Figure 6.

PP-1 isoforms distribute to different compartments in mitotic HeLa cells as assayed in immunoblots. PP-1 γ1, but not PP-1 α or δ, is enriched in microtubule fractions after cell lysis (A). Residual pellets from cells treated with taxol contain assembled microtubules, but pellets from cells treated with 1.0 μg/ml nocodazole (Noc.) are devoid of microtubules. PP-1 γ1 is preferentially associated with the residual cell pellet that contains microtubules after treatment with taxol. Whole cell lysates and residual cell pellets from cells treated with either taxol or nocodazole and loaded on the basis of equal cell number are shown. PP-1 δ is associated with chromosomes, as determined by release from the residual cell pellet after digestion with DNase I (B). Mitotic cells were collected by selective detachment, permeabilized, and incubated in digestion buffer either with (+) or without (−) DNase I, and supernatants (Sup.) were loaded onto gels. The residual cell pellet after lysis, loaded on the basis of equal cell number, is shown for reference. The majority of PP-1 δ is present in the cell supernatant from nocodazole-treated cells (see Fig. 6 A). Therefore, the blot of PP-1 δ shown here was exposed extensively to bring out the pellet fraction to examine its extractability by digestion with DNase I.