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. Author manuscript; available in PMC: 2020 Mar 2.
Published in final edited form as: Cell Rep. 2020 Feb 11;30(6):1848–1861.e5. doi: 10.1016/j.celrep.2020.01.050

Figure 1. Mutating endu-2 in C. elegans Suppresses NT Deficiency-Induced Germline Proliferation Arrest and Renders Worms Hypersensitive to NT Supplementation and Genotoxic Drugs.

Figure 1.

(A) Results of fertility test of worms fed low-pyrimidine food (E. coli OP50). Loss of endu-2 gene function suppressed sterility caused by cdd-1(−);cdd-2(−) double knockout (cddko). A transgene of wild-type (WT) endu-2, but not the endu-2(H470A) mutant gene, fully reversed the suppression. Expression of endu-2 specifically in the intestine using the Pges-1 promoter also fully reversed the suppression. The endu-2(−) allele is ku544. “Ex” indicates extrachromosomal transgenes. Error bars, standard deviation. Asterisks indicate significant differences. Tukey’s range test, family-wise error rate (FWER) = 0.01; see Method Details for details. The suppression of the germline morphology defect is described in the text and Figure S1A.

(B) Bright-field and fluorescence images showing the expression of a single-copy insertion of the Pendu-2::gfp reporter in the intestine. Dotted lines outline the distal region of the germline.

(C and D) Lifespan assays of worms with indicated RNAi treatment from L1 (C) or adult (D). *p < 0.01; N.S., not significant (p > 0.05), log-rank test. See text for details.

(E and F) High-level single-NT supplementation or genotoxic drugs caused severe developmental delay in endu-2(−) animals. WT or endu-2(−) worms (150–250 per replicate) were supplemented with 100 mM NTs uridine (U), thymidine (T), or cytidine (C) in (E), or hydroxyurea (HU) (10 mM), 5-FU (50 μM) or FUDR (50 μM) in (F).

(G) Fluorescence image showing the DNA bridge phenotype in a DAPI-stained worm intestine. The arrow points to the bridge connecting 2 daughter nuclei. Scale bar represents 10 μm.

(H) Percentage of adult worms having DNA bridges in the intestine.

(E, F, and H) Error bars, standard deviation. Asterisks indicate significant differences. Student’s t test between WT and endu-2(−) with the same treatment, unpaired, two-tailed, with Bonferroni correction, α = 0.0018 (E), 0.0033 (F and H); see Method Details for details.

Unless noted specifically, 50–100 worms were examined in each replicate, with at least 2 replicates for each experiment.