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. 2022 Nov 14;25(12):105573. doi: 10.1016/j.isci.2022.105573

Figure 5.

Figure 5

sdha-1, sdha-2 and icl-1 mutants have fewer sperm in hermaphrodite spermatheca

(A) Number of sperm in one hermaphrodite spermatheca at day 1 of adulthood. Genotype/+ indicates balanced strains, where sperm were counted in a heterozygote hermaphrodite. Data are mean ± SEM; n = 8-11. Comparisons are between the indicated strain and wild-type and were performed using one-way ANOVA with Dunnett’s post hoc test ∗∗p ≤ 0.01, ∗∗∗p ≤ 0.001, ∗∗∗∗p ≤ 0.0001.

(B) Representative maximum intensity Z-projections of a spermatheca in DAPI-stained day 1 adult hermaphrodites.

(C) Representative images of DAPI-stained day 1 adult hermaphrodites. Sperm can be detected throughout the uterus of sdha-2(tm1420) and sdhc-1(ok909) animals, indicated by arrowheads. (B and C) The spermatheca is outlined in pink. Scale bar = 50 μm.

(D) SDH is comprised of four subunits: SDHA, SDHB, SDHC, and SDHD. In C. elegans, these are encoded by sdha-1 and -2, sdhb-1, sdhc-1, and sdhd-1 respectively. FUM-1 is the C. elegans fumarase, and ICL-1 is the C. elegans isocitrate lysase/malate synthase.

(E–H) Brood size assay to assess icl-1 fertility. (E and F) The average number of (E) live progeny and (F) unfertilized oocytes produced per animal per day. (G and H) The total number of (G) live progeny and (H) unfertilized oocytes per animal. All animals were derived from a single cross between sdha-2(tm1420) and icl-1(ok531) to ensure a consistent genetic background between strains. Three lines were selected from the single cross for each genotype, used in three independent brood size assays (n = 8 worms per assay). Data are mean ± SEM; n = 24. Comparisons were performed using one-way ANOVA with Tukey’s post hoc test ∗∗p ≤ 0.01, ∗∗∗p ≤ 0.001, ∗∗∗∗p ≤ 0.0001.

(I) Representative images of MitoTracker Red sperm in the uterus of wild-type hermaphrodites. The spermatheca is outlined in blue and V = vulva. Scale bar = 20 μm.

(J) The percentage of sperm in the three zones of the hermaphrodite uterus 1 h after mating (zone 1 = vulval region, zone 2 = central zone, zone 3 = spermathecal region). n = 11 per strain.

(K) All visible sperm in zone 2 were tracked and averaged to give the average sperm velocity per gonad arm 30 min-1 hr post-mating. n = 3 (wild-type) and n = 4 (sdha-2(tm1420)) gonad arms.