Determination of the in vivo structural DNA loop organization in the genomic region of the rat albumin locus by means of a topological approach

Abstract

Nuclear DNA of metazoans is organized in supercoiled loops anchored to a proteinaceous substructure known as the nuclear matrix (NM). DNA is anchored to the NM by non-coding sequences known as matrix attachment regions (MARs). There are no consensus sequences for identification of MARs and not all potential MARs are actually bound to the NM constituting loop attachment regions (LARs). Fundamental processes of nuclear physiology occur at macromolecular complexes organized on the NM; thus, the topological organization of DNA loops must be important. Here, we describe a general method for determining the structural DNA loop organization in any large genomic region with a known sequence. The method exploits the topological properties of loop DNA attached to the NM and elementary topological principles such as that points in a deformable string (DNA) can be positionally mapped relative to a position-reference invariant (NM), and from such mapping, the configuration of the string in third dimension can be deduced. Therefore, it is possible to determine the specific DNA loop configuration without previous characterization of the LARs involved. We determined in hepatocytes and B-lymphocytes of the rat the DNA loop organization of a genomic region that contains four members of the albumin gene family.

DNA moves sequentially towards the nuclear matrix during DNA replication in vivo

“…A typical structural DNA loop can be divided into four topological zones according to their relative proximity to the NM [23]. Each of these zones would manifest an identifiable behavior when exposed to non-specific nucleases that are sensitive to the local DNA topology such as DNase I (Figure 1B).…”

The post-mitotic state in neurons correlates with a stable nuclear higher-order structure

“…In situ MARs have been operationally classified into structural-constitutive, resistant to high-salt extraction, and functional-facultative, non-resistant to high-salt extraction 31 , 32 . Therefore the resulting DNA loops can be also classified into structural and functional 30 , 33 , 34 . The high-salt resistant MARs attaching the structural DNA loops to the NM are also known as loop anchorage regions or LARs 31 .…”

The organization of a large transcriptional unit (Fyn) into structural DNA loops is cell-type specific and independent of transcription

“…In nucleoid preparations the relative resistance of a given loop-DNA sequence to a limited concentration of DNase I, a non-specific nuclease that is sensitive to the local DNA topology (Lewin, 1980), is directly proportional to its proximity to the NM anchoring point Maya-Mendoza et al, 2004;Rivera-Mulia and Aranda-Anzaldo, 2010). Two main factors determine such property: (1) Steric hindrance resulting from the proteinaceous NM that acts as a physical barrier that relatively protects the naked loop DNA that is closer to the NM from endonuclease action.…”