Reisz-Porszasz S, Probst M R, Fukunaga B N, Hankinson O
Laboratory of Structural Biology and Molecular Medicine, University of California, Los Angeles 90024-1786.
Mol Cell Biol. 1994 Sep;14(9):6075-86. doi: 10.1128/mcb.14.9.6075-6086.1994.
The activated aryl hydrocarbon receptor (AHR) and the AHR nuclear translocator (ARNT) bind DNA as a heterodimer. Both proteins represent a novel class of basic helix-loop-helix (bHLH)-containing transcription factors in that (i) activation of AHR requires the binding of ligand (e.g., 2,3,7,8-tetrachlorodibenzo-p-dioxin [TCDD]), (ii) the xenobiotic responsive element (XRE) recognized by the AHR/ARNT heterodimer differs from the recognition sequence for nearly all other bHLH proteins, and (iii) both proteins contain a PAS homology region, which in the Drosophila PER and SIM proteins functions as a dimerization domain. A cDNA for mouse ARNT has been cloned, and potential functional domains of ARNT were investigated by deletion analysis. A mutant lacking all regions of ARNT other than the bHLH and PAS regions is unimpaired in TCDD-dependent dimerization and subsequent XRE binding and only modestly reduced in ability to complement an ARNT-deficient mutant cell line, c4, in vivo. Both the first and second alpha helices of the bHLH region are required for dimerization. The basic region is required for XRE binding but not for dimerization. Deletion of either the A or B segments of the PAS region slightly affects TCDD-induced heterodimerization, while deletion of the complete PAS region severely affects (but does not eliminate) dimerization. Thus, ARNT possesses multiple domains required for maximal heterodimerization. Mutants deleted for PAS A, PAS B, and the complete PAS region all retain some degree of XRE binding, yet none can rescue the c4 mutant. Therefore, both the PAS A and PAS B segments, besides contributing to dimerization, apparently fulfill additional, unknown functions required for biological activity of ARNT.
活化的芳烃受体（AHR）和AHR核转运蛋白（ARNT）作为异源二聚体与DNA结合。这两种蛋白质代表了一类新型的含碱性螺旋-环-螺旋（bHLH）的转录因子，因为：（i）AHR的激活需要配体（如2,3,7,8-四氯二苯并-p-二恶英[TCDD]）的结合；（ii）AHR/ARNT异源二聚体识别的外源性反应元件（XRE）与几乎所有其他bHLH蛋白的识别序列不同；（iii）这两种蛋白质都含有一个PAS同源区域，在果蝇的PER和SIM蛋白中该区域作为二聚化结构域发挥作用。小鼠ARNT的cDNA已被克隆，并通过缺失分析研究了ARNT的潜在功能结构域。一个除了bHLH和PAS区域外缺少ARNT所有其他区域的突变体，在TCDD依赖性二聚化和随后的XRE结合方面未受损害，并且在体内补充ARNT缺陷型突变细胞系c4的能力仅略有降低。bHLH区域的第一个和第二个α螺旋是二聚化所必需的。碱性区域是XRE结合所必需的，但不是二聚化所必需的。PAS区域的A或B片段的缺失对TCDD诱导的异源二聚化有轻微影响，而完整PAS区域的缺失则严重影响（但不消除）二聚化。因此，ARNT拥有最大程度异源二聚化所需的多个结构域。缺失PAS A、PAS B和完整PAS区域的突变体都保留了一定程度的XRE结合能力，但没有一个能够拯救c4突变体。因此，PAS A和PAS B片段除了有助于二聚化外，显然还履行了ARNT生物活性所需的其他未知功能。
