Germline <i>uev-2</i> overexpression differentially impacts DSB levels in young and aged germlines.

A) Schemes used to isolate young (1 day post-L4) and aged (4 days post L4) worms for experiments. Days count ~18–24 hour periods after hermaphrodites were isolated as L4 larvae and are separated by alternating grey shaded boxes. B) Representative images of RAD-51 foci in meiotic nuclei of young pie-1p::uev-2, aged pie-1::uev-2, and young wildtype germlines. Scale bars represent 5μm. C) RAD-51 foci per nucleus in oocytes. Line plots represent the mean RAD-51 foci per nucleus along the length of the germline in a sliding window encompassing 0.1 units of normalized germline distance with a step size of 0.01 germline distance units. Mean RAD-51 foci were calculated from nuclei analyzed in n = 9 total germlines derived from ≥3 experimental replicates within each age group. Shaded areas around each line represent ± SEM. Total nuclei analyzed (Bins 1/2/3/4/5/6/7) Young Wildtype: 185/117/146/107/117/97/83; Aged Wildtype Mated: 234/205/192/173/154/177/96; Young pie-1p::uev-2: 182/192/162/125/116/97; Aged pie-1p::uev-2 Mated: 140/149/136/127/126/95/102. Germlines distances were normalized to the start (0) and end (1) of pachytene based on DAPI morphology (see Methods). For analysis, the germline was divided into 7 bins encompassing the transition zone (Bin 1), early pachytene (Bins 2–3), mid pachytene (Bins 4–5), and late pachytene (Bins 6–7). The germline positions at which each bin start and end are marked on the X axis as vertical grey lines. Heat maps below each bin display the p values of pairwise comparisons of RAD-51 foci per nucleus counts within that bin. P values were calculated using Mann-Whitney U tests with Holm-Bonferroni correction for multiple comparisons. Blue tiles indicate significant differences (adjusted p value 0.05). Young and aged mated wild type data is shared with Figs 1 and 6. Numerical data associated with this figure is presented in S1 Data.