Large neurological component to genetic differences underlying biased sperm use in Drosophila

Abstract

Sperm competition arises as a result of complex interactions among male and female factors. While the roles of some male factors are known, little is known of the molecules or mechanisms that underlie the female contribution to sperm competition. The genetic tools available for Drosophila allow us to identify, in an unbiased manner, candidate female genes that are critical for mediating sperm competition outcomes. We first screened for differences in female sperm storage and use patterns by characterizing the natural variation in sperm competition in a set of 39 lines from the sequenced Drosophila Genetic Reference Panel (DGRP) of wild-derived inbred lines. We found extensive female variation in sperm competition outcomes. To generate a list of candidate female genes for functional studies, we performed a genome-wide association mapping, utilizing the common single-nucleotide polymorphisms (SNPs) segregating in the DGRP lines. Surprisingly, SNPs within ion channel genes and other genes with roles in the nervous system were among the top associated SNPs. Knockdown studies of three candidate genes (para, Rab2, and Rim) in sensory neurons innervating the female reproductive tract indicate that some of these candidate female genes may affect sperm competition by modulating the neural input of these sensory neurons to the female reproductive tract. More extensive functional studies are needed to elucidate the exact role of all these candidate female genes in sperm competition. Nevertheless, the female nervous system appears to have a previously unappreciated role in sperm competition. Our results indicate that the study of female control of sperm competition should not be limited to female reproductive tract-specific genes, but should focus also on diverse biological pathways.

Figure 1

DGRP lines vary widely in female effect on P1 scores. Each box plot represents P1 scores from 30 females from a single DGRP line. P1 score is the proportion of first-male progeny after the second mating. Mean P1 scores vary up to sixfold among genotypes. Female DGRP genotype has a significant effect on P1 score (P = 0.0003). In the box plots, the boxes represent the interquartile range, the whiskers represent 1.5 × interquartile range, and open circles are outliers. See Table S1 for line identity.

Figure 2

Average expression of candidate genes associated with sperm competition in different tissues. The mean expression levels of the 33 candidate genes are higher in neuronal tissues (dark shading) than in nonneuronal tissues (light shading) (P = 0.0019). Tissue-specific expression data were taken from FlyAtlas. TaG, thoracicoabdominal ganglion; SG, salivary gland; Spt, spermatheca; L_CNS, larval central nervous system. Mean ± SD is shown.

Figure 3

Level of first-male sperm precedence (P1 score) in RNAi knockdown of select neurological candidates. Three of the four candidates chosen for functional analysis affect P1 score when knocked down in sensory neurons innervating the female reproductive tract. KD, knockdown; *P < 0.05.