Replication Initiation Patterns in the β-Globin Loci of Totipotent and Differentiated Murine Cells: Evidence for Multiple Initiation Regions

Abstract

The replication initiation pattern of the murine β-globin locus was analyzed in totipotent embryonic stem cells and in differentiated cell lines. Initiation events in the murine β-globin locus were detected in a region extending from the embryonic Ey gene to the adult βminor gene, unlike the restricted initiation observed in the human locus. Totipotent and differentiated cells exhibited similar initiation patterns. Deletion of the region between the adult globin genes did not prevent initiation in the remainder of the locus, suggesting that the potential to initiate DNA replication was not contained exclusively within the primary sequence of the deleted region. In addition, a deletion encompassing the six identified 5′ hypersensitive sites in the mouse locus control region had no effect on initiation from within the locus. As this deletion also did not affect the chromatin structure of the locus, we propose that the sequences determining both chromatin structure and replication initiation lie outside the hypersensitive sites removed by the deletion.

We gratefully acknowledge D. J. Ciavatta and T. M. Townes for the ES cells containing the adult β-globin deletion, C. Schildkraut and N. Lam for sharing data prior to publication and helpful comments, and Kurt Kohn, Yves Pommier, and Fred Indig for insightful discussions.

This work was supported by grants from the NIH to G.M.W. (CA48405 and GM51104) and to M.G. (DK44746), a grant from the G. Harold and Leila Y. Mathers Charitable Foundation to G.M.W., and a fellowship from the American Cancer Society to D.M.C. M.I.A. was a Special Fellow of the Leukemia Society of America.