Cell density regulates intracellular localization of aryl hydrocarbon receptor
Cell density regulates intracellular localization of aryl hydrocarbon receptor
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DOI：
10.1074/jbc.m310492200
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发表时间：
2004-04-30
影响因子：
4.8
通讯作者：
Kawajiri, K
中科院分区：
文献类型：
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作者：
Ikuta, T;Kobayashi, Y;Kawajiri, K
芳烃受体（AhR）是一种配体激活的转录因子，在异种生物信号通路中起着细胞内介质的作用。AhR包含核定位信号和核输出信号。本研究的目的是证明AhR在细胞内的分布是如何受到生理调节的。我们发现，在角质细胞细胞系中，细胞密度而不是细胞周期影响AhR的亚细胞分布，HaCaT: AhR在稀疏细胞密度下主要是核性的，在亚汇合时是核性和细胞质性的，在汇合时主要是细胞质性的。携带与外源应答元件融合的报告基因的HaCaT稳定转染表明，外源应答元件介导的转录与AhR重定位之间存在关联。Leptomycin B促进AhR的核积累，而不考虑细胞密度，这表明这种改变可能是由于AhR核输出调节的改变。我们发现，活化的AhR在细胞核积累后，AhR NES中的Ser-68被磷酸化，嵌合GST-AhR-GFP融合蛋白的核输出被丝氨酸残基（Ser-68）取代为天冬氨酸，这模拟了磷酸化的负电荷。这种新的细胞密度依赖性AhR重定位受到暴露于SB203580、冈田酸和低Ca2+浓度的影响。这些发现强烈表明，细胞密度通过调节核输出活动调节AhR的细胞内定位和功能。p38 mapk介导的NES磷酸化及其去磷酸化，受细胞-细胞接触信号调节，可能在新的AhR重定位中起关键作用。
The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor that plays a role as an intracellular mediator of the xenobiotic signaling pathway. AhR contains signals for both nuclear localization and nuclear export (NES). The objective of this study was to demonstrate how AhR intracellular distribution was regulated physiologically in cells. We found that cell density, but not the cell cycle, influenced the subcellular distribution of AhR in a keratinocyte cell line, HaCaT: AhR was predominantly nuclear at sparse cell densities, both nuclear and cytoplasmic at subconfluence, and predominantly cytoplasmic at confluence. Stable transfectants of HaCaT carrying a reporter gene fused with xenobiotic responsive element showed an association between xenobiotic responsive element-mediated transcription and AhR relocalization. Leptomycin B promoted nuclear accumulation of AhR irrespective of cell density, suggesting that this alteration may be because of a change of the regulation of the nuclear export of AhR. We found that Ser-68 in the NES of AhR was phosphorylated after nuclear accumulation of activated AhR and the nuclear export of a chimeric GST-AhR-GFP fusion protein was suppressed by substitution of a serine residue (Ser-68) to aspartic acid, which mimics the negative charge of phosphorylation. This novel cell density-dependent AhR relocalization was affected by exposure to SB203580, okadaic acid, and low Ca2+ concentrations. These findings strongly suggest that cell density regulates the intracellular localization and function of AhR, because of modulation of nuclear export activity. The p38 MAPK-mediated phosphorylation of the NES and its dephosphorylation, regulated by cell-cell contact signals, may have pivotal roles in the novel AhR relocalization.