Male-specific roles of lincRNA in C. elegans fertility

Abstract

The testis is the mammalian tissue with the highest expression levels of long intergenic non-coding RNAs (lincRNAs). However, most in vivo models have not found significant reductions in male fertility when highly expressed lincRNA genes were removed. This suggests that certain lincRNAs may act redundantly or lack functional roles. In the genome of the nematode Caenorhabditis elegans, there is an order of magnitude fewer lincRNA genes than in mammals. This characteristic lowers the potential for redundancy, making it an ideal model to test these possibilities. We identified five highly and dynamically expressed lincRNAs in male C. elegans gonads and quantified the fertility of worm strains in which these genes were removed. In contrast to the hermaphrodites of deletion strains, which exhibited no significant reductions in broods, smaller brood sizes were observed in the progeny of males of three of the lincRNA deleted strains. This demonstrates reduced male fertility in worms with those genes removed. Interestingly, reduced brood size was statistically significant only in the last days of egg laying in two of these strains. This suggests the effect is due to early deterioration and aging of the transferred sperm. We detected a mild increase in embryonic lethality in only one of the strains, supporting the possibility that these lincRNAs do not affect fertility through critical roles in essential meiotic processes. Together our results indicate a sexually dimorphic outcome on fertility when lincRNA are removed and show that, unlike mammals, individual lincRNAs in C. elegans do play significant roles in male fertility.

Keywords: C. elegans; fertility; lincRNA; lncRNA; long intergenic non-coding RNA; spermatogenesis.

FIGURE 1

Expression patterns of lincRNAs in the male gonad. Log₂ of normalized expression values of five lincRNAs with high levels of expression along the male gonad from proliferative to mature sperm stages. X-axis numbers correspond to the segments used for the analysis and refer to the following stages: 1-2 proliferative, 2–4 leptotene/zygotene, 5-6 pachytene, 7-8 condensation, and division, 9-10 spermiogenesis. Adapted from (Tzur et al., 2018).

FIGURE 2

Deletion of three lincRNA genes leads to reduced male fertility. Average progeny brood sizes for females mated with males of the indicated lines. Mann-Whitney p-value: * <0.05, ** <0.01.

FIGURE 3

Progeny of males with deletions in lincRNA genes do not undergo substantial embryonic lethality. Average embryonic lethality of the progeny of males of the indicated genotypes. Mann-Whitney p-value: * <0.05.

FIGURE 4

Dynamics of brood size along the reproductive term of lincRNA deleted males. (A) Average progeny brood size for females mated with males of the indicated genotypes along six 24 h intervals. X-axis timepoints refer to days post fertilization. (B) Percentage of embryos laid on day 2 and day 3 by progeny of WT and linc-168 males.