Over the last few decades, an increasing number of vertebrate taxa have been identified that undergo programmed genome rearrangement, or programmed DNA loss, during development. In these organisms, the genome of germ cells is often reproducibly different from the genome of all other cells within the body. Although we clearly have not identified all vertebrate taxa that undergo programmed genome loss, the list of species known to undergo loss now represents ∼10% of vertebrate species, including several basally diverging lineages. Recent studies have shed new light on the targets and mechanisms of DNA loss and their association with canonical modes of DNA silencing. Ultimately, expansion of these studies into a larger collection of taxa will aid in reconstructing patterns of shared/independent ancestry of programmed DNA loss in the vertebrate lineage, as well as more recent evolutionary events that have shaped the structure and content of eliminated DNA.

Aedes aegypti, the primary vector of dengue, yellow fever and Zika flaviviruses, consists of at least two subspecies. Aedes aegypti (Aaa) is light in color, has pale scales on the first abdominal tergite, oviposits in artificial containers, and preferentially feeds on humans. Aedes aegypti formosus (Aaf), has a dark cuticle, is restricted to sub-Saharan Africa, has no pale scales on the first abdominal tergite and frequently oviposits in natural containers. Scale patterns correlate with cuticle color in East Africa but not in Senegal, West Africa where black cuticle mosquitoes display a continuum of scaling patterns and breed domestically indoors. An earlier laboratory study did not indicate any pre- or postzygotic barriers to gene flow between Aaa and Aaf in East Africa. However, similar attempts to construct F₁ intercross families between Aaa laboratory strains and Senegal Ae. aegypti (SenAae) failed due to poor F₁ oviposition and low F₂ egg-to-adult survival. Insemination and assortative mating experiments failed to identify prezygotic mating barriers. Backcrosses were performed to test for postzygotic isolation patterns consistent with Haldane’s rule modified for species, like Aedes, that have an autosomal sex determining locus (SDL). Egg-pupal survival was predicted to be low in females mated to hybrid F₁ males but average when a male mates with a hybrid F₁ female. Survival was in fact significantly reduced when females mated to hybrid males but egg-pupal survival was significantly increased when males were mated to hybrid F₁ females. These observations are therefore inconclusive with regards to Haldane’s rule. Basic cytogenetic analyses and Fluorescent In Situ Hybridization (FISH) experiments were performed to compare SenAae strains with the IB12 strain of Aaa that was used for genome sequencing and physical mapping. Some SenAae strains had longer chromosomes than IB12 and significantly different centromeric indices on chromosomes 1 and 3. DAPI staining was used to identify AT-rich regions, chromomycin A3 following pretreatment with barium hydroxide stained for GC-rich regions and stained the ribosomal RNA locus and YOYO-1 was used to test for differential staining. Chromosome patterns in SenAae strains revealed by these three stains differed from those in IB12. For FISH, 40 BAC clones previously physically mapped on Aaa chromosomes were used to test for chromosome rearrangements in SenAae relative to IB12. Differences in the order of markers identified two chromosomal rearrangements between IB12 and SenAae strains. The first rearrangement involves two overlapping pericentric (containing the centromere) inversions in chromosome 3 or an insertion of a large fragment into the 3q arm. The second rearrangement is close to the centromere on the p arm of chromosome 2. Linkage analysis of the SDL and the white-eye locus identified a likely chromosomal rearrangement on chromosome 1. The reproductive incompatibility observed within SenAae and between SenAae and Aaa may be generally associated with chromosome rearrangements on all three chromosomes and specifically caused by pericentric inversions on chromosomes 2 and 3. Aedes aegypti is one of the best studied mosquito species and it is the principal vector of dengue, Zika, and yellow fever flaviviruses and the Chikungunya alphavirus. Aedes aegypti occurs throughout all tropical and subtropical regions of the world, and previous population genetic studies have shown that the highest genetic diversity occurs in Africa. Aedes aegypti from Senegal, West Africa (SenAae) have a low oviposition rate; those that do oviposit have a low fecundity and poor egg-to-adult survival. Furthermore rearrangements were detected on all three chromosomes in SenAae. These observations are consistent with the presence of at least two cryptic subspecies of Ae. aegypti in Senegal arising from reproductive isolation due to chromosome rearrangements. Genetic control strategies are being considered for the suppression of Ae. aegypti populations worldwide. Barriers to gene flow in African Ae. aegypti populations could compromise these future control efforts.

The sea lamprey (Petromyzon marinus) is one of few vertebrate species known to reproducibly eliminate large fractions of its genome during normal embryonic development. This germline-specific DNA is lost in the form of large fragments, including entire chromosomes, and available evidence suggests that DNA elimination acts as a permanent silencing mechanism that prevents the somatic expression of a specific subset of “germline” genes. However, reconstruction of eliminated regions has proven to be challenging due to the complexity of the lamprey karyotype. We applied an integrative approach aimed at further characterization of the large-scale structure of eliminated segments, including: (1) in silico identification of germline-enriched repeats; (2) mapping the chromosomal location of specific repetitive sequences in germline metaphases; and (3) 3D DNA/DNA-hybridization to embryonic lagging anaphases, which permitted us to both verify the specificity of elements to physically eliminated chromosomes and characterize the subcellular organization of these elements during elimination. This approach resulted in the discovery of several repetitive elements that are found exclusively on the eliminated chromosomes, which subsequently permitted the identification of 12 individual chromosomes that are programmatically eliminated during early embryogenesis. The fidelity and specificity of these highly abundant sequences, their distinctive patterning in eliminated chromosomes, and subcellular localization in elimination anaphases suggest that these sequences might contribute to the specific targeting of chromosomes for elimination or possibly in molecular interactions that mediate their decelerated poleward movement in chromosome elimination anaphases, isolation into micronuclei and eventual degradation.

Heterochromatin is identified as a potential factor driving diversification of species. To understand the magnitude of heterochromatin variation within the Anopheles gambiae complex of malaria mosquitoes, we analyzed metaphase chromosomes in An. arabiensis, An. coluzzii, An. gambiae, An. merus, and An. quadriannulatus. Using fluorescence in situ hybridization (FISH) with ribosomal DNA (rDNA), a highly repetitive fraction of DNA, and heterochromatic Bacterial Artificial Chromosome (BAC) clones, we established the correspondence of pericentric heterochromatin between the metaphase and polytene X chromosomes of An. gambiae. We then developed chromosome idiograms and demonstrated that the X chromosomes exhibit qualitative differences in their pattern of heterochromatic bands and position of satellite DNA (satDNA) repeats among the sibling species with postzygotic isolation, An. arabiensis, An. merus, An. quadriannulatus, and An. coluzzii or An. gambiae. The identified differences in the size and structure of the X chromosome heterochromatin point to a possible role of repetitive DNA in speciation of mosquitoes. We found that An. coluzzii and An. gambiae, incipient species with prezygotic isolation, share variations in the relative positions of the satDNA repeats and the proximal heterochromatin band on the X chromosomes. This previously unknown genetic polymorphism in malaria mosquitoes may be caused by a differential amplification of DNA repeats or an inversion in the sex chromosome heterochromatin.

As the only surviving lineages of jawless fishes, hagfishes and lampreys provide a critical window into early vertebrate evolution. Here, we investigate the complex history, timing, and functional role of genome-wide duplications in vertebrates in the light of a chromosome-scale genome of the brown hagfish Eptatretus atami . Using robust chromosome-scale (paralogon-based) phylogenetic methods, we confirm the monophyly of cyclostomes, document an auto-tetraploidization (1R _V ) that predated the origin of crown group vertebrates ∼517 Mya, and establish the timing of subsequent independent duplications in the gnathostome and cyclostome lineages. Some 1R _V gene duplications can be linked to key vertebrate innovations, suggesting that this early genomewide event contributed to the emergence of pan-vertebrate features such as neural crest. The hagfish karyotype is derived by numerous fusions relative to the ancestral cyclostome arrangement preserved by lampreys. These genomic changes were accompanied by the loss of genes essential for organ systems (eyes, osteoclast) that are absent in hagfish, accounting in part for the simplification of the hagfish body plan; other gene family expansions account for hagfishes’ capacity to produce slime. Finally, we characterise programmed DNA elimination in somatic cells of hagfish, identifying protein-coding and repetitive elements that are deleted during development. As in lampreys, the elimination of these genes provides a mechanism for resolving genetic conflict between soma and germline by repressing germline/pluripotency functions. Reconstruction of the early genomic history of vertebrates provides a framework for further exploration of vertebrate novelties.

Many eukaryotic species undergo programmed elimination of specific chromosomes during embryogenesis, typically retaining these chromosomes only in their germ cells. In some species, programmatic elimination of GRCs, or sex chromosomes, also occurs in a sex-specific manner, with specific chromosomes being transmitted or eliminated by only one sex. As such, these chromosomes provide a unique perspective on the evolution of gene functions that are advantageous to the germline and genetic tradeoffs between somatic vs germline or oocyte vs sperm biology. While GRCs have been extensively characterized in male sea lampreys ( Petromyzon marinus ), the status of GRCs in females has not yet been resolved, though it has been hypothesized that male-specific expression/transmission of these chromosomes might provide a solution to resolving the long-standing mystery of lamprey sex determining mechanisms. To gain insight into the roles of GRCs in female lampreys, we performed several karyological, transcriptomic, and genomic analyses, which demonstrate that GRCs are present in the female lamprey germline, transmitted by oocytes and somatically eliminated in both sexes. These analyses also show that GRCs play important roles in the maintenance and development of female germline but provide no evidence for sex-specific variation in the elimination and transmission of lamprey GRCs. These findings underscore the diversity of germline functions that are carried out by GRCs in both male and female lampreys and highlight the fact that sex-specific transmission/retention of GRCs likely follows no universal rules across the diverse lineages that have independently evolved to undergo developmentally programmed DNA elimination.