Use of enzymatic assay to evaluate uv-induced DNA repair in human and embryonic chick fibroblasts and multinucleate heterokaryons derived from both

Conference · Wed Jan 01 04:00:00 EST 1975

OSTI ID:4120207

Paterson, M C; Lohman, P H.M.; Hanawalt, P C ^[1]

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1. ed.

A sensitive enzymatic assay has been utilized to monitor repair of uv- induced damage to DNA in primary human and embryonic chick cells in multinucleate heterokaryons artificially derived from both. The assay exploits the unique ability of a purified repair endonuclease to attack uv-irradiated DNA at sites containing pyrimidine dimers. These nuclease-susceptible sites are subsequently observed as single-strand scissions by velocity sedimentation in alkaline sucrose gradients. Incubation of uv-damaged cultures followed by extraction and enzymatic analysis of the radioactively labeled DNA enables one to trace the disappearance of such sites in vivo and hence to monitor endogenous repair activity. When uv-irradiated human cells are incubated in the dark, the curve for site removal exhibits a two-phase exponential decline; i.e., there exists a fast component responsible for elimination of 60 percent of the initial damage and a second one approximately 7 times slower in rate. The removal of sites is not further enhanced by exposing cells to blacklight during post-uv incubation. Conversely, uv-damaged chick cells rid their DNA of all nuclease-susceptible sites rapidly (i.e., at an exponential rate approximately 13 times faster than the fast component of site removal in human cells) when incubated under blacklight but not when kept in the dark. These data indicate the presence in human and embryonic chick cells of distinct enzymatic mechanisms for the elimination of dimer-containing sites. Whereas human fibroblasts rely heavily on a light-independent process, excision-repair, chick fibroblasts possess a light- dependent mechanism, presumably photoenzymatic repair. (auth)

• Research Organization: Atomic Energy of Canada, Ltd., Chalk River, Ont.
• NSA Number: NSA-33-012243
• OSTI ID: 4120207
• Country of Publication: United States
• Language: English

Related Subjects

*DNA-- BIOLOGICAL REPAIR
*FIBROBLASTS-- BIOLOGICAL RADIATION EFFECTS
*NUCLEASES-- PHOTOCHEMISTRY
560121* --Biomedical Sciences
Applied Studies--Radiation Effects--Radiation Effects on Cells--External Source
BIOASSAY
CHICKENS
DIMERS
MAN
N48210* --Life Sciences--Radiation Effects on Cells-- External Source
ULTRAVIOLET RADIATION