THE PROMOTER REGION OF THE YEAST KAR2 (BIP) GENE CONTAINS A REGULATORY DOMAIN THAT RESPONDS TO THE PRESENCE OF UNFOLDED PROTEINS IN THE ENDOPLASMIC-RETICULUM
THE PROMOTER REGION OF THE YEAST KAR2 (BIP) GENE CONTAINS A REGULATORY DOMAIN THAT RESPONDS TO THE PRESENCE OF UNFOLDED PROTEINS IN THE ENDOPLASMIC-RETICULUM
复制标题
DOI：
10.1128/mcb.13.2.877
复制
发表时间：
1993-02-01
影响因子：
5.3
通讯作者：
MORI, K
中科院分区：
文献类型：
--
作者：
KOHNO, K;NORMINGTON, K;MORI, K
真核细胞的内质网(ER)含有丰富的78,000-Da蛋白质(BiP)，参与分泌蛋白和跨膜蛋白的转位、折叠和组装。在酿酒酵母中，与哺乳动物细胞一样，Bip mRNA以较高的基础速率合成，并因内质网中存在更多的未折叠蛋白质而进一步诱导。然而，与哺乳动物Bip不同的是，酵母Bip也会被热休克诱导数倍，尽管是短暂的。为了确定编码Bip的酵母KAR2基因中对这些刺激做出反应的调控序列，我们从Bip编码序列的上游区域克隆了一段1.3kb的DNA，并将其与报告基因--大肠杆菌β-半乳糖苷酶基因融合在一起。对一系列渐进的5‘截断以及上游序列的内部缺失的分析表明。准确调控酿酒酵母KAR2基因转录所需的信息包含在一个约230个碱基的XhoI-Drai片段(核苷酸-245至-9)中，该片段至少包含两个顺式作用元件，一个(热休克元件[HSE])响应热休克，另一个(未折叠蛋白响应元件[UPR])响应内质网中未折叠蛋白质的存在。HSE和UPR元件在功能上相互独立，但为了最大限度地诱导酵母KAR2基因的表达，它们是相加的。位于这两个元件之间的是一个GC富集区，它在序列上与哺乳动物转录因子Sp1的共同结合元件相似，并参与KAR2基因的基础表达。最后，我们提供的证据表明，酵母细胞监控内质网中游离Bip的浓度，并相应地调节KAR2基因的转录水平；这种作用是通过KAR2启动子中的UPR元件介导的。
The endoplasmic reticulum (ER) of eukaryotic cells contains an abundant 78,000-Da protein (BiP) that is involved in the translocation, folding, and assembly of secretory and transmembrane proteins. In the yeast Saccharomyces cerevisiae, as in mammalian cells, BiP mRNA is synthesized at a high basal rate and is further induced by the presence of increased amounts of unfolded proteins in the ER. However, unlike mammalian BiP, yeast BiP is also induced severalfold by heat shock, albeit in a transient fashion. To identify the regulatory sequences that respond to these stimuli in the yeast KAR2 gene that encodes BiP, we have cloned a 1.3-kb segment of DNA from the region upstream of the sequences coding for BiP and fused it to a reporter gene, the Escherichia coli beta-galactosidase gene. Analysis of a series of progressive 5' truncations as well as internal deletions of the upstream sequence showed that the. information required for accurate transcriptional regulation of the KAR2 gene in S. cerevisiae is contained within a approximately 230-bp XhoI-DraI fragment (nucleotides -245 to -9) and that this fragment contains at least two cis-acting elements, one (heat shock element [HSE]) responding to heat shock and the other (unfolded protein response element [UPR]) responding to the presence of unfolded proteins in the ER. The HSE and UPR elements are functionally independent of each other but work additively for maximum induction of the yeast KAR2 gene. Lying between these two elements is a GC-rich region that is similar in sequence to the consensus element for binding of the mammalian transcription factor Sp1 and that is involved in the basal expression of the KAR2 gene. Finally, we provide evidence suggesting that yeast cells monitor the concentration of free BiP in the ER and adjust the level of transcription of the KAR2 gene accordingly; this effect is mediated via the UPR element in the KAR2 promoter.