Steps in DNA Chain Elongation and Joining after Ultra-violet Irradiation of Human Cells

Summary

The DNA synthesized in human cells shortly after u.v.-irradiation is of lower molecular weight than that in unirradiated cells. Within several hours after irradiation, these smaller DNA units are both elongated and joined together. To determine if this process involves incorporation of exogenous precursors, cells were irradiated, pulse-labelled, and incubated in medium containing bromodeoxyuridine. They were then exposed to different fluxes of 313-nm radiation, and the number of breaks in the DNA was determined by sedimentation in alkaline sucrose. If bromodeoxyuridine, rather than exclusively premade DNA, is used to elongate and join the small DNA segments synthesized after u.v.-irradiation, photolysis by 313-nm radiation would yield the lower molecular weight segments that are present immediately after u.v.-irradiation. This expectation was fulfilled, and our results indicate that the mechanism of filling the gaps in DNA synthesized after irradiation involves the insertion of ∼ 10³ nucleotides per gap in both normal human cells and those from individuals with xeroderma pigmentosum.