Multiple independent origins of the female W chromosome in moths and butterflies

Abstract

Lepidoptera, the most diverse group of insects, exhibit female heterogamy (Z0 or ZW), which is different from most other insects (male heterogamy, XY). Previous studies suggest a single origin of the Z chromosome. However, the origin of the lepidopteran W chromosome remains poorly understood. Here, we assemble the genome from females down to the chromosome level of a model insect (Bombyx mori) and identify a W chromosome of approximately 10.1 megabase using a newly developed tool. In addition, we identify 3593 genes that were not previously annotated in the genomes of B. mori. Comparisons of 21 lepidopteran species (including 17 ZW and four Z0 systems) and three trichopteran species (Z0 system) reveal that the formation of Ditrysia W involves multiple mechanisms, including previously proposed canonical and noncanonical models, as well as a newly proposed mechanism called single-Z turnover. We conclude that there are multiple independent origins of the W chromosome in the Ditrysia (most moths and all butterflies) of Lepidoptera.

Fig. 1.. The assembled chromosomes and identification of the W chromosome in B. mori.

(A) The syntenic relationships between chromosomes of three genomes (light green), including female P50 genome assembled in this study (dilute purple) and two male P50 genomes released in the database of SilkDB (https://silkdb.bioinfotoolkits.net/) (green) and Silkbase (https://silkbase.ab.a.u-tokyo.ac.jp/) (purple). Chromosome 29 has no homolog chromosome in the male genomes. (B) The identification of B. mori sex chromosomes using the CQ method. The CQ value represents the ratio of male (ZZ) to female (ZW) coverage reads on a chromosome. Theoretically, the CQ values of Z and W should be equal to two and zero, respectively. (C) The distribution of Fem (a Piwi-interacting RNA) precursor on the W chromosome. Each black vertical line indicates a Fem precursor, and the numbers show the copy number of Fem.

Fig. 2.. TEs and protein-coding genes in the silkworm.

(A) The distribution of protein-coding genes and the different types of TEs on different chromosomes of female B. mori (in 1-Mb windows). The vertical coordinate of the gene distribution is the number of genes. The vertical coordinate of the TEs distribution is the number of bases in kilobase (kb). (B) Stacked bar charts with the proportion of different TEs on the autosomal (A), Z, and W chromosomes. LINEs and LTRs content on the W chromosome are much higher than that on autosomal and Z chromosomes. (C) The Venn diagram at the right represents the comparison results of three different predicted gene sets from the genomes of the same strain. The red circle represents the female silkworm gene set identified in this study, while the green and blue circles represent the previously predicted male silkworm gene sets in SilkDB and Silkbase. The top and bottom pie charts in the right panel represent the proportions of transposase-encoding genes and genes without evidence of full-length transcripts in specific genes from Silkbase and SilkDB, respectively. The middle pie chart represents the proportions of transposase-encoding genes and genes without evidence of full-length transcripts in genes that are simultaneously present in both Silkbase and SilkDB but are missing in the predicted gene set of the female silkworm genome.

Fig. 3.. The deduced origins of the W chromosomes are based both on the gene synteny within the Z chromosomes and on the sequence homology between Z and W chromosomes in the same species.

(A) The species distribution of sex chromosome systems, containing female Z0 and ZW systems in Lepidoptera and Trichoptera based on prior studies. The W chromosome is mainly found in Ditrysia (Lepidoptera). (B) The three possible mechanisms of W chromosome formation. The ancestral female Z0 system has a Z chromosome (in red) and pairs of autosomes (A_i, in blue). For the canonical models, the W and Z chromosomes are descended from a single autosome pair and should be more similar than the W and autosomes in a ZW system. In the noncanonical model, the W is derived from a B chromosome that arose by rearrangements or duplications from one or more autosomes. The white regions with blue dotted lines represent the further degradation of the neo-W chromosome. (C) The tree on the left represents the phylogenetic relationships of the species. The topological structure of the tree and the divergence times between the species were estimated by maximum likelihood (ML) based on single-copy genes of lepidopteran BUSCO set. On the right, the female karyotype of each female is presented after the corresponding species name. In the middle (in green) are shown the results of the gene synteny for the Z chromosomes. The red five-pointed stars indicate the species with Z-autosome fusion. (D) The scatter plots represent the number (x axis) and total length (y axis, in unit of kilobase) of synteny blocks between the W (masked repetitive sequence) and autosomes (in gray) and Z (in yellow), respectively, in four ZW systems.

Fig. 4.. The single-Z turnover model, a newly proposed mechanism of W origin.

Three conditions would be required for the occurrence of this mechanism. First, the W chromosome of a species has high homology with its own Z chromosome. Second, no Z-autosome fusion has occurred. Third, there is no homology between the W of this species and the W chromosomes generated through other mechanisms in closely related species. In such a situation, we suggest that the W chromosome of this species has been generated through a single-Z turnover mechanism.

Fig. 5.. The homology between the W of each species and each chromosome of a closely related species.

The synteny blocks between chromosomes serve as indicators of chromosome homology. They were estimated using Satsuma2synteny with the minimum alignment length set to the default value (0). The homology between the W of a species and each chromosome of its closely related species in (A) Nymphalidae, (B) Noctuidae and Bombycidae, and (C) two species including one Tortricidae and one Zygaenidae. In each scatter plot, both the W query and target genome are displayed with the vertical axis representing the total length of synteny blocks and the horizontal axis the number of synteny blocks. Each dot represents the number and total length of synteny blocks for a comparison between the indicated W chromosome with an autosome (gray dot), Z (yellow point), and W (red point) of the closely related species. The double-headed arrow lines connect the reciprocal comparisons between two species. For example, in the comparison between B. selene and D. iulia, one direction of comparison involves using the B. selene W as the query to search against each chromosome of D. iulia, while the other direction involves using the D. iulia W as the query to search against each chromosome of B. selene. The color code for the double-headed arrow lines represent bidirectional highest homology (green), unidirectional highest homology (blue), and bidirectional lack of homology between two W chromosomes in reciprocal comparisons (black). The scatter plots with a light green background represent the W chromosomes between the two species with the highest number and total length of synteny blocks. The tree on the left is from Fig. 3C.

Fig. 6.. Summary of the inferred origins of W chromosomes in species of Ditrysia.

The color code for the triangles under each branch of the phylogenetic tree represent the mechanism by which the W chromosome is generated. The blue rectangles under each branch represent the generation of neo-Z through Z-autosome fusion events. The composition of the sex chromosomes for each species is displayed above each branch. The W of C. pomonella is derived from Z-autosome fusion and single Z turnover. The W2 of M. jurtina and the W3 of A. cardamines were formed by single Z turnover. The W4 of P. brassicae were generated through Z-autosome fusion mechanism. The Ws of the other 13 ZW systems are likely the result of recruitment of a B chromosome or an unclear mechanism. The W chromosomes of B. selene, D. iulia, K. inachus, and V. cardui are named W1 (green rectangle), a single origin for W chromosomes of the species. Similarly, the W chromosomes named W5 (gray rectangle) in three species of Noctuidae evolved from a single ancestor, and the W chromosomes, named W6 (purple rectangle), from the two species of Crambidae are also derived from a common ancestor. The W chromosomes of the remaining four species are named Wi (black rectangle) to indicate that we are unsure whether their W chromosomes are derived from a common ancestor due to their lack of homology. In addition, we are uncertain whether the W chromosomes, including W1, W5, W6, and Wi, generated by the noncanonical model are derived from a single ancestor because of lack of homology between them. The right side of the phylogenetic tree displays the sex chromosome composition of each species, whether Z-autosome fusion has occurred, and whether the W and Z chromosomes within the genome are homologous. The phylogenetic tree is from Fig. 3C.