Repeatome Dynamics and Sex Chromosome Differentiation in the XY and XY1Y2 Systems of the Fish Hoplias malabaricus (Teleostei; Characiformes)

Abstract

The wolf fish Hoplias malabaricus is a Neotropical species characterized by remarkable karyotypic diversity, including seven karyomorphs (KarA-G) with distinct sex chromosome systems. This study investigated the homologous XY (KarF) and XY₁Y₂ (KarG) sex chromosome systems present in this species by integrating cytogenetics and genomics to examine sex chromosomes' composition through characterization of repeatome (satellite DNA and transposable elements) and sex-linked markers. Our analysis indicated that both karyomorphs are little differentiated in their sex chromosomes content revealed by satDNA mapping and putative sex-linked markers. Both repeatomes were mostly composed of transposable elements, but neither intra- (male versus female) nor interspecific (KarF x KarG) variations were found. In both systems, we demonstrated the occurrence of sex-specific sequences probably located on the non-recombining region of the Y chromosome supported by the accumulation of sex-specific haplotypes of HmfSat10-28/HmgSat31-28. This investigation offered valuable insights by highlighting the composition of homologous XY and XY₁Y₂ multiple sex chromosomes. Although homologous, the large Y chromosome in KarF corresponds to two separate linkage groups (Y₁ and Y₂) in KarG implying a specific meiotic arrangement involving the X chromosome in a meiotic trivalent chain. This scenario likely influenced recombination rates and, as a result, the genomic composition of these chromosomes.

Keywords: multiple sex systems; repeatome; single nucleotide polymorphism.

Figure 1

Geographic distribution and phylogenetic relationships of H. malabaricus karyomorphs. The map on the left illustrates the hydrographic basins in Brazil, indicating the karyomorph sampling locations in pink circles. The boxes highlight the corresponding karyomorphs, their distinct 2n, and sex chromosome morphologies. On the right, a phylogenetic tree proposes the evolutionary trajectory of the sex chromosome systems in these karyomorphs based on [8], with blue circles denoting key chromosomal events, following: (1) expansion of the X chromosome by repetitive DNA accumulation [7,9]; (2) sex chromosome–autosome translocation leading to multiple sex chromosomes [6,7]; and (3) conservation of the ancestral rearrangement (sex chromosome–autosome fusion) in a heterozygous condition resulting in multiple XX/XY₁Y₂ sex systems [10]. Legend on the lower right corner indicates the colors used in the idiograms for linkage groups and ribosomal DNA distribution, so as the main chromosomal events at the phylogeny branches.

Figure 2

Female (a–d) and male (e–h) metaphase plates of KarF of H. malabaricus highlighting the chromosomal location of HmfSatDNAs. While HmfSat07-149, HmfSat25-941, and HmfSat38-1394 showed positive hybridization signals in autosomes, HmfSat10-28 was clustered in the Y chromosome (f). The satDNA family names are indicated on the left top, in red (Atto550-dUTP-labeled). The asterisk symbol (*) denotes that HmfSat10-28 and HmgSat31-28 exhibit 100% homology. Bar = 5 μm.

Figure 3

Female (a–e) and male (f–j) metaphase plates of KarG of H. malabaricus highlighting the chromosomal location of HmgSatDNAs. All HmgSatDNAs showed positive hybridization signals in the autosomes. Additionally, HmgSat02-513 was found in the Y₂ (f) and HmgSat31-28 (HmfSat10-28) found in the X (h). The satDNA family names are indicated on the top left, in green (Atto488-dUTP-labeled) or red (Atto550-dUTP-labeled). The asterisk symbol (*) denotes that HmgSat31-28 and HmfSat10-28 exhibit 100% homology. Bar = 5 μm.

Figure 4

(A) Linear MST of HmfSat10-28 obtained from karyomorphs F (pink) and G (green) reads. Males are represented in dark green and dark pink, and females are represented in light green and light pink. Three major haplotypes were shared by both sexes of the two karyomorphs. Additionally, several abundant haplotypes were exclusively identified in KarF, indicating accumulation on the Y chromosome and lack of recombination with the X chromosome. Black dots indicate one nucleotide mutational step between each haplotype. (B) Alignment between HmfSat10-28 and HmgSat31-28.

Figure 5

Analysis of the DNApipeTE repeatome content in both karyomorphs (F and G) of Hoplias malabaricus. Distinct coverages of 0.1, 0.25, and 0.5× were employed for each karyomorph. The legend delineates the color scheme for each repetition class. The comprehensive results are provided in Table S4.