Figure 5. Dynamic chromatin binding of... | Rockefeller University Press

Figure 5.

$Figure 5. Dynamic chromatin binding of licensing factors. (A) Time-lapse images of embryos progressing through anaphase, expressing GFP–CDC-6, –ORC-1, –ORC-2, or –MCM-3 as indicated. The anterior (left) mass of DNA was photobleached 20–30 s after anaphase onset. The posterior (right) mass of DNA was used as control. Embryos are shown at −1, 1, 4, and 8 s after photobleaching. Bar, 10 µm. (B) Normalized fluorescence intensity of bleached (solid lines) or unbleached control (dashed lines) DNA (intensity prephotobleaching = 1, and cytoplasmic intensity = 0). (C) Turnover rates obtained by subtracting the accumulation of GFP on chromatin (given by the unbleached control DNA) from the normalized fluorescence intensity. (D and E) Mobile fractions of chromatin-bound licensing proteins (left graphs) and half-lives of protein recovery (right graphs) for wild-type (D) and cdt-1 RNAi (E) embryos. The mobile fraction is the fraction of GFP recovered 20 s after photobleaching. The half-life is the time to recover half of the mobile fraction. Error bars represent SD. n = 5.$

Dynamic chromatin binding of licensing factors. (A) Time-lapse images of embryos progressing through anaphase, expressing GFP–CDC-6, –ORC-1, –ORC-2, or –MCM-3 as indicated. The anterior (left) mass of DNA was photobleached 20–30 s after anaphase onset. The posterior (right) mass of DNA was used as control. Embryos are shown at −1, 1, 4, and 8 s after photobleaching. Bar, 10 µm. (B) Normalized fluorescence intensity of bleached (solid lines) or unbleached control (dashed lines) DNA (intensity prephotobleaching = 1, and cytoplasmic intensity = 0). (C) Turnover rates obtained by subtracting the accumulation of GFP on chromatin (given by the unbleached control DNA) from the normalized fluorescence intensity. (D and E) Mobile fractions of chromatin-bound licensing proteins (left graphs) and half-lives of protein recovery (right graphs) for wild-type (D) and cdt-1 RNAi (E) embryos. The mobile fraction is the fraction of GFP recovered 20 s after photobleaching. The half-life is the time to recover half of the mobile fraction. Error bars represent SD. n = 5.