Genomic structural variants have long been implicated in phenotypic diversity and human disease, but dissecting the mechanisms by which they exert their functional impact has proven elusive. Recently however, developments in high-throughput DNA sequencing and chromosomal engineering technology have facilitated the analysis of structural variants in human populations and model systems in unprecedented detail. In this Review, we describe how structural variants can affect molecular and cellular processes, leading to complex organismal phenotypes, including human disease. We further present advances in delineating disease-causing elements that are affected by structural variants, and we discuss future directions for research on the functional consequences of structural variants.

Phenotypic impact of genomic structural variation: insights from and for human disease

A Regulatory Archipelago Controls Hox Genes Transcription in Digits

Hox genes are major determinants of the animal body plan, where they organize structures along both the trunk and appendicular axes. During mouse limb development, Hoxd genes are transcribed in two waves: early on, when the arm and forearm are specified, and later, when digits form. The transition between early and late regulations involves a functional switch between two opposite topological domains. This switch is reflected by a subset of Hoxd genes mapping centrally into the cluster, which initially interact with the telomeric domain and subsequently swing toward the centromeric domain, where they establish new contacts. This transition between independent regulatory landscapes illustrates both the modularity of the limbs and the distinct evolutionary histories of its various pieces. It also allows the formation of an intermediate area of low HOX proteins content, which develops into the wrist, the transition between our arms and our hands. This regulatory strategy accounts for collinear Hox gene regulation in land vertebrate appendages.

https://science.sciencemag.org/content/sci/340/6137/1234167.full.pdf

A Switch Between Topological Domains Underlies HoxD Genes Collinearity in Mouse Limbs

http://materiais.dbio.uevora.pt/BD/Crescimento/ERVs.pdf

Dynamic control of endogenous retroviruses during development

I will argue that transcription 'factories' are central organizers of the human genome during interphase, and that proximity to an appropriate factory determines the activity of a gene. The nucleolus is the prototypic factory; it is a place where many rRNA genes are efficiently co-transcribed by local concentrations of RNA polymerase I. Analogous clusters of RNA polymerase II in nucleoplasmic factories make protein-coding transcripts. I begin by describing new forces able to drive genome organization uncovered using Brownian dynamic simulations. For example, a diffusion-based \" osmotic ratchet \" can force bound \" slip-links \" like cohesin to \" convergent \" CTCF sites without the need for any motor activity. Additional simulations point to an unforeseen 'bridging-induced attraction' that can assemble factories and organize interphase chromosomes at all scales. This organization leads naturally to an explanation of how gene activity is regulated: a promoter is only likely to initiate if tethered near a factory containing appropriate factors. As motifs like enhancers, silencers, insulators, barriers, and boundaries are transcription units, they would work by tethering target promoters close to, or distant from, suitable factories; although we might name the motifs differently, they are all just transcription units influencing promoter-factory distance (and so initiation frequency).

/pdf/transcription-factories-genome-organization-and-gene-5a0y4fezhn.pdf

Transcription factories: genome organization and gene regulation.

Finding sequences that control expression of genes is central to understanding genome function. Previous studies have used evolutionary conservation as an indicator of regulatory potential. Here, we present a method for the unbiased in vivo screen of putative enhancers in large DNA regions, using the mouse as a model. We cloned a library of 142 overlapping fragments from a 200 kb-long murine BAC in a lentiviral vector expressing LacZ from a minimal promoter, and used the resulting vectors to infect fertilized murine oocytes. LacZ staining of E11 embryos obtained by first using the vectors in pools and then testing individual candidates led to the identification of 3 enhancers, only one of which shows significant evolutionary conservation. In situ hybridization and 3C/4C experiments suggest that this enhancer, which is active in the neural tube and posterior diencephalon, influences the expression of the Olig1 and/or Olig2 genes. This work provides a new approach for the large-scale in vivo screening of transcriptional regulatory sequences, and further demonstrates that evolutionary conservation alone seems too limiting a criterion for the identification of enhancers.

/pdf/a-systematic-enhancer-screen-using-lentivector-transgenesis-d6ipto25d6.pdf

A systematic enhancer screen using lentivector transgenesis identifies conserved and non-conserved functional elements at the olig1 and olig2 locus

SHFM3 is a limb malformation characterized by the absence of central digits. It has been shown that this condition is associated with tandem duplications of about 500 kb at 10q24. The Dactylaplasia mice display equivalent limb defects and the two corresponding alleles (Dac1j and Dac2j) map in the region syntenic with the duplications in SHFM3. Dac1j was shown to be associated with an insertion of an unspecified ETn-like mouse endogenous transposon upstream of the Fbxw4 gene. Dac2j was also thought to be an insertion or a small inversion in intron 5 of Fbxw4, but the breakpoints and the exact molecular lesion have not yet been characterized. Here we report precise mapping and characterization of these alleles. We failed to identify any copy number differences within the SHFM3 orthologous genomic locus between Dac mutant and wild-type littermates, showing that the Dactylaplasia alleles are not associated with duplications of the region, in contrast with the described human SHFM3 cases. We further show that both Dac1j and Dac2j are caused by insertions of MusD retroelements that share 98% sequence identity. The differences between the nature of the human and mouse genomic abnormalities argue against models proposed so far that either envisioned SHFM3 as a local trisomy or Dac as a mutant allele of Fbxw4. Instead, both genetic conditions might lead to complex alterations of gene regulation mechanisms that would impair limb morphogenesis. Interestingly, the Dac2j mutation occurs within a highly conserved element that may represent a regulatory sequence for a neighboring gene.

/pdf/characterization-of-mouse-dactylaplasia-mutations-a-model-32vpqt1j7i.pdf

Characterization of mouse Dactylaplasia mutations: a model for human ectrodactyly SHFM3

Comparative analyses of various mammalian genomes have identified numerous conserved non-coding (CNC) DNA elements that display striking conservation among species, suggesting that they have maintained specific functions throughout evolution. CNC function remains poorly understood, although recent studies have identified a role in gene regulation. We hypothesized that the identification of genomic loci that interact physically with CNCs would provide information on their functions. We have used circular chromosome conformation capture (4C) to characterize interactions of 10 CNCs from human chromosome 21 in K562 cells. The data provide evidence that CNCs are capable of interacting with loci that are enriched for CNCs. The number of trans interactions varies among CNCs; some show interactions with many loci, while others interact with few. Some of the tested CNCs are capable of driving the expression of a reporter gene in the mouse embryo, and associate with the oligodendrocyte genes OLIG1 and OLIG2. Our results underscore the power of chromosome conformation capture for the identification of targets of functional DNA elements and raise the possibility that CNCs exert their functions by physical association with defined genomic regions enriched in CNCs. These CNC-CNC interactions may in part explain their stringent conservation as a group of regulatory sequences.

/pdf/chromosome-conformation-capture-uncovers-potential-genome-4lki0hf7a9.pdf

Mélanie Arcangeli

Papers

A systematic enhancer screen using lentivector transgenesis identifies conserved and non-conserved functional elements at the olig1 and olig2 locus

Characterization of mouse Dactylaplasia mutations: a model for human ectrodactyly SHFM3

Chromosome Conformation Capture Uncovers Potential Genome-Wide Interactions between Human Conserved Non-Coding Sequences