Overexpression of HAC1 gene increased levels of both intracellular and secreted human kringle fragment in Saccharomyces cerevisiae
Overexpression of HAC1 gene increased levels of both intracellular and secreted human kringle fragment in Saccharomyces cerevisiae
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DOI：
10.1016/j.procbio.2012.09.006
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发表时间：
2012-12-01
影响因子：
4.4
通讯作者：
Seo, Jin-Ho
中科院分区：
文献类型：
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作者：
Lee, Tae-Hee;Bae, Yi-Hyun;Seo, Jin-Ho
在含有16个拷贝的人Kringle蛋白基因LK8的重组酿酒酵母菌株中，研究了未折叠蛋白反应（UPR）基因过表达的影响。In S. Had p作为转录激活因子发挥重要作用，Ire 1 p作为内切核酸酶作用于HAC1转录物以去除内含子并诱导UPR。这两个基因的破坏对LK8表达是有害的，有趣的是，hac1 Delta菌株不能利用半乳糖作为碳源，并且与野生型和ire1 Delta菌株相比，伴随着延迟细胞生长。在互补试验中，生长缺陷通过GAL 1启动子控制的HAC1基因的过表达而部分恢复。UPR的额外激活由GAL 1启动子介导，该启动子驱动HAC1基因的共表达，并分别将LK8的细胞内和分泌水平提高了4倍和1.6倍。UPR是异源生产LK8所必需的。此外，Hac1p是促进重组S.酿酒酵母菌株。这一结果表明，UPR的额外激活可能是一个很好的选择过量生产异源蛋白在S。啤酒。(C)2012爱思唯尔有限公司保留所有权利。
Effects of overexpression of an unfolded protein response (UPR) gene were investigated in recombinant Saccharomyces cerevisiae strains harboring 16 copies of the gene for human kringle protein, LK8. In S. cerevisiae, Had p plays a crucial role as a transcription activator and Ire1 p as endonuclease acting on the HAC1 transcript to remove an intron and induce the UPR. The disruption of the two genes was detrimental to LK8 expression, and interestingly, the hac1 Delta strain was not able to utilize galactose as a carbon source and concomitantly delayed cell growth compared with the wild type and the ire1 Delta, strains. In a complementation test, the growth defect was partially recovered by the overexpression of the HAC1 gene controlled by the GAL1 promoter. Additional activation of UPR was mediated by the GAL1 promoter driven coexpression of the HAC1 gene and enhanced the intracellular and secreted levels of LK8 by 4- and 1.6-fold, respectively. The UPR was essentially required for the heterologous production of LK8. Furthermore, Hac1p is the factor promoting LK8 protein production as well as cell growth in recombinant S. cerevisiae strains. This result indicates that the additional activation of UPR might be a good option for overproduction of heterologous proteins in S. cerevisiae. (C) 2012 Elsevier Ltd. All rights reserved.