Identification and functional modelling of plausibly causative cis-regulatory variants in a highly-selected cohort with X-linked intellectual disability.

Identifying causative variants in cis-regulatory elements (CRE) in neurodevelopmental disorders has proven challenging. We have used in vivo functional analyses to categorize rigorously filtered CRE variants in a clinical cohort that is plausibly enriched for causative CRE mutations: 48 unrelated males with a family history consistent with X-linked intellectual disability (XLID) in whom no detectable cause could be identified in the coding regions of the X chromosome (chrX). Targeted sequencing of all chrX CRE identified six rare variants in five affected individuals that altered conserved bases in CRE targeting known XLID genes and segregated appropriately in families. Two of these variants, FMR1CRE and TENM1CRE, showed consistent site- and stage-specific differences of enhancer function in the developing zebrafish brain using dual-color fluorescent reporter assay. Mouse models were created for both variants. In male mice Fmr1CRE induced alterations in neurodevelopmental Fmr1 expression, olfactory behavior and neurophysiological indicators of FMRP function. The absence of another likely causative variant on whole genome sequencing further supported FMR1CRE as the likely basis of the XLID in this family. Tenm1CRE mice showed no phenotypic anomalies. Following the release of gnomAD 2.1, reanalysis showed that TENM1CRE exceeded the maximum plausible population frequency of a XLID causative allele. Assigning causative status to any ultra-rare CRE variant remains problematic and requires disease-relevant in vivo functional data from multiple sources. The sequential and bespoke nature of such analyses renders them time-consuming and challenging to scale for routine clinical use

In-Depth Annotation of the Drosophila Bithorax-Complex Reveals the Presence of Several Alternative ORFs That Could Encode for Motif-Rich Peptides

It is recognized that a large proportion of eukaryotic RNAs and proteins is not produced from conventional genes but from short and alternative (alt) open reading frames (ORFs) that are not captured by gene prediction programs. Here we present an in silico prediction of altORFs by applying several selecting filters based on evolutionary conservation and annotations of previously characterized altORF peptides. Our work was performed in the Bithorax-complex (BX-C), which was one of the first genomic regions described to contain long non-coding RNAs in Drosophila. We showed that several altORFs could be predicted from coding and non-coding sequences of BX-C. In addition, the selected altORFs encode for proteins that contain several interesting molecular features, such as the presence of transmembrane helices or a general propensity to be rich in short interaction motifs. Of particular interest, one altORF encodes for a protein that contains a peptide sequence found in specific isoforms of two Drosophila Hox proteins. Our work thus suggests that several altORF proteins could be produced from a particular genomic region known for its critical role during Drosophila embryonic development. The molecular signatures of these altORF proteins further suggests that several of them could make numerous protein–protein interactions and be of functional importance in vivo

Endogenous retroviruses in fish genomes: from relics of past infections to evolutionary innovations?

The increasing availability of fish genome sequences has allowed to gain new insights into the diversity and host distribution of retroviruses in fish and other vertebrates. This distribution can be assessed through the identification and analysis of endogenous retroviruses, which are proviral remnants of past infections integrated in genomes. Retroviral sequences are probably important for evolution through their ability to induce rearrangements and to contribute regulatory and coding sequences; they may also protect their host against new infections. We argue that the current mass of genome sequences will soon strongly improve our understanding of retrovirus diversity and evolution in aquatic animals, with the identification of new/re-emerging elements and host resistance genes that restrict their infectivity

Sex and the TEs: transposable elements in sexual development and function in animals

Transposable elements are endogenous DNA sequences able to integrate into and multiply within genomes. They constitute a major source of genetic innovations, as they can not only rearrange genomes but also spread ready-to-use regulatory sequences able to modify host gene expression, and even can give birth to new host genes. As their evolutionary success depends on their vertical transmission, transposable elements are intrinsically linked to reproduction. In organisms with sexual reproduction, this implies that transposable elements have to manifest their transpositional activity in germ cells or their progenitors. The control of sexual development and function can be very versatile, and several studies have demonstrated the implication of transposable elements in the evolution of sex. In this review, we report the functional and evolutionary relationships between transposable elements and sexual reproduction in animals. In particular, we highlight how transposable elements can influence expression of sexual development genes, and how, reciprocally, they are tightly controlled in gonads. We also review how transposable elements contribute to the organization, expression and evolution of sexual development genes and sex chromosomes. This underscores the intricate co-evolution between host functions and transposable elements, which regularly shift from a parasitic to a domesticated status useful to the host

Interspecies Insertion Polymorphism Analysis Reveals Recent Activity of Transposable Elements in Extant Coelacanths

<div><p>Coelacanths are lobe-finned fish represented by two extant species, <i>Latimeria chalumnae</i> in South Africa and Comoros and <i>L. menadoensis</i> in Indonesia. Due to their intermediate phylogenetic position between ray-finned fish and tetrapods in the vertebrate lineage, they are of great interest from an evolutionary point of view. In addition, extant specimens look similar to 300 million-year-old fossils; because of their apparent slowly evolving morphology, coelacanths have been often described as « living fossils ». As an underlying cause of such a morphological stasis, several authors have proposed a slow evolution of the coelacanth genome. Accordingly, sequencing of the <i>L. chalumnae</i> genome has revealed a globally low substitution rate for protein-coding regions compared to other vertebrates. However, genome and gene evolution can also be influenced by transposable elements, which form a major and dynamic part of vertebrate genomes through their ability to move, duplicate and recombine. In this work, we have searched for evidence of transposition activity in coelacanth genomes through the comparative analysis of orthologous genomic regions from both <i>Latimeria</i> species. Comparison of 5.7 Mb (0.2%) of the <i>L. chalumnae</i> genome with orthologous Bacterial Artificial Chromosome clones from <i>L. menadoensis</i> allowed the identification of 27 species-specific transposable element insertions, with a strong relative contribution of CR1 non-LTR retrotransposons. Species-specific homologous recombination between the long terminal repeats of a new coelacanth endogenous retrovirus was also detected. Our analysis suggests that transposon activity is responsible for at least 0.6% of genome divergence between both <i>Latimeria</i> species. Taken together, this study demonstrates that coelacanth genomes are not evolutionary inert: they contain recently active transposable elements, which have significantly contributed to post-speciation genome divergence in <i>Latimeria</i>.</p></div

Example of a polymorphic insertion of a CR1 retrotransposon (element 7 in Table 2) present in <i>Latimeria menadoensis</i> but absent from <i>L. chalumnae</i>.

<p>Target Site Duplications (TSDs) are framed in red. CR1  =  Chicken Repeat 1; ORF  =  Open Reading Frame; RT  =  Reverse Transcriptase; APE  =  Apurinic/Apyrimidic Endonuclease.</p

Phylogenetic relationship between coelacanth CoeERV1-1 and reptile retroviruses.

<p>Vertebrate retrovirus phylogeny was reconstructed on an alignment of RT (210 amino acids) using Maximum Likelihood with optimized parameters (best of NNI and SPR; optimized invariable sites <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0114382#pone.0114382-Guindon1" target="_blank">[37]</a>. Branch values represent supporting aLRT non-parametric statistics. The dashed line highlights the group of Epsilon viruses containing turtle, crocodile, coelacanth and lungfish sequences. Gypsy LTR retrotransposon sequences were used as an outgroup.</p

Single-pass classification of all noncoding sequences in a bacterial genome using phylogenetic profiles

Identification and characterization of functional elements in the noncoding regions of genomes is an elusive and time-consuming activity whose output does not keep up with the pace of genome sequencing. Hundreds of bacterial genomes lay unexploited in terms of noncoding sequence analysis, although they may conceal a wide diversity of novel RNA genes, riboswitches, or other regulatory elements. We describe a strategy that exploits the entirety of available bacterial genomes to classify all noncoding elements of a selected reference species in a single pass. This method clusters noncoding elements based on their profile of presence among species. Most noncoding RNAs (ncRNAs) display specific signatures that enable their grouping in distinct clusters, away from sequence conservation noise and other elements such as promoters. We submitted 24 ncRNA candidates from Staphylococcus aureus to experimental validation and confirmed the presence of seven novel small RNAs or riboswitches. Besides offering a powerful method for de novo ncRNA identification, the analysis of phylogenetic profiles opens a new path toward the identification of functional relationships between co-evolving coding and noncoding elements