The Evolution and Functional Significance of Nested Gene Structures in Drosophila melanogaster

Nearly ten percent of the genes in the genome of Drosophila melanogaster are in nested structures, in which one gene is completely nested within the intron of another gene (nested and including gene, respectively). Even though the coding sequences and UTRs of these nested/including gene pairs do not overlap, their intimate structures and the possibility of shared regulatory sequences raise questions about the evolutionary forces governing the origination, and subsequent functional and evolutionary impacts of these structures. In this study, we show that nested genes experience weaker evolutionary constraint, have faster rates of protein evolution and are expressed in fewer tissues than other genes, while including genes show the opposite patterns. Surprisingly, despite completely overlapping with each other, nested and including genes are less likely to display correlated gene expression and biological function than the nearby yet non-overlapping genes. Interestingly, significantly fewer nested genes are transcribed from the same strand as the including gene. We found that same-strand nested genes are more likely to be single-exon genes. In addition, same-strand including genes are less likely to have known lethal or sterile phenotypes than opposite-strand including genes only when the corresponding nested genes have introns. These results support our hypothesis that selection against potential erroneous mRNA splicing when nested and including genes are on the same strand plays an important role in the evolution of nested gene structures

Phylogenomic Analysis Reveals Dynamic Evolutionary History of the Drosophila Heterochromatin Protein 1 (HP1) Gene Family

Heterochromatin is the gene-poor, satellite-rich eukaryotic genome compartment that supports many essential cellular processes. The functional diversity of proteins that bind and often epigenetically define heterochromatic DNA sequence reflects the diverse functions supported by this enigmatic genome compartment. Moreover, heterogeneous signatures of selection at chromosomal proteins often mirror the heterogeneity of evolutionary forces that act on heterochromatic DNA. To identify new such surrogates for dissecting heterochromatin function and evolution, we conducted a comprehensive phylogenomic analysis of the Heterochromatin Protein 1 gene family across 40 million years of Drosophila evolution. Our study expands this gene family from 5 genes to at least 26 genes, including several uncharacterized genes in Drosophila melanogaster. The 21 newly defined HP1s introduce unprecedented structural diversity, lineage-restriction, and germline-biased expression patterns into the HP1 family. We find little evidence of positive selection at these HP1 genes in both population genetic and molecular evolution analyses. Instead, we find that dynamic evolution occurs via prolific gene gains and losses. Despite this dynamic gene turnover, the number of HP1 genes is relatively constant across species. We propose that karyotype evolution drives at least some HP1 gene turnover. For example, the loss of the male germline-restricted HP1E in the obscura group coincides with one episode of dramatic karyotypic evolution, including the gain of a neo-Y in this lineage. This expanded compendium of ovary- and testis-restricted HP1 genes revealed by our study, together with correlated gain/loss dynamics and chromosome fission/fusion events, will guide functional analyses of novel roles supported by germline chromatin

Widespread Polymorphism in the Positions of Stop Codons in Drosophila melanogaster

The mechanisms underlying evolutionary changes in protein length are poorly understood. Protein domains are lost and gained between species and must have arisen first as within-species polymorphisms. Here, we use Drosophila melanogaster population genomic data combined with between species divergence information to understand the evolutionary forces that generate and maintain polymorphisms causing changes in protein length in D. melanogaster. Specifically, we looked for protein length variations resulting from premature termination codons (PTCs) and stop codon losses (SCLs). We discovered that 438 genes contained polymorphisms resulting in truncation of the translated region (PTCs) and 119 genes contained polymorphisms predicted to lengthen the translated region (SCLs). Stop codon polymorphisms (SCPs) (especially PTCs) appear to be more deleterious than other polymorphisms, including protein amino acid changes. Genes harboring SCPs are in general less selectively constrained, more narrowly expressed, and enriched for dispensable biological functions. However, we also observed exceptional cases such as genes that have multiple independent SCPs, alleles that are shared between D. melanogaster and Drosophila simulans, and high-frequency alleles that cause extreme changes in gene length. SCPs likely have an important role in the evolution of these genes

The Role of <i>piRNA</i>-Mediated Epigenetic Silencing in the Population Dynamics of Transposable Elements in <i>Drosophila melanogaster</i>

The piwi-interacting RNAs (piRNA) are small RNAs that target selfish transposable elements (TEs) in many animal genomes. Until now, piRNAs’ role in TE population dynamics has only been discussed in the context of their suppression of TE transposition, which alone is not sufficient to account for the skewed frequency spectrum and stable containment of TEs. On the other hand, euchromatic TEs can be epigenetically silenced via piRNA-dependent heterochromatin formation and, similar to the widely known “Position-effect variegation”, heterochromatin induced by TEs can “spread” into nearby genes. We hypothesized that the piRNA-mediated spread of heterochromatin from TEs into adjacent genes has deleterious functional effects and leads to selection against individual TEs. Unlike previously identified deleterious effects of TEs due to the physical disruption of DNA, the functional effect we investigated here is mediated through the epigenetic influences of TEs. We found that the repressive chromatin mark, H3K9me, is elevated in sequences adjacent to euchromatic TEs at multiple developmental stages in Drosophila melanogaster. Furthermore, the heterochromatic states of genes depend not only on the number of and distance from adjacent TEs, but also on the likelihood that their nearest TEs are targeted by piRNAs. These variations in chromatin status probably have functional consequences, causing genes near TEs to have lower expression. Importantly, we found stronger selection against TEs that lead to higher H3K9me enrichment of adjacent genes, demonstrating the pervasive evolutionary consequences of TE-induced epigenetic silencing. Because of the intrinsic biological mechanism of piRNA amplification, spread of TE heterochromatin could result in the theoretically required synergistic deleterious effects of TE insertions for stable containment of TE copy number. The indirect deleterious impact of piRNA-mediated epigenetic silencing of TEs is a previously unexplored, yet important, element for the evolutionary dynamics of TEs

RESEARCH ARTICLE The Role of piRNA-Mediated Epigenetic Silencing in the Population Dynamics of Transposable Elements in Drosophila melanogaster

The piwi-interacting RNAs (piRNA) are small RNAs that target selfish transposable ele-ments (TEs) in many animal genomes. Until now, piRNAs ’ role in TE population dynamics has only been discussed in the context of their suppression of TE transposition, which alone is not sufficient to account for the skewed frequency spectrum and stable containment of TEs. On the other hand, euchromatic TEs can be epigenetically silenced via piRNA-de-pendent heterochromatin formation and, similar to the widely known “Position-effect varie-gation”, heterochromatin induced by TEs can “spread ” into nearby genes. We hypothesized that the piRNA-mediated spread of heterochromatin from TEs into adjacent genes has dele-terious functional effects and leads to selection against individual TEs. Unlike previously identified deleterious effects of TEs due to the physical disruption of DNA, the functional ef-fect we investigated here is mediated through the epigenetic influences of TEs. We found that the repressive chromatin mark, H3K9me, is elevated in sequences adjacent to euchro-matic TEs at multiple developmental stages in Drosophila melanogaster. Furthermore, the heterochromatic states of genes depend not only on the number of and distance from adja

The Role of <i>piRNA</i>-Mediated Epigenetic Silencing in the Population Dynamics of Transposable Elements in <i>Drosophila melanogaster</i>

<div><p>The <i>piwi</i>-interacting RNAs (<i>piRNA</i>) are small RNAs that target selfish transposable elements (TEs) in many animal genomes. Until now, <i>piRNAs’</i> role in TE population dynamics has only been discussed in the context of their suppression of TE transposition, which alone is not sufficient to account for the skewed frequency spectrum and stable containment of TEs. On the other hand, euchromatic TEs can be epigenetically silenced via <i>piRNA</i>-dependent heterochromatin formation and, similar to the widely known “Position-effect variegation”, heterochromatin induced by TEs can “spread” into nearby genes. We hypothesized that the <i>piRNA</i>-mediated spread of heterochromatin from TEs into adjacent genes has deleterious functional effects and leads to selection against individual TEs. Unlike previously identified deleterious effects of TEs due to the physical disruption of DNA, the functional effect we investigated here is mediated through the epigenetic influences of TEs. We found that the repressive chromatin mark, H3K9me, is elevated in sequences adjacent to euchromatic TEs at multiple developmental stages in <i>Drosophila melanogaster</i>. Furthermore, the heterochromatic states of genes depend not only on the number of and distance from adjacent TEs, but also on the likelihood that their nearest TEs are targeted by piRNAs. These variations in chromatin status probably have functional consequences, causing genes near TEs to have lower expression. Importantly, we found stronger selection against TEs that lead to higher H3K9me enrichment of adjacent genes, demonstrating the pervasive evolutionary consequences of TE-induced epigenetic silencing. Because of the intrinsic biological mechanism of <i>piRNA</i> amplification, spread of TE heterochromatin could result in the theoretically required synergistic deleterious effects of TE insertions for stable containment of TE copy number. The indirect deleterious impact of <i>piRNA</i>-mediated epigenetic silencing of TEs is a previously unexplored, yet important, element for the evolutionary dynamics of TEs.</p></div

Differences in expression levels between alleles with and without TEs.

<p>1. Median for the differences in expression rank between alleles with and without TEs. Rank differences were calculated as the mean expression rank of “with TE” alleles minus that of “without TE” alleles. A positive rank difference means “with TE” alleles have larger expression rank (and thus lower expression) than “without TE” alleles.</p><p>2. The number of genes whose “with TE” alleles have significantly (permutation <i>p-value</i> < 0.05) larger expression rank (i.e. lower expression) than “without TE” alleles.</p><p>3. The expected number of genes whose “with TE” alleles have significantly larger expression rank (i.e. lower expression) than “without TE” alleles under the null hypothesis with 5% false positive rate.</p><p>Differences in expression levels between alleles with and without TEs.</p