Skip to main content
. Author manuscript; available in PMC: 2024 Jul 24.
Published in final edited form as: Curr Biol. 2023 Jul 14;33(14):3056–3064.e5. doi: 10.1016/j.cub.2023.06.051

Figure 1. spe-36 mutants are sterile due to a sperm defect where sperm are present but do not fertilize the eggs.

Figure 1.

(A) Hermaphrodite and male fertility was determined for spe-36(as6) mutants. Majority of unmated spe-36(as6) hermaphrodites produce no self-progeny (avg = 0.6 progeny, n = 19) and an extrachromosomal array containing a PCR product of F40F11.4 (asEx96) restores fertility to spe-36(as6) hermaphrodites. spe-36(as6) hermaphrodites also produce progeny when mated to wild-type males, showing that the sterility is caused by a defect in spe-36(as6) sperm. When spe-36(as6) males are mated to feminized hermaphrodites they fail to produce progeny, indicating that spe-36 is also necessary for male fertility. Error bars indicate standard error. (B) Schematic of the spe-36 gene structure including the locations of the it 114, as1, and as6 mutations and the region that encodes the EGF motif (amino acids 282 – 329). The it114 and as1 alleles have the same nucleotide change at position chrIV: 11,591,836 resulting in a C -> Y change at amino acid 104. as6 has a nucleotide change at position chrIV: 11,592,109 that alters the splice donor site of the first intron (C. elegans Feb. 2013 (WBcel235/ce11) Assembly). Scale bar represents 100 bp. (C) DIC imaging of spe-36(as6) hermaphrodites shows that oocytes are present in the oviduct and sperm are present in the spermatheca. However unfertilized eggs (arrows) are present in the uterus instead of developing embryos, despite the fact that the sperm must have contacted the eggs when the eggs moved through the spermatheca (brackets) to enter the uterus. (D) DAPI staining confirms the presence of sperm in the spermatheca (small bright spots within brackets) of 1 day adult hermaphrodites. Single DNA masses in the eggs in the uterus (arrowheads) are consistent with a lack of sperm entry into the egg. (E-F) DAPI staining of 4 day adult hermaphrodites. By day 4 of adulthood wild-type hermaphrodites (E) have depleted their self-sperm through fertilization of their eggs. In contrast, spe-36 hermaphrodites of the same age (F) still have abundant sperm in their spermatheca. This indicates that the mutant sperm do not fertilize and are capable of maintaining their position in the spermatheca. (G-J) To confirm that spe-36 males exhibit normal mating, sperm transfer and sperm migratory behavior cylc-2 (mon-2[c41g7.6::mNĜ3xFlag]) was crossed into him-5 and spe-36(as6); him-5. The him-5 mutation was used to generate males within the strain. Males with mNeonGreen (mNG) marked sperm were crossed with N2 L4 hermaphrodites and hermaphrodites were imaged ~24 hours later. Both wild-type and mutant sperm were transferred to hermaphrodites and successfully migrated to the region of the spermatheca. (G-H) N2 hermaphrodite mated to him-5; cylc-2::mNG males (I-J) N2 hermaphrodite mated to spe-36(as6); him-5; cylc-2::mNG males. Scale bars = 50 μm, See also Figure S1 and S4.