Damage to nuclear DNA induced by Shiga toxin 1 and ricin in human endothelial cells

被引:112
作者
Brigotti, M
Alfieri, R
Sestili, P
Bonelli, M
Petronini, PG
Guidarelli, A
Barbieri, L
Stirpe, F
Sperti, S
机构
[1] Univ Bologna, Dipartimento Patol Sperimentale, I-40126 Bologna, Italy
[2] Univ Parma, Dipartimento Med Sperimentale, Sez Patol Mol & Immunol, I-43100 Parma, Italy
[3] Univ Urbino, Fac Sci Motorie, Catedra Farmacol, I-61029 Urbino, Italy
[4] Univ Urbino, Ist Farmacol & Farmacognosia, I-61029 Urbino, Italy
关键词
ribosome-inactivating proteins; alkaline-halo assay; AP sites; cytotoxins;
D O I
10.1096/fj.01-0521com
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Ribosome-inactivating proteins (RIPs) remove a specific adenine from 28S rRNA leading to inactivation of ribosomes and arrest of translation. Great interest as to a possible second physiological substrate for RIPs came from the observation that in vitro RIPs remove adenine from DNA. This paper addresses the problem of nuclear lesions induced by RIPs in human endothelial cells susceptible to the bacterial RIP Shiga toxin 1 and the plant RIP ricin. With both toxins, nuclear DNA damage as evaluated by two independent techniques (alkaline-halo assay and alkaline filter elution) appears early, concomitant with (ricin) or after (Shiga toxin 1) the inhibition of protein synthesis. At this time, the annexin V binding assay, caspase 3 activity, the formation of typical less than or equal to 50 Kb DNA fragments, and changes in morphology associated with apoptosis were negative. Furthermore, a block of translation comparable to that induced by RIPs, but obtained with cycloheximide, did not induce nuclear damage. Such damage is consistent with the enzymatic activity (removal of adenine) of RIPs acting in vitro on RNA-free chromatin and DNA. The results unequivocally indicate that RIPs can damage nuclear DNA in whole cells by means that are not secondary to ribosome inactivation or apoptosis.
引用
收藏
页码:365 / 372
页数:8
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