Pinniped Karyotype Evolution Substantiated by Comparative Chromosome Painting of 10 Pinniped Species (Pinnipedia, Carnivora)

Numerous Carnivora karyotype evolution investigations have been performed by classical and molecular cytogenetics and were supplemented by reconstructions of the Ancestral Carnivora Karyotype (ACK). However, the group of Pinnipedia was not studied in detail. Here we reconstruct pinniped karyotype evolution and refine ACK using published and our new painting data for 10 pinniped species. The combination of human (HSA) and domestic dog (CFA) whole-chromosome painting probes was used for the construction of the comparative chromosome maps for species from all three pinniped families: Odobenidae– Odobenus rosmarus Linnaeus, 1758, Phocidae – Phoca vitulina Linnaeus, 1758, Pusa sibirica Gmelin, 1788, Erignathus barbatus Erxleben, 1777, Phoca largha Pallas, 1811, Phoca hispida Schreber, 1775 and Otariidae – Eumetopias jubatus Schreber, 1775, Callorhinus ursinus Linnaeus, 1758, Phocarctos hookeri Gray, 1844, Arctocephalus forsteri Lesson, 1828. HSA and CFA autosome painting probes have delineated 32 and 68 conservative autosome segments in the studied genomes. The comparative painting in Pinnipedia supports monophyletic origin of pinnipeds, shows that pinniped karyotype evolution was characterized by slow rate of genome rearrangements (less then one rearrangement per 10 million years), provides strong support for refined structure of ACK with 2n = 38 and specifies plausible order of dog chromosome synthenic segments on ancestral Carnivora chromosomes. The heterochromatin, telomere and ribosomal DNA distribution was studied in all 10 species

Segmental paleotetraploidy revealed in sterlet (Acipenser ruthenus) genome by chromosome painting

Backgroun

Limited Lifespan of Fragile Regions in Mammalian Evolution

An important question in genome evolution is whether there exist fragile regions (rearrangement hotspots) where chromosomal rearrangements are happening over and over again. Although nearly all recent studies supported the existence of fragile regions in mammalian genomes, the most comprehensive phylogenomic study of mammals (Ma et al. (2006) Genome Research 16, 1557-1565) raised some doubts about their existence. We demonstrate that fragile regions are subject to a "birth and death" process, implying that fragility has limited evolutionary lifespan. This finding implies that fragile regions migrate to different locations in different mammals, explaining why there exist only a few chromosomal breakpoints shared between different lineages. The birth and death of fragile regions phenomenon reinforces the hypothesis that rearrangements are promoted by matching segmental duplications and suggests putative locations of the currently active fragile regions in the human genome

X Chromosome Evolution in Cetartiodactyla

The mammalian X chromosome is characterized by high level of conservation. On the contrary the Cetartiodactyl X chromosome displays variation in morphology and G-banding pattern. It is hypothesized that X chromosome has undergone multiple rearrangements during Cetartiodactyla speciation. To investigate the evolution of this sex chromosome we have selected 26 BAC clones from cattle CHORI-240 library evenly distributed along the cattle X chromosome. High-resolution maps were obtained by fluorescence in situ hybridisation in a representative range of cetartiodactyl species from different families: pig (Suidae), gray whale (Eschrichtiidae), pilot whale (Delphinidae), hippopotamus (Hippopotamidae), Java mouse deer (Tragulidae), pronghorn (Antilocapridae), Siberian musk deer (Moschidae), giraffe (Giraffidae). To trace the X chromosome evolution during fast radiation in speciose families, we mapped more than one species in Cervidae (moose, Siberian roe deer, fallow deer and Pere David’s deer) and Bovidae (musk ox, goat, sheep, sable antelope, nilgau, gaur, saola, and cattle). We have identified three major conserved synteny blocks and based on this data reconstructed the structure of putative ancestral cetartiodactyl X chromosome. We demonstrate that intrachromosomal rearrangements such as inversions and centromere reposition are main drivers of cetartiodactyl’s chromosome X evolution

X Chromosome Evolution in Cetartiodactyla

The phenomenon of a remarkable conservation of the X chromosome in eutherian mammals has been first described by Susumu Ohno in 1964. A notable exception is the cetartiodactyl X chromosome, which varies widely in morphology and G-banding pattern between species. It is hypothesized that this sex chromosome has undergone multiple rearrangements that changed the centromere position and the order of syntenic segments over the last 80 million years of Cetartiodactyla speciation. To investigate its evolution we have selected 26 evolutionarily conserved bacterial artificial chromosome (BAC) clones from the cattle CHORI-240 library evenly distributed along the cattle X chromosome. High-resolution BAC maps of the X chromosome on a representative range of cetartiodactyl species from different branches: pig (Suidae), alpaca (Camelidae), gray whale (Cetacea), hippopotamus (Hippopotamidae), Java mouse-deer (Tragulidae), pronghorn (Antilocapridae), Siberian musk deer (Moschidae), and giraffe (Giraffidae) were obtained by fluorescent in situ hybridization. To trace the X chromosome evolution during fast radiation in specious families, we performed mapping in several cervids (moose, Siberian roe deer, fallow deer, and Pere David’s deer) and bovid (muskox, goat, sheep, sable antelope, and cattle) species. We have identified three major conserved synteny blocks and rearrangements in different cetartiodactyl lineages and found that the recently described phenomenon of the evolutionary new centromere emergence has taken place in the X chromosome evolution of Cetartiodactyla at least five times. We propose the structure of the putative ancestral cetartiodactyl X chromosome by reconstructing the order of syntenic segments and centromere position for key groups

Evolution of gene regulation in ruminants differs between evolutionary breakpoint regions and homologous synteny blocks

The role of chromosome rearrangements in driving evolution has been a long-standing question of evolutionary biology. Here we focused on ruminants as a model to assess how rearrangements may have contributed to the evolution of gene regulation. Using reconstructed ancestral karyotypes of Cetartiodactyls, Ruminants, Pecorans, and Bovids, we traced patterns of gross chromosome changes. We found that the lineage leading to the ruminant ancestor after the split from other cetartiodactyls was characterized by mostly intrachromosomal changes, whereas the lineage leading to the pecoran ancestor (including all livestock ruminants) included multiple interchromosomal changes. We observed that the liver cell putative enhancers in the ruminant evolutionary breakpoint regions are highly enriched for DNA sequences under selective constraint acting on lineage-specific transposable elements (TEs) and a set of 25 specific transcription factor (TF) binding motifs associated with recently active TEs. Coupled with gene expression data, we found that genes near ruminant breakpoint regions exhibit more divergent expression profiles among species, particularly in cattle, which is consistent with the phylogenetic origin of these breakpoint regions. This divergence was significantly greater in genes with enhancers that contain at least one of the 25 specific TF binding motifs and located near bovidae-to-cattle lineage breakpoint regions. Taken together, by combining ancestral karyotype reconstructions with analysis of cis regulatory element and gene expression evolution, our work demonstrated that lineage-specific regulatory elements colocalized with gross chromosome rearrangements may have provided valuable functional modifications that helped to shape ruminant evolution

An integrated chromosome-scale genome assembly of the Masai giraffe (Giraffa camelopardalis tippelskirchi)

Background The Masai giraffe (Giraffa camelopardalis tippelskirchi) is the largest-bodied giraffe and the world's tallest terrestrial animal. With its extreme size and height, the giraffe's unique anatomical and physiological adaptations have long been of interest to diverse research fields. Giraffes are also critical to ecosystems of sub-Saharan Africa, with their long neck serving as a conduit to food sources not shared by other herbivores. Although the genome of a Masai giraffe has been sequenced, the assembly was highly fragmented and suboptimal for genome analysis. Herein we report an improved giraffe genome assembly to facilitate evolutionary analysis of the giraffe and other ruminant genomes. Findings Using SOAPdenovo2 and 170 Gbp of Illumina paired-end and mate-pair reads, we generated a 2.6-Gbp male Masai giraffe genome assembly, with a scaffold N50 of 3 Mbp. The incorporation of 114.6 Gbp of Chicago library sequencing data resulted in a HiRise SOAPdenovo + Chicago assembly with an N50 of 48 Mbp and containing 95% of expected genes according to BUSCO analysis. Using the Reference-Assisted Chromosome Assembly tool, we were able to order and orient scaffolds into 42 predicted chromosome fragments (PCFs). Using fluorescence in situ hybridization, we placed 153 cattle bacterial artificial chromosomes onto giraffe metaphase spreads to assess and assign the PCFs on 14 giraffe autosomes and the X chromosome resulting in the final assembly with an N50 of 177.94 Mbp. In this assembly, 21,621 protein-coding genes were identified using both de novo and homology-based predictions. Conclusions We have produced the first chromosome-scale genome assembly for a Giraffidae species. This assembly provides a valuable resource for the study of artiodactyl evolution and for understanding the molecular basis of the unique adaptive traits of giraffes. In addition, the assembly will provide a powerful resource to assist conservation efforts of Masai giraffe, whose population size has declined by 52% in recent years

Comparative Chromosome Mapping of Musk Ox and the X Chromosome among Some Bovidae Species

Bovidae, the largest family in Pecora infraorder, are characterized by a striking variability in diploid number of chromosomes between species and among individuals within a species. The bovid X chromosome is also remarkably variable, with several morphological types in the family. Here we built a detailed chromosome map of musk ox (Ovibos moschatus), a relic species originating from Pleistocene megafauna, with dromedary and human probes using chromosome painting. We trace chromosomal rearrangements during Bovidae evolution by comparing species already studied by chromosome painting. The musk ox karyotype differs from the ancestral pecoran karyotype by six fusions, one fission, and three inversions. We discuss changes in pecoran ancestral karyotype in the light of new painting data. Variations in the X chromosome structure of four bovid species nilgai bull (Boselaphus tragocamelus), saola (Pseudoryx nghetinhensis), gaur (Bos gaurus), and Kirk’s Dikdik (Madoqua kirkii) were further analyzed using 26 cattle BAC-clones. We found the duplication on the X in saola. We show main rearrangements leading to the formation of four types of bovid X: Bovinae type with derived cattle subtype formed by centromere reposition and Antilopinae type with Caprini subtype formed by inversion in XSB1

The Case of X and Y Localization of Nucleolus Organizer Regions (NORs) in Tragulus javanicus (Cetartiodactyla, Mammalia)

There are differences in number and localization of nucleolus organizer regions (NORs) in genomes. In mammalian genomes, NORs are located on autosomes, which are often situated on short arms of acrocentric chromosomes and more rarely in telomeric, pericentromeric, or interstitial regions. In this work, we report the unique case of active NORs located on gonоsomes of a eutherian mammal, the Javan mouse-deer (Tragulus javanicus). We have investigated the position of NORs by FISH experiments with ribosomal DNA (rDNA) sequences (18S, 5.8S, and 28S) and show the presence of a single NOR site on the X and Y chromosomes. The NOR is localized interstitially on the p-arm of the X chromosome in close proximity with prominent C-positive heterochromatin blocks and in the pericentromeric area of mostly heterochromatic Y. The NOR sites are active on both the X and Y chromosomes in the studied individual and surrounded by GC enriched heterochromatin. We hypothesize that the surrounding heterochromatin might have played a role in the transfer of NORs from autosomes to sex chromosomes during the karyotype evolution of the Javan mouse-deer