RESUMO
Chromosome conformation capture techniques have revolutionized our understanding of chromatin architecture and dynamics at the genome-wide scale. In recent years, these methods have been applied to a diverse array of species, revealing fundamental principles of chromosomal organization. However, structural organization of the extrachromosomal entities, like viral genomes or plasmids, and their interactions with the host genome, remain relatively underexplored. In this work, we introduce an enhanced 4C-protocol tailored for probing plasmid DNA interactions. We design specific plasmid vector and optimize protocol to allow high detection rate of contacts between the plasmid and host DNA.
Assuntos
Plasmídeos , Plasmídeos/metabolismo , Plasmídeos/genética , DNA/química , DNA/genética , Cromatina/genética , Cromatina/metabolismo , Cromatina/química , GenomaRESUMO
Structural variations are a pervasive feature of human genomes, and there is growing recognition of their role in disease development through their impact on spatial chromatin architecture. This understanding has led us to investigate the clinical significance of CNVs in noncoding regions that influence TAD structures. In this study, we focused on the Epb41l4a locus, which contains a highly conserved TAD boundary present in both human chromosome 5 and mouse chromosome 18, and its association with neurodevelopmental phenotypes. Analysis of human data from the DECIPHER database indicates that CNVs within this locus, including both deletions and duplications, are often observed alongside neurological abnormalities, such as dyslexia and intellectual disability, although there is not enough evidence of a direct correlation or causative relationship. To investigate these possible associations, we generated mouse models with deletion and inversion mutations at this locus and carried out RNA-seq analysis to elucidate gene expression changes. We found that modifications in the Epb41l4a TAD boundary led to dysregulation of the Nrep gene, which plays a crucial role in nervous system development. These findings underscore the potential pathogenicity of these CNVs and highlight the crucial role of spatial genome architecture in gene expression regulation.