Invasion and Amplification of Endogenous Retroviruses in Dasyuridae Marsupial Genomes.

Retroviruses are an ancient viral family that have globally coevolved with vertebrates and impacted their evolution. In Australia, a continent that has been geographically isolated for millions of years, little is known about retroviruses in wildlife, despite the devastating impacts of a retrovirus on endangered koala populations. We therefore sought to identify and characterize Australian retroviruses through reconstruction of endogenous retroviruses from marsupial genomes, in particular the Tasmanian devil due to its high cancer incidence. We screened 19 marsupial genomes and identified over 80,000 endogenous retrovirus fragments which we classified into eight retrovirus clades. The retroviruses were similar to either Betaretrovirus (5/8) or Gammaretrovirus (3/8) retroviruses, but formed distinct phylogenetic clades compared to extant retroviruses. One of the clades (MEBrv 3) lost an envelope but retained retrotranspositional activity, subsequently amplifying throughout all Dasyuridae genomes. Overall, we provide insights into Australian retrovirus evolution and identify a highly active endogenous retrovirus within Dasyuridae genomes.

The admixed brushtail possum genome reveals invasion history in New Zealand and novel imprinted genes.

Combining genome assembly with population and functional genomics can provide valuable insights to development and evolution, as well as tools for species management. Here, we present a chromosome-level genome assembly of the common brushtail possum (Trichosurus vulpecula), a model marsupial threatened in parts of their native range in Australia, but also a major introduced pest in New Zealand. Functional genomics reveals post-natal activation of chemosensory and metabolic genes, reflecting unique adaptations to altricial birth and delayed weaning, a hallmark of marsupial development. Nuclear and mitochondrial analyses trace New Zealand possums to distinct Australian subspecies, which have subsequently hybridised. This admixture allowed phasing of parental alleles genome-wide, ultimately revealing at least four genes with imprinted, parent-specific expression not yet detected in other species (MLH1, EPM2AIP1, UBP1 and GPX7). We find that reprogramming of possum germline imprints, and the wider epigenome, is similar to eutherian mammals except onset occurs after birth. Together, this work is useful for genetic-based control and conservation of possums, and contributes to understanding of the evolution of novel mammalian epigenetic traits.

Between the Devil and the Deep Blue Sea: Non-Coding RNAs Associated with Transmissible Cancers in Tasmanian Devil, Domestic Dog and Bivalves.

Currently there are nine known examples of transmissible cancers in nature. They have been observed in domestic dog, Tasmanian devil, and six bivalve species. These tumours can overcome host immune defences and spread to other members of the same species. Non-coding RNAs (ncRNAs) are known to play roles in tumorigenesis and immune system evasion. Despite their potential importance in transmissible cancers, there have been no studies on ncRNA function in this context to date. Here, we present possible applications of the CRISPR/Cas system to study the RNA biology of transmissible cancers. Specifically, we explore how ncRNAs may play a role in the immortality and immune evasion ability of these tumours.

Pseudogenes: A Novel Source of Trans-Acting Antisense RNAs.

Several recent studies support a functional role for pseudogenes, a copy of a parent gene that has lost protein-coding potential, which was for a long time thought to represent only "junk" DNA. Several hundreds of pseudogenes have now been reported as transcribed RNAs in a large variety of tissues and tumor types. Most studies have focused on pseudogenes expressed in sense direction, relative to their protein-coding gene counterpart, but some reports suggest that pseudogenes can be also transcribed as antisense RNAs (asRNAs). Key regulatory genes, such as PTEN and OCT4, have in fact been reported to be under the regulation of pseudogene-expressed asRNAs. Here, we review what is known about pseudogene-expressed asRNAs, we discuss the functional role that these transcripts may have in gene regulation and we summarize the techniques that are available to study them.

Evolution of the Antisense Overlap between Genes for Thyroid Hormone Receptor and Rev-erbα and Characterization of an Exonic G-Rich Element That Regulates Splicing of TRα2 mRNA.

The α-thyroid hormone receptor gene (TRα) codes for two functionally distinct proteins: TRα1, the α-thyroid hormone receptor; and TRα2, a non-hormone-binding variant. The final exon of TRα2 mRNA overlaps the 3' end of Rev-erbα mRNA, which encodes another nuclear receptor on the opposite strand of DNA. To understand the evolution of this antisense overlap, we sequenced these genes and mRNAs in the platypus Orthorhynchus anatinus. Despite its strong homology with other mammals, the platypus TRα/Rev-erbα locus lacks elements essential for expression of TRα2. Comparative analysis suggests that alternative splicing of TRα2 mRNA expression evolved in a stepwise fashion before the divergence of eutherian and marsupial mammals. A short G-rich element (G30) located downstream of the alternative 3'splice site of TRα2 mRNA and antisense to the 3'UTR of Rev-erbα plays an important role in regulating TRα2 splicing. G30 is tightly conserved in eutherian mammals, but is absent in marsupials and monotremes. Systematic deletions and substitutions within G30 have dramatically different effects on TRα2 splicing, leading to either its inhibition or its enhancement. Mutations that disrupt one or more clusters of G residues enhance splicing two- to three-fold. These results suggest the G30 sequence can adopt a highly structured conformation, possibly a G-quadruplex, and that it is part of a complex splicing regulatory element which exerts both positive and negative effects on TRα2 expression. Since mutations that strongly enhance splicing in vivo have no effect on splicing in vitro, it is likely that the regulatory role of G30 is mediated through linkage of transcription and splicing.

Does the human X contain a third evolutionary block? Origin of genes on human Xp11 and Xq28.

Comparative gene mapping of human X-borne genes in marsupials defined an ancient conserved region and a recently added region of the eutherian X, and the separate evolutionary origins of these regions was confirmed by their locations on chicken chromosomes 4p and 1q, respectively. However, two groups of genes, from the pericentric region of the short arm of the human X (at Xp11) and a large group of genes from human Xq28, were thought to be part of a third evolutionary block, being located in a single region in fish, but mapping to chicken chromosomes other than 4p and 1q. We tested this hypothesis by comparative mapping of genes in these regions. Our gene mapping results show that human Xp11 genes are located on the marsupial X chromosome and platypus chromosome 6, indicating that the Xp11 region was part of original therian X chromosome. We investigated the evolutionary origin of genes from human Xp11 and Xq28, finding that chicken paralogs of human Xp11 and Xq28 genes had been misidentified as orthologs, and their true orthologs are represented in the chicken EST database, but not in the current chicken genome assembly. This completely undermines the evidence supporting a separate evolutionary origin for this region of the human X chromosome, and we conclude, instead, that it was part of the ancient autosome, which became the conserved region of the therian X chromosome 166 million years ago.

LINE-1 amplification accompanies explosive genome repatterning in rodents.

Transposable elements (TEs) sometimes induce karyotypic changes following recombination, breakage and rearrangement. We used FISH and Southern blot analyses to investigate the amount and distribution of LINE-1 retrotransposons in rodents (genus Taterillus, Muridae, Gerbillinae) that have recently undergone an important genome repatterning. Our results were interpreted in a known phylogenetic framework and clearly showed that LINE-1 elements were greatly amplified and non-randomly distributed in the most rearranged karyotypes. A comparison between FISH and conventional banding patterns provided evidence that LINE-1 insertion sites and chromosome breakpoints were not strongly correlated, thus suggesting that LINE-1 amplification subsequently accompanied Taterillus chromosome evolution. Similar patterns are observed in some cases of genomic stresses (hybrid genomes, cancer and DNA-damaged cells) and usually associated with DNA hypomethylation. We propose that intensively repatterned genomes face transient stress phases during which some epigenetic features, such as DNA methylation, are relaxed, thus allowing TE amplification.