DNA loop domains in a 1.4-Mb region around the human hprt gene mapped by cleavage mediated by nuclear matrix-associated topoisomerase II

Abstract

We have mapped the positions in a approximately 1.4-Mb region of genomic DNA around the human hprt gene which are accessible in vivo to cleavage by topoisomerase II associated with the nuclear matrix. These positions, which are interpreted as the boundaries of DNA loop domains, were mapped in K562 cells by examining the truncation of rare-cutter restriction fragments separated by pulsed field gel electrophoresis after topoisomerase II-mediated cleavage, using seven linked markers mapped in this region as probes for indirect end-labeling. Eleven cleavage positions were detected and were interpreted as defining ten loop domains of lengths between 70 and 210 kb (average approximately 135 kb); the hprt gene resides in a 150-kb loop domain. Loop domain boundaries coincided with three of the fifteen deletion breakpoints mapped in a 600-kb sector of this region in human lymphocytes, within the limits of resolution of pulsed field gel electrophoresis; this correlation was not statistically significant.

Visualization of individual DNA loops and a map of loop domains in the human dystrophin gene

“…We believe that this apparent length re¯ects the existence of long regions of DNA attachment to the nuclear matrix and inability to obtain complete cleavage by the enzyme, because the length distribution of these fragments becomes more homogeneous as the concentration of VM-26 increases. Similar incomplete cleavage has been observed in other regions (23,25). Alternatively, the cell population could be heterogeneous with different attachment sites being selected from a region of potential attachment in different cells and/or at different times in the cell cycle.…”

Beziehungen zwischen der Chromatinstruktur und Apoptose-bedingter DNA-Fragmentierung

“…Durch die Anheftung bilden sich sogenannte DNA-Schleifen aus, die als Domänen einer gemeinsamen Regulation unterliegen könnten (LAEMMLI et al, 1992;RAZIN et al, 1993) RAZIN et al, 1993;GROMOVA et al, 1995a;IAROVAIA, O. et al, 1996;FAJKUS et al, 1998). Sie wird auch als "Topoisomerase II mediated DNA loop excision protocol" (Methode zur Topoisomerase II-vermittelten Isolierung von DNA-Schleifen) bezeichnet.…”