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Calmodulin, poly(ADP–ribose)polymerase and p53 are targets for modulating the effects of sulfur mustard
Author(s) -
Rosenthal Dean S.,
SimbulanRosenthal Cynthia M.,
Iyer Sudha,
Smith William J.,
Ray Radharaman,
Smulson Mark E.
Publication year - 2000
Publication title -
journal of applied toxicology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.784
H-Index - 87
eISSN - 1099-1263
pISSN - 0260-437X
DOI - 10.1002/1099-1263(200012)20:1+<::aid-jat691>3.0.co;2-s
Subject(s) - poly adp ribose polymerase , involucrin , apoptosis , biology , microbiology and biotechnology , calmodulin , programmed cell death , keratinocyte , in vitro , polymerase , biochemistry , dna , enzyme
We describe two pathways by which the vesicating agent sulfur mustard (HD) may cause basal cell death and detachment: induction of terminal differentiation and apoptosis. Following treatment of normal human epidermal keratinocytes (NHEK) with 10 or 100 μmM HD, the differentiation‐specific keratin pair K1/K10 was induced and the cornified envelope precursor protein, involucrin, was cross‐linked by epidermal transglutaminase. Fibronectin levels were reduced in a time‐ and dose‐dependent manner. The rapid increase in p53 and decrease in Bcl‐2 levels was consistent not only with epidermal differentiation but with apoptosis as well. Further examination of biochemical markers of apoptosis following treatment of either NHEK or human papillomavirus (HPV)‐immortalized keratinocytes revealed a burst of poly(ADP–ribose) synthesis, specific cleavage of poly(ADP–ribose)polymerase (PARP) in vivo and in vitro into characteristic 89 and 24 kDa fragments, processing of caspase‐3 into its active form and the formation of DNA ladders. The intracellular calcium chelator BAPTA suppressed the differentiation markers, whereas antisense oligonucleotides and chemical inhibitors specific for calmodulin blocked both markers of differentiation and apoptosis. Modulation of p53 levels utilizing retroviral constructs expressing the E6, E7 or E6 + E7 genes of HPV‐16 revealed that HD‐induced apoptosis was partially p53‐dependent. Finally, immortalized fibroblasts derived from PARP −/− ‘knockout mice’ were exquisitely sensitive to HD‐induced apoptosis. These cells became HD resistant when wild‐type PARP was stably expressed in these cells. These results indicate that HD exerts its effects via calmodulin, p53 and PARP‐sensitive pathways. Copyright © 2000 John Wiley & Sons, Ltd.