Lindebro M C, Poellinger L, Whitelaw M L
Department of Medical Nutrition, Karolinska Institute, Huddinge University Hospital, Sweden.
EMBO J. 1995 Jul 17;14(14):3528-39. doi: 10.1002/j.1460-2075.1995.tb07359.x.
Gene regulation by dioxins is mediated by the dioxin receptor-Arnt heterodimer, a ligand generated complex of two basic helix-loop-helix (bHLH)/Per-Arnt-Sim (PAS) transcription factors. By using dioxin receptor chimeras where the dimerization and DNA binding bHLH motif has been replaced by a heterologous DNA binding domain, we have detected an ability of Arnt to interact with the dioxin receptor via the PAS domain in a mammalian 'hybrid interaction' system. By coimmunoprecipitation assays, we have confirmed the ability of PAS domains of the dioxin receptor and Arnt to mediate independent heterodimerization in vitro. Selectivity for PAS dimerization was noted in our hybrid interaction system, as dioxin receptor or Arnt PAS-mediated homodimers were not detected. Surprisingly, however, the PAS domain of Per could dimerize with both the dioxin receptor and Arnt subunits in vitro, and disrupt the ability of these subunits to form a DNA binding heterodimer. Moreover, ectopic expression of Per blocked dioxin signalling in mammalian cells. The PAS domains of the dioxin receptor and Arnt are therefore novel dimerizing regions critical in formation of a functional dioxin receptor-Arnt complex, while the PerPAS domain is a potential negative regulator of bHLH/PAS factor function.
二噁英对基因的调控是由二噁英受体 - Arnt异二聚体介导的，该异二聚体是由两个碱性螺旋 - 环 - 螺旋（bHLH）/Per - Arnt - Sim（PAS）转录因子形成的配体复合物。通过使用二噁英受体嵌合体，其中二聚化和DNA结合bHLH基序已被异源DNA结合结构域取代，我们在哺乳动物的“杂交相互作用”系统中检测到Arnt通过PAS结构域与二噁英受体相互作用的能力。通过免疫共沉淀分析，我们证实了二噁英受体和Arnt的PAS结构域在体外介导独立异二聚化的能力。在我们的杂交相互作用系统中发现了对PAS二聚化的选择性，因为未检测到二噁英受体或Arnt PAS介导的同二聚体。然而，令人惊讶的是，Per的PAS结构域在体外可与二噁英受体和Arnt亚基都形成二聚体，并破坏这些亚基形成DNA结合异二聚体的能力。此外，Per的异位表达在哺乳动物细胞中阻断了二噁英信号传导。因此，二噁英受体和Arnt的PAS结构域是形成功能性二噁英受体 - Arnt复合物的关键新二聚化区域，而Per PAS结构域是bHLH/PAS因子功能的潜在负调节剂。
