Satellitome Analysis of Adalia bipunctata (Coleoptera): Revealing Centromeric Turnover and Potential Chromosome Rearrangements in a Comparative Interspecific Study

Abstract

Eukaryotic genomes exhibit a dynamic interplay between single-copy sequences and repetitive DNA elements, with satellite DNA (satDNA) representing a substantial portion, mainly situated at telomeric and centromeric chromosomal regions. We utilized Illumina next-generation sequencing data from Adalia bipunctata to investigate its satellitome. Cytogenetic mapping via fluorescence in situ hybridization was performed for the most abundant satDNA families. In silico localization of satDNAs was carried out using the CHRISMAPP (Chromosome In Silico Mapping) pipeline on the high-fidelity chromosome-level assembly already available for this species, enabling a meticulous characterization and localization of multiple satDNA families. Additionally, we analyzed the conservation of the satellitome at an interspecific scale. Specifically, we employed the CHRISMAPP pipeline to map the satDNAs of A. bipunctata onto the genome of Adalia decempunctata, which has also been sequenced and assembled at the chromosome level. This analysis, along with the creation of a synteny map between the two species, suggests a rapid turnover of centromeric satDNA between these species and the potential occurrence of chromosomal rearrangements, despite the considerable conservation of their satellitomes. Specific satDNA families in the sex chromosomes of both species suggest a role in sex chromosome differentiation. Our interspecific comparative study can provide a significant advance in the understanding of the repeat genome organization and evolution in beetles.

Keywords: Adalia bipunctata; Adalia decempunctata; Chromosome In Silico Mapping (CHRISMAPP); Coccinellidae; Coleoptera; RepeatExplorer2; fluorescence in situ hybridization (FISH); karyotype evolution; ladybird beetle; satellite DNA; satellitome.

Figure 1

(A) Male mitotic metaphase and (B) karyotype of A. bipunctata. The karyotype consists of nine pairs of autosomes and the sex pair, an X chromosome and a dot-shaped Y chromosome. (C) Male mitotic metaphase and (D) karyotype after C-banding technique. Heterochromatin blocks are observed in the pericentromeric regions of all chromosomes except for the Y chromosome. (E) Meiotic metaphase after C-banding technique and subsequent DAPI staining (inverted image), with nine autosomal bivalents and the sex chromosomes in an Xy_p “parachute” shape. Arrows indicate the sex chromosomes. Bar = 5 μm.

Figure 2

DAPI staining (A) and FISH (B) on male meiotic metaphase I of A. bipunctata using a specific probe for the AbipSat01-187 satDNA, revealing positive hybridization signals (in red) on all autosomes and the two sex chromosomes. DAPI staining (C) and FISH (D) on male meiotic prophase showing the coincidence of the hybridization signals and the location of the DAPI-positive heterochromatic blocks. Bar = 5 μm.

Figure 3

A. bipunctata pseudochromosomes showing the distribution of different satDNA families obtained through the CHRISMAPP approach. For each chromosome, two schemes are displayed. The top one illustrates the distribution of the three most abundant satDNA families, while the bottom one shows the distribution of the remaining satDNA families. Asterisks indicate the presence of short arrays of the AbipSat01-187 and AbipSat03-8 out of the heterochromatic blocks.

Figure 4

DAPI staining (A) and FISH (B) on male meiotic metaphase I of A. bipunctata using a probe specific to the AbipSat03-8 satDNA, revealing positive hybridization signals (in red) on all chromosomes and the two sex chromosomes. DAPI staining (C) and FISH (D) on male meiotic prophase showing the coincidence of the hybridization signals and the location of the DAPI positive heterochromatic blocks. Bar = 5 μm.

Figure 5

A region of the A. bipunctata chromosome 1 heterochromatin block showing alternative arrays of AbipSat01-187 (red) and AbipSat03-8 (blue).

Figure 6

Double FISH using the AbipSat01-187 satDNA (labeled in red) and the satDNA AbipSat02-497 (labeled in green) as probes on A. bipunctata meiotic chromosomes. DAPI staining (A,C) and double FISH (B,D). On male meiotic metaphase I (B), positive hybridization signals with AbipSat01-187 are visible on the extremes of the bivalents (pericentromeric regions), the X and the Y chromosome. Hybridization signals with AbipSat02-497 are visible in the middle of the bivalents. On male meiotic prophase (D), hybridization signals with AbipSat01-187 are coincident with the DAPI positive heterochromatin, and the hybridization signals with AbipSat02-497 are located at one of the ends of the bivalents. Insert in C and D depicts an isolated bivalent showing the distribution of both satDNAs. Bar = 5 μm.

Figure 7

DAPI staining (A) and FISH (B) on male meiotic metaphase I of A. bipunctata using a probe for AbipSat04-193 (labeled in red) with two amplification rounds, revealing a diffuse labeling along the chromosomes. Bar = 5 μm.

Figure 8

CHRISMAPP analysis on unplaced scaffolds of the A. bipunctata genome, showing the presence of six scaffolds with satDNA AbipSat06-176. This satDNA is associated with satDNAs AbipSat01-178 and AbipSat03-8 in four of the unplaced scaffolds.

Figure 9

DAPI staining (A) and FISH (B) on male meiotic metaphase I of A. bipunctata using AbipSat06-176 as a probe. Hybridization signals (in red) are visible on three autosomal bivalents and the Y chromosome. Bar = 5 μm.

Figure 10

(A) Location of the satDNA AbipSat02-497 in the A. bipunctata and in the A. decempunctata genome assembly using CHRISMAPP. (B) Synteny map of the A. bipunctata and A. decempunctata genomes.