We have examined a possible role for protein poly(ADP-ribosylation) during in vivo DNA replication by studying the metabolism of poly(ADP-ribose) in the nuclear matrix fraction from normal and regenerating rat liver. This fraction contains the newly replicated DNA and thus allows for the examination of the events closely associated with the replication process. It was found that 55% of the total nuclear protein-bound poly(ADP-ribose) and 15-35% of the total nuclear poly(ADP-ribose)-polymerase activity were tightly associated with this subnuclear compartment in normal liver. Surgical removal of two-thirds of the liver initiated a time-dependent decrease in nuclear matrix associated polymers of ADP-ribose and poly(ADP-ribose) polymerase activity which reached a minimum of 40% of control livers after 24 h, before returning to normal levels at 41 h post-partial hepatectomy. In contrast, the total levels of poly(ADP-ribose) in intact liver and the total polymerase activity of isolated nuclei exhibited a 2-fold increase over basal levels. These results are consistent with the conclusion that the nuclear matrix is a major poly(ADP-ribosylation) site within the nucleus and that this metabolic reaction may be closely connected with the events modulating DNA replication in this fraction.