Identification of nucleotides responsible for enhancer activity of sterol regulatory element in low density lipoprotein receptor gene.
Identification of nucleotides responsible for enhancer activity of sterol regulatory element in low density lipoprotein receptor gene.
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DOI：
10.1016/s0021-9258(19)39976-4
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发表时间：
1990-02
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影响因子：
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通讯作者：
Jeffrey R. Smith;T. Osborne;J. Goldstein;Michael S. Brown
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文献类型：
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作者：
Jeffrey R. Smith;T. Osborne;J. Goldstein;Michael S. Brown
低密度脂蛋白（LDL）受体启动子的甾醇依赖性调节先前已定位于该基因5 '侧翼区的16个碱基对序列，命名为重复序列2。在目前的研究中，我们发现重复序列2的中心10个核苷酸对甾醇调节活性至关重要。该序列包括八聚体，命名为固醇调节元件1（SRE-1），其先前在胆固醇生物合成的固醇调节酶3-羟基-3-甲基戊二酰辅酶A合酶的基因启动子中被鉴定。我们在完整LDL受体启动子的1471个碱基对片段内进行了一系列单碱基替换，通过转染将突变质粒引入仓鼠细胞，并在不存在和存在甾醇的情况下测量mRNA水平。重复序列2中的10个碱基对序列内的取代在很大程度上阻止了在没有甾醇的情况下发生的转录诱导。这些点突变都不影响甾醇存在下的转录。像增强子一样，重复序列2中的SRE-1以不依赖于方向的方式发挥作用。我们解释这些研究结果表明，SRE-1的LDL受体启动子是一个条件性的积极因素，与其他元素合作，以提高转录在没有甾醇和甾醇的存在下失去其功能。
Sterol-dependent regulation of the low density lipoprotein (LDL) receptor promoter has been localized previously to a 16-base pair sequence, designated repeat 2, in the 5'-flanking region of the gene. In the current study, we show that the central 10 nucleotides of repeat 2 are crucial for the sterol regulatory activity. This sequence includes an octamer, designated sterol regulatory element 1 (SRE-1), which was identified previously in the promoter of the gene for 3-hydroxy-3-methylglutaryl coenzyme A synthase, a sterol-regulated enzyme of cholesterol biosynthesis. We made a series of single-base substitutions within a 1471-base pair fragment of the intact LDL receptor promoter, introduced the mutant plasmids into hamster cells by transfection, and measured mRNA levels in the absence and presence of sterols. Substitutions within the 10-base pair sequence in repeat 2 largely prevented the induction of transcription which occurs in the absence of sterols. None of these point mutations affected transcription in the presence of sterols. Like an enhancer, the SRE-1 in repeat 2 functioned in an orientation-independent manner. We interpret these findings to indicate that the SRE-1 of the LDL receptor promoter is a conditional positive element that cooperates with other elements to enhance transcription in the absence of sterols and loses its function in the presence of sterols.