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Yoshihito Oura, Motokazu Tsujikawa, Kohji Nishida; Overexpression of OASIS/CREB3L1 in photoreceptor of zebrafish Induces Photoreceptor Cell Death without Classical Endoplasmic Reticulum Stress Response. Invest. Ophthalmol. Vis. Sci. 2013;54(15):1798.
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© ARVO (1962-2015); The Authors (2016-present)
Old astrocyte specifically induced substance (OASIS) is a putative endoplasmic reticulum (ER) stress sensor in astrocytes with a mechanism of activation that is similar to ATF6. In this study we investigated the effect of overexpression of OASIS in photoreceptor cells death induced by ER stress.
We generated a transgenic zebrafish line of OASIS using the tol2 transposon system by which we can express any exogenous genes in fish rod photoreceptors. We measured the cell viability by two methods, counting photoreceptor cells (PRC) numbers and TUNEL assay. ER stress was induced by 1ug/ml of tunicamycin. We investigated the expression levels of unfolded protein response (UPR) genes in OASIS transgenic and wild type (wt) fish. From 3 to 7 days after fertilization (dpf), mRNA expression levels were tested by RT-PCR.
OASIS mRNA expression was detected in eye of 7 dpf embryo in both transgenic embryo and wt. Unexpectedly, photoreceptor cell viability of OASIS transgenic embryo was decreased significantly (mean PRC number±SE; OASIS+:27.7±2.9, Wt:81.9±3.9, p<0.05 student t-test). TUNEL assay positive cells were seen in outer nuclear layer of OASIS transgenic embryos but not seen in that of wt. When treated with tunicamycin, numbers of PRC were significantly decreased in wt, whereas there is no significant change in OASIS transgenic embryo (mean PRC number±SE; OASIS+:20.9±3.9, Wt:40.1±3.5). In mRNA levels of transgenic embryo was increased in 19% compare with wt, and other UPR genes were also increased except for XBP1 (relative quantity normalized by β-actin(RQ): OASIS 1.19; Bip 1.12; CHOP 1.42; XBP1 0.71; sXBP1 0.99; ATF4 1.38; ATF6 1.31). Treated with tunicamycin, mRNA levels of UPR genes were significantly increased in both wt and OASIS transgenic (RQ of OASIS transgenic/wt, Bip 7.4/6.6, CHOP 21.3/23.2, XBP1 1.86/2.40, sXBP1 6.15/8.02, ATF4 1.31/1.18, ATF6 2.23/2.48, respectively). There was no significant difference between OASIS transgenic and wt.
Overexpression of OASIS in PRC induced apoptosis of PRC. It is not by classical endoplasmic reticulum stress response but by other signaling pathway, because the expression level of UPR genes were not affected.
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