractions were analyzed with the indicated antibodies. C, co-IP was performed using cell total extract from U2OS cells transfected with siRNA control, siRNA Brg1, and a Brg1−/− U2OS cell line. Molecular weights for each blot are indicated. D, U2OS nuclear cell lysates were subjected to sucrose gradient fractionation (5–20%) and analyzed by Western blotting with the indicated antibodies. Only fractions 14–25 of a total of 30 fractions are shown. Stars mark the peaks of Baf200 and Brg1. One representative experiment of three independent replicates (utilizing independently cultured cells, processed and resolved in sucrose gradients) is shown. E, quantitative analysis of results in D. F, 293T nuclear cells lysates were subjected to sucrose gradient fractionation (5–20%) and analyzed by Western blotting with the indicated antibodies. Only fractions 14–31 of a total of 38 fractions are shown.