REQUIREMENT OF TYROSINE AND SERINE THREONINE KINASE IN THE TRANSCRIPTIONAL ACTIVATION OF THE MAMMALIAN GRP78/BIP PROMOTER BY THAPSIGARGIN/

Depletion of endoplasmic reticulum (ER) Ca2+ store by thapsigargin (Tg ) in mammalian cells induces a set of ER protein genes known as the gl ucose-regulated proteins. Recently, IRE1p, a transmembrane protein pos tulated to have a serine/threonine kinase activity, has been identifie d as required for the induction of ER resident proteins genes in Sacch aromyces cerevisiae. To investigate whether IRE1p can stimulate mammal ian grp transcription, a stable Chinese hamster ovary cell line contai ning amplified copies of IRE1p has been created. The IRE1p expressing transfectants exhibited a modest (2-fold) enhancement of both the basa l and Tg induced level of grp78 and grp94, two coordinately regulated grp genes. Using okadaic acid as a specific inhibitor for the endogeno us serine/threonine protein phosphatase activities, a mild (2-fold) st imulative effect was observed for Tg induction of grp78 transcription. The okadaic acid potentiating ef feet requires a 50-base pair region in the vicinity of the grp78 TATA element. In contrast, the transcript ional activation of grp78 by Tg is almost totally eliminated by genist ein, a tyrosine kinase inhibitor. The grp core, the C3 and C1 elements which are major Tg response elements of the rat grp78 promoter, are a lso major targets of the inhibitive effects of genistein.