Far beyond variant discovery: NGS applied to the study of spatial organization of the mouse genome

When it comes to NGS, we usually think about genetic variant discovery in individuals with medical conditions.

However the potentials of the new sequencing technologies can go much further as demonstrated by this masterpiece published on Cell. In this paper by Zhang et al. authors used high-throughput genome-wide translocation sequencing (HTGTS) together with Genome-wide chromosome conformation capture (Hi-C) technique to obtain a detailed map of chromosome physical interactions in the mouse genome and evaluate the impact of chromosome spatial distribution on translocation events.

Reading the Methods section we find where the NGS provide its support. First, authors used Roche 454 for the HTGTS application. Then they applied NGS to Hi-C, with a really interesting method. Briefly, after cross-linking, they have digested the mouse genomic DNA with the HindIII restriction enzyme, labeled the fragment obtained with a biotinylated dCTP and then ligated adjacent fragments. These steps resulted in short sequences composed by two pieces of DNA that were in spatial proximity with the restriction sites and the labeled dCTP in the middle. Then they reversed cross-ligation and proceeded with the standard protocol for NGS library preparation: fragmentation by sonication and hang-repair. They added a bead-capture step to select only fragments containing the biotinylated dCTP, and then finalized the library by size-selection and adapter ligation. Using Illumina NGS paired-end sequencing they were able to massively sequence both extremities of these fragments and then map them against all the HindIII restriction sites in the mouse genome obtaining a precise map of all the proximity regions occurring between different chromosomes!

The results obtained are really interesting! They demonstrated that spatial distribution, together with the double strand breaks (DSB) events probability, is a major factor in determining preferential translocation. Besides providing an accurate map of spatial distribution of chromosomes in mammals, as authors stated in the discussion "this finding has great relevance to translocations in cancer". The paper shows that "formation of translocations between randomly generated DSBs, such as those induced by chemotherapies and radiotherapies, will likely reflect a strong influence of spatial proximity [...]. [The results] also suggest that spatial proximity may be a major driving force for the activation of certain oncogenes via translocation to a wide range of recurrent partners".