MODULATION OF A CARDIOGENIC-SHOCK INDUCIBLE RNA BY CHEMICAL STRESS - ADAPT73 PIGHEP3/

Background. Understanding how mammalian cells respond to sh-ess is imp ortant in the study, detection, and therapy of stress-related disorder s. We have been studying cellular stress response in hamster HA-1 cell s by using an adaptive response model. HA-1 cells respond to a minimal ly toxic ''pretreatment'' dose of hydrogen peroxide by synthesizing RN As and proteins that protect them against subsequent exposure to a hig her cytotoxic concentration of peroxide. The purpose of our studies is to identify and partially characterize any mRNA whose steady state le vel is significantly modulated during adaptation. Methods, HA-I cells were exposed to a pretreatment dose of hydrogen peroxide and RNA extra cted. The differential display technique was used to identify modulate d mRNAs. The effects of calcium ionophore A23187 and c's (II)-platinum on the modulation of mRNA from HA-I cells and A23187 on the modulatio n of mRNA from human IMR-90 cells were also determined. Results. One o f the RNAs induced by a pretreatment concentration of hydrogen peroxid e was designated adapt73. The size of the induced adapt73 RNA was dete rmined to be 2.1 kb. Induction of adapt73 was maximal 5 hours after pe roxide treatment, but elevated Levels were still obvious at 10 hours. This induction was not specific to oxidative stress, because other str ess agents including cis (II)-platinum and especially calcium ionophor e. A23187 also induced adapt73 mRNA levels. Partial sequencing of adap t73 and a subsequent GenBank homology search revealed extensive homolo gy to a novel RNA from Pig; designated PigHep3, that was identified as a cardiogenic shock response gene fi-om liver in pigs that were under going resuscitation after circulatory shock. Homology to a completely sequenced but uncharacterized human homolog was found. Using a partial ly sequenced expressed sequence tag (EST) human clone to Adapt73, we p robed Northern blots containing RNA from IMR-90 human fibroblasts trea ted with A23187. A strongly induced human adapt 73 mRNA homolog was ob served, almost identical in size to ifs hamster homolog. In vitro tran scription and translation of the human EST clone revealed a translatab le Adapt73 protein product. Conclusions, These data indicate that adap t73/PigHep3 RNA can be induced by multiple chemical stress, that these inductions occur under protective or adaptive response conditions, th at there is an inducible human homolog to adapt73 and suggest that ada pt73 may be an important physiologic mediator of organ and cellular sh ock response in mammals.