Role of poly(ADP-ribose) polymerase (PARP) in DNA repair in sulfur mustard-exposed normal human epidermal keratinocytes (NHEK).
We previously reported that, in normal human epidermal keratinocytes (NHEK) cultures exposed to the alkylating compound sulfur mustard (bis-(2-chloroethyl) sulfide, HD, 0.3-1 mM), there is a rapid (< or =1 h) activation (100% above unexposed control) of the DNA repair enzyme DNA ligase I (130 kD) followed by a first-order decay (1-5 h). The DNA ligase activation is accompanied by a time-dependent (0.5-4 h) and significant DNA repair. Inhibition of another putative DNA repair enzyme, poly(ADP-ribose) polymerase (PARP), by using 3-amino benzamide does not affect DNA ligase activation following HD exposure, but increases the half-life of the activated enzyme threefold. To examine the role of PARP in HD-induced DNA ligase activation and subsequent DNA repair, we conducted studies using cultured keratinocytes in which the level of PARP had been selectively lowered (> or =85%) by the use of induced expression of antisense RNA. In these cells, there was no stimulation of DNA ligase up to 3 h, and a small stimulation (ca. 30% above unexposed control at 5-6 h after HD exposure. A time-course (0.5-6 h) study of DNA repair in HD-exposed PARP-deficient keratinocytes revealed a much slower rate of repair compared with HD-exposed NHEK. The results suggest an active role of PARP in DNA ligase activation and DNA repair in mammalian cells, and also indicate that modulation of PARP-mediated mechanisms may provide a useful approach in preventing HD toxicity.
Bhat
K R
KR
Lincoln University, PA, USA.
Benton
B J
BJ
Rosenthal
D S
DS
Smulson
M E
ME
Ray
R
R
D018929:Cell Culture Techniques; D000072481:DNA Ligase ATP; D011088:DNA Ligases / Q000096:biosynthesis / Q000378:metabolism; D004260:DNA Repair; D003879:Dermatologic Agents / Q000009:adverse effects*; D005786:Gene Expression Regulation; D006801:Humans; D015603:Keratinocytes / Q000187:drug effects / Q000473:pathology; D009151:Mustard Gas / Q000009:adverse effects*; D011065:Poly(ADP-ribose) Polymerases / Q000378:metabolism*
D003879:Dermatologic Agents; C000607892:LIG1 protein, human; D011065:Poly(ADP-ribose) Polymerases; D011088:DNA Ligases; D000072481:DNA Ligase ATP; D009151:Mustard Gas