The DNA molecule is the molecular basis of heredity. Each species has its own specific DNA, and within a species each individual carries its own characteristic DNA. Nonetheless, it is far from being well understood how DNA encodes structures as sophisticated as those of complex multicellular organisms. DNA carries the genes, but the organism complexity does not only depend on genes. For example, in humans, genes are encoded by roughly 1% of the DNA. The 99% of DNA that are left are necessarily involved in the determination of the structure of our body, but the underlying molecular mechanisms still need to be discovered.

One commonly admitted hypothesis is that the organization and folding of DNA in chromosomes depend on the cell type, leading to a cell type specific expression of genes. This is why the study of DNA topology and of the DNA spatial arrangement in chromosomes is of great interest. In this context, our research focuses more specifically on original structures called DNA hemicatenanes (Figure 1), in which two distinct DNA molecules are stably associated, allowing the formation of DNA loops (Figure 2). Could DNA hemicatenanes participate in the structuration of the DNA loops of chromosomes?

Figure 1. DNA hemicatenane. Two distinct double strand DNA molecules are stably associated, with one of the strands of one DNA molecule going in between the two strands of the other DNA molecule, and reciprocally.

Figure 2. DNA loops prepared in vitro and observed by atomic force microscopy (Lyubchenko et al. 2002). These loops are stable, as they are maintained by a DNA hemicatenane at their bases, as represented schematically on the right.

We have recently developed a method to assemble DNA hemicatenanes from two small circular DNA molecules. Our aim is now twofold: 

  • On the one hand, we want to identify nuclear proteins that specifically bind DNA hemicatenanes. Our strategy consists in fractionating protein nuclear extracts on a variety of chromatographic supports and in identifying, for each chromatography, fractions that contain proteins able to specifically bind our structures. Once purified, the proteins of interest will be further characterized both in vivo and in vitro. 
  • On the other hand, we want to improve our method of construction of DNA hemicatenanes in order to be able to assemble such structures from large circular DNA molecules. We will thus be able to introduce such constructs into culture cells, characterize their cellular fate and functions, and shed new light on their molecular properties.