Gene-Level, but Not Chromosome-Wide, Divergence between a Very Young House Fly Proto-Y Chromosome and Its Homologous Proto-X Chromosome

Abstract

X and Y chromosomes are usually derived from a pair of homologous autosomes, which then diverge from each other over time. Although Y-specific features have been characterized in sex chromosomes of various ages, the earliest stages of Y chromosome evolution remain elusive. In particular, we do not know whether early stages of Y chromosome evolution consist of changes to individual genes or happen via chromosome-scale divergence from the X. To address this question, we quantified divergence between young proto-X and proto-Y chromosomes in the house fly, Musca domestica. We compared proto-sex chromosome sequence and gene expression between genotypic (XY) and sex-reversed (XX) males. We find evidence for sequence divergence between genes on the proto-X and proto-Y, including five genes with mitochondrial functions. There is also an excess of genes with divergent expression between the proto-X and proto-Y, but the number of genes is small. This suggests that individual proto-Y genes, but not the entire proto-Y chromosome, have diverged from the proto-X. We identified one gene, encoding an axonemal dynein assembly factor (which functions in sperm motility), that has higher expression in XY males than XX males because of a disproportionate contribution of the proto-Y allele to gene expression. The upregulation of the proto-Y allele may be favored in males because of this gene's function in spermatogenesis. The evolutionary divergence between proto-X and proto-Y copies of this gene, as well as the mitochondrial genes, is consistent with selection in males affecting the evolution of individual genes during early Y chromosome evolution.

Keywords: Diptera; allele-specific expression; insect; sex chromosome evolution; sexual conflict.

Fig. 1.

Elevated heterozygosity on the third and X chromosomes in genotypic (III^M/III) males relative to sex-reversed (III/III) males. The boxplots show the distributions of the percent of heterozygous variants per gene in the genotypic males relative to the sex-reversed males (% III^M heterozygous variants) on each chromosome. Muller element nomenclature for each chromosome is shown in parentheses (Meisel and Scott 2018). See Materials and Methods for the calculation of % III^M heterozygous variants. Values >50% indicate more heterozygous variants in genotypic (III^M/III) males, and <50% indicates more heterozygous variants in sex-reversed males. The median across autosomes I, II, IV, and V is represented by a dashed line.

Fig. 2.

Two proto-X and proto-Y loci identified in the III^M genome assembly. (A) One locus was identified with a truncated Mdmd on the proto-Y (ctg2382) and the same three genes on both the proto-X (ctg1607) and proto-Y. (B) One locus was identified with four genes on both the proto-X (ctg1519) and proto-Y (ctg2522). The two contigs (ctg2522 and ctg1519) were assigned to the proto-Y and proto-X based on sequences that are enriched in the male relative to female reads (see supplementary fig. 1, Supplementary Material online).

Fig. 3.

Evidence for moderately elevated ASE on the third (proto-Y) chromosome in III^M males. (A) Proportions of genes with ASE in genotypic (III^M) or sex-reversed (III) males on each chromosome. There is not a significant difference on any chromosome between the two genotypes. (B) Proportions of genes with ASE in genotypic males and non-ASE in sex-reversed males on the third chromosome and all other chromosomes (left two bars). Proportions of genes with non-ASE in genotypic males and ASE in sex-reversed males on the third chromosome and all other chromosomes (right two bars). The asterisk indicates a significant difference (P < 0.05) in the number of genes in these categories as determined by Fisher’s exact test.

Fig. 4.

Allele-specific expression (ASE) of Md-HEATR2. (A) Diagnostic variable sites for ASE in the Md-HEATR2 gene are based on haplotypes estimated in IDP-ASE. Read depth is measured as fragments per million mapped reads (FPM) in III^M males, sex-reversed (SR) males, and the III^M or III allele in III^M males. (B) Variable sites that differ between genotypic (III^M) males and sex-reversed males (triangles) across 1,273 bp upstream of Md-HEATR2 were identified using Oxford Nanopore long reads only. (C) Transcription factor (TF)-binding motifs predicted within 1,273 bp upstream of Md-HEATR2. The starting position of each motif is given, along with its length (in parentheses). None of the variable sites overlaps with predicted TF motifs. All positions are coordinates in scaffold NW_004764965.1 of the reference genome assembly (Scott et al. 2014).