Figure 3.

Cell-cycle-regulated chromatin binding of Cdc46p and Cdc47p. (a) Whole-cell extracts (WCE), supernatants (Su), and chromatin fractions (Ch) were prepared as described in Materials and Methods. SDS/PAGE was performed as described in Fig. 1, and immunoblots were performed as described in Materials and Methods. (a) Cdc46p is chromatin-bound in G₁ but not G₂/M. PY26 containing tagged Cdc46p was blocked with either nocodazole (NOC) or alpha factor (α) before chromatin isolation. (b) Cdc47p is chromatin-bound throughout G₁ but not in the S, G₂/M, or late M phases. W303–1a containing tagged Cdc47p and no additional mutation (lanes 1–3, 7–9, and 13–15), the cdc15–2 mutation (lanes 4–6), or the cdc7–1 mutation (lanes 10–12) were blocked either with the indicated inhibitors or by raising cultures of mutant strains to the nonpermissive temperature until a uniform arrest was achieved before chromatin isolation.