Dynamic association of replicating DNA fragments with the nuclear matrix of regenerating liver

Approx. 1-2% of the total nuclear DNA remains anchored to the proteinaceous nucleoskeletal matrix following endogenous DNA cleavage in isolated regenerating rat liver nuclei. The matrix-attached DNA fragments have an average size of 1.6 kilobases compared with 0.8 kilobases for total nuclear DNA, and are initially enriched with in vivo pulse-labeled, replicating DNA. Use of protease inhibitors during nuclear matrix isolation results in a several-fold enhancement of matrix-attached, newly replicating DNA. The percentage of replicating DNA which is anchored to the matrix rapidly declines from approx. 50% (one min pulse) to 5% after a 10 min pulse. The replicating DNA fragments are remarkably sensitive to release from the matrix structure by exogenous nucleases including pancreatic DNase I and micrococcal nuclease. Moreover, a preferential release of replicating as opposed to non-replicating DNA fragments is measured as long as substantial amounts of replicating DNA are matrix-attached. These findings suggest that the DNA replicational sites are continuously bound to the nuclear matrix. Two alternative mechanistic views termed the fixed and sliding matrix models are proposed to describe this association. In both models it is proposed that the DNA of the rat liver nucleus consists of ~ 125 000 matrix-attached DNA loops with an average size of 80 kilobases. The replication of DNA in replicon subunits then follows as a natural consequence of this inherent spatial organization.