Figure 5.
Figure 5. Matched doses of nocodazole reveal differences in APC activity with reduced oocyte volume. (A) B0 oocytes were matured in the indicated concentrations of nocodazole and assessed for bivalent biorientation 6 h after NEB. Colors indicate the number of SDs a bivalent is, according to its worst-performing measure, from mean values defined by metaphase oocytes treated without nocodazole (green <1, yellow ≤2, orange ≤3, and red >3). (B and C) 3D scatterplots showing the extent of chromosome biorientation in B3 oocytes (25 nM nocodazole; B) and B0 oocytes (35 nM nocodazole; C) as in A. (D) Percentage of oocytes of the indicated size extruding polar bodies after 15-h culture in the indicated concentration of nocodazole. (E) Duration of MI, defined as the time from NEB to anaphase. (F) Individual (pale) and mean (bold) securin traces from B0 and B3 oocytes, arranged relative to the time of APC activation. Horizontal bars show the corresponding timing of anaphase. (G) Representative traces of two modes of securin degradation found in B3 oocytes matured in 25 nM nocodazole. Arrowheads indicate the timing of anaphase. Red color indicates periods of stalling. (H) Proportion of MI-arrested oocytes that exhibited stalling. (I) Proportion of oocytes that completed MI that exhibited stalling. (J) Results from experiments with and without nocodazole addition are presented together for comparison of the effects of oocyte volume reduction and addition of matched doses of nocodazole. Blue points, B3 oocytes; red, B0; round points, no stalling event was detected; square points, stalling. Back border indicates the presence of nocodazole (25 nM for B3 and 35 nM for B0). Parentheses indicate number of oocytes. Error bars are SDs. Gray dashed lines indicate points where the duration of MI (prometaphase + metaphase) is the same. Statistical comparisons between all points are indicated in Fig. S3. In A, D, E, and G–I, numbers of oocytes are indicated in parenthesis. Statistical tests used were Fisher’s exact test (D, H, and I) and unpaired t test (E).

Matched doses of nocodazole reveal differences in APC activity with reduced oocyte volume. (A) B0 oocytes were matured in the indicated concentrations of nocodazole and assessed for bivalent biorientation 6 h after NEB. Colors indicate the number of SDs a bivalent is, according to its worst-performing measure, from mean values defined by metaphase oocytes treated without nocodazole (green <1, yellow ≤2, orange ≤3, and red >3). (B and C) 3D scatterplots showing the extent of chromosome biorientation in B3 oocytes (25 nM nocodazole; B) and B0 oocytes (35 nM nocodazole; C) as in A. (D) Percentage of oocytes of the indicated size extruding polar bodies after 15-h culture in the indicated concentration of nocodazole. (E) Duration of MI, defined as the time from NEB to anaphase. (F) Individual (pale) and mean (bold) securin traces from B0 and B3 oocytes, arranged relative to the time of APC activation. Horizontal bars show the corresponding timing of anaphase. (G) Representative traces of two modes of securin degradation found in B3 oocytes matured in 25 nM nocodazole. Arrowheads indicate the timing of anaphase. Red color indicates periods of stalling. (H) Proportion of MI-arrested oocytes that exhibited stalling. (I) Proportion of oocytes that completed MI that exhibited stalling. (J) Results from experiments with and without nocodazole addition are presented together for comparison of the effects of oocyte volume reduction and addition of matched doses of nocodazole. Blue points, B3 oocytes; red, B0; round points, no stalling event was detected; square points, stalling. Back border indicates the presence of nocodazole (25 nM for B3 and 35 nM for B0). Parentheses indicate number of oocytes. Error bars are SDs. Gray dashed lines indicate points where the duration of MI (prometaphase + metaphase) is the same. Statistical comparisons between all points are indicated in Fig. S3. In A, D, E, and G–I, numbers of oocytes are indicated in parenthesis. Statistical tests used were Fisher’s exact test (D, H, and I) and unpaired t test (E).

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