A requirement for RNA, protein and DNA synthesis in the establishment of DNA replicase activity in synchronized HeLa cells

Abstract

DNA replicase activity of synchronized cultures of HeLa cells was assayed by a permeable cell technique during normal S-phase and under conditions of restricted RNA, protein, or DNA synthesis. Inhibition studies with puromycin, cycloheximide, actinomycin D, and 2-mercapto-1-(beta-4-pyridethyl)benzimidazole revealed that the establishment as well as the maintenance of DNA replicase activity in S-phase cells was dependent on the continued synthesis of both RNA and protein. Measurements during limitation of DNA replication by hydroxyurea, cytosine arabinoside, or restricted availability of thymidine indicate that a low level of DNA synthesis is required to activate or assemble some subunits of DNA replicase. Evidence for the existence of short-lived RNA and protein factors essential for DNA replicase activity is discussed.