Association of human origin recognition complex 1 with chromatin DNA and nuclease-resistant nuclear structures

An origin recognition complex (ORC) consisting of six polypeptides has been identified as a DNA replication origin-binding factor in Saccharomyces cer evisiae, Homologues of ORC subunits have been discovered among eukaryotes, and we have prepared monoclonal antibodies against a human homologue of ORC 1 (hORC1) to study its localization in human cells. It was thus found to as sociate with nuclei throughout the cell cycle and to be resistant to nonion ic detergent treatment, in contrast to MCM proteins, which are other replic ation factors, the association of which with nuclei is clearly dependent on the phase of the cell cycle, A characteristic feature of hORC1 is dissocia tion by NaCl in a narrow concentration range around 0.25 hi, suggesting int eraction with some specific partner(s) in nuclei. Nuclease treatment experi ments and UV cross-linking experiments further indicated interaction with b oth nuclease-resistant nuclear structures and chromatin DNA. Although its D NA binding was unaffected, some variation in the cell cycle was apparent, t he association with nuclear structures being less stable in the M phase. In terestingly, the less stable association occurred concomitantly with hyperp hosphorylation of hORC1, suggesting that this hyperphosphorylation may be i nvolved in M phase changes.