The standard chromosome set of individuals includes autosomes and, frequently, sex chromosomes. However, certain species harbor non-essential chromosomes known as B chromosomes, which can influence cell metabolism by affecting gene expression. Studies in Drosophila melanogaster identified B chromosomes that are predominantly heterochromatic and likely originated from Chromosome 4. Here we advanced in detail about the repetitive composition of B chromosomes and impact of B chromosomes on the expression of genes and Transposable Elements (TEs) located on standard chromosomes. Using tools like RepeatExplorer and RepeatMasker, we quantified the abundance and divergence of repetitive sequences from genomes B+/0B and from the B chromosome isolated by flow cytometry. Fluorescent in situ hybridization (FISH) was subsequently used to understand sequence-sharing patterns between B and standard chromosomes. RSEM, TEspeX, Deseq2, and EdgeR were used to quantify and perform differential expression analysis of coding transcripts and TEs.

Overall, the isolated B chromosome is composed of approximately 17.04% TEs, 16.88% satellite DNAs (satDNAs) and microsatellites, and 1.9% multigene families, totaling 35.82% of repetitive composition. FISH results showed no sequence was shared between the Bs and other chromosomes besides 4th, supporting the B chromosome's origin from the 4th chromosome. Concerning the expression analysis, the addition of a B chromosome to the 0B genome exhibited a minimal immediate effect on coding transcripts (0.8%), which after approximately 30 generations resulted in a substantial increased effect (7%). Interestingly, TEs displayed differential expression following B chromosome addition (7.3%), maintaining a consistent effect (8.5%) over generations. Moreover, TEs were more susceptible to B dose (5.8% by B dose and 12.4% by presence) compared to coding transcripts (0.7% altered by B dose and 9.8% altered by presence). These findings shed light on the dynamic interactions between B chromosomes and the genomic landscape, offering insights into their role in gene and TE expression modulation.