LEU3 OF SACCHAROMYCES-CEREVISIAE ACTIVATES MULTIPLE GENES FOR BRANCHED-CHAIN AMINO-ACID BIOSYNTHESIS BY BINDING TO A COMMON DECANUCLEOTIDE CORE SEQUENCE
LEU3 OF SACCHAROMYCES-CEREVISIAE ACTIVATES MULTIPLE GENES FOR BRANCHED-CHAIN AMINO-ACID BIOSYNTHESIS BY BINDING TO A COMMON DECANUCLEOTIDE CORE SEQUENCE
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DOI：
10.1128/mcb.8.7.2690
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发表时间：
1988-07-01
影响因子：
5.3
通讯作者：
SCHIMMEL, P
中科院分区：
文献类型：
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作者：
FRIDEN, P;SCHIMMEL, P
酿酒酵母的LEU 3编码一种886个氨基酸的多肽，该多肽调节参与亮氨酸生物合成的一组基因的转录，并且已经显示特异性结合LEU 2上游区的114个碱基对的DNA片段（P. Friden和P. Schimmel，Mol. Cell. 7：2707-2717，1987）。我们表明，除了LEU 2，LEU 3结合在体外序列的启动子区的LEU 1，LEU 4，ILV 2，并推断，ILV 5。这些基因中结合位点所共有的大部分保守的十核苷酸核心序列是CCGGNNCCGG。甲基化干扰足迹实验表明，LEU 3与位于大沟中的保守碱基进行对称性竞争。将含有LEU 1、LEU 2、LEU 4和ILV 2的核心十核苷酸和侧翼序列的合成寡核苷酸（19至29个碱基对）分别置于LEU 3不敏感的测试启动子的上游。每个构建体的表达都被LEU 3激活，尽管激活程度根据引入的特定寡核苷酸而变化很大。然而，在核心序列中具有取代的启动子构建体保持LEU 3不敏感。还测试了其中一种寡核苷酸（基于LEU 2序列），并显示其在测试启动子上赋予亮氨酸敏感性表达。结果表明，只有一个短的序列元素是必需的LEU 3依赖性启动子的结合和激活，并提供了直接的证据，扩大了由LEU 3激活的基因库。
LEU3 of Saccharomyces cerevisiae encodes an 886-amino-acid polypeptide that regulates transcription of a group of genes involved in leucine biosynthesis and has been shown to bind specifically to a 114-base-pair DNA fragment of the LEU2 upstream region (P. Friden and P. Schimmel, Mol. Cell. Biol. 7:2707-2717, 1987). We show that, in addition to LEU2, LEU3 binds in vitro to sequences in the promoter regions of LEU1, LEU4, ILV2, and, by inference, ILV5. The largely conserved decanucleotide core sequence shared by the binding sites in these gene is CCGGNNCCGG. Methylation interference footprinting experiments show that LEU3 makes symmetrical contasts with the conserved bases that lie in the major groove. Synthetic oligonucleotides (19 to 29 base pairs) which contain the core decanucleotide and flanking sequences of LEU1, LEU2, LEU4, and ILV2 have individually been placed upstream of a LEU3-insensitive test promoter. The expression of each construction is activated by LEU3, although the degree of activation varies considerably according to the specific oligonucleotide which is introduced. A promoter construction with substitutions in the core sequence remains LEU3 insensitive, however. One of the oligonucleotides (based on a LEU2 sequence) was also tested and shown to confer leucine-sensitive expression on the test promoter. The results demonstrate that only a short sequence element is necessary for LEU3-dependent promoter binding and activation and provide direct evidence for an expanded repertoire of genes that are activated by LEU3.