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首页> 外文期刊>Molecular and Cellular Biology >Evidence for Homodimerization of the c-Fos Transcription Factor in Live Cells Revealed by Fluorescence Microscopy and Computer Modeling
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Evidence for Homodimerization of the c-Fos Transcription Factor in Live Cells Revealed by Fluorescence Microscopy and Computer Modeling

机译：荧光显微镜和计算机建模揭示活细胞中c-Fos转录因子均一化的证据

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The c-Fos and c-Jun transcription factors, members of the activator protein 1 (AP-1) complex, form heterodimers and bind to DNA via a basic leucine zipper and regulate the cell cycle, apoptosis, differentiation, etc. Purified c-Jun leucine zipper fragments could also form stable homodimers, whereas c-Fos leucine zipper homodimers were found to be much less stable in earlier in vitro studies. The importance of c-Fos overexpression in tumors and the controversy in the literature concerning c-Fos homodimerization prompted us to investigate Fos homodimerization. F?rster resonance energy transfer (FRET) and molecular brightness analysis of fluorescence correlation spectroscopy data from live HeLa cells transfected with fluorescent-protein-tagged c-Fos indicated that c-Fos formed homodimers. We developed a method to determine the absolute concentrations of transfected and endogenous c-Fos and c-Jun, which allowed us to determine dissociation constants of c-Fos homodimers (Kd = 6.7 ± 1.7 μM) and c-Fos–c-Jun heterodimers (on the order of 10 to 100 nM) from FRET titrations. Imaging fluorescence cross-correlation spectroscopy (SPIM-FCCS) and molecular dynamics modeling confirmed that c-Fos homodimers were stably associated and could bind to the chromatin. Our results establish c-Fos homodimers as a novel form of the AP-1 complex that may be an autonomous transcription factor in c-Fos-overexpressing tissues and could contribute to tumor development.
机译：激活蛋白1（AP-1）复合物的成员c-Fos和c-Jun转录因子形成异二聚体，并通过碱性亮氨酸拉链与DNA结合，并调节细胞周期，凋亡，分化等。 Jun亮氨酸拉链片段也可以形成稳定的同二聚体，而c-Fos亮氨酸拉链同二聚体在早期的体外研究中被发现稳定性较差。 c-Fos过度表达在肿瘤中的重要性以及有关c-Fos同型二聚化的文献中的争议促使我们研究Fos同型二聚化。用荧光蛋白标记的c-Fos转染的活HeLa细胞的Fster共振能量转移（FRET）和荧光相关光谱数据的分子亮度分析表明，c-Fos形成了同二聚体。我们开发了一种确定转染和内源c-Fos和c-Jun的绝对浓度的方法，这使我们能够确定c-Fos同二聚体（ K d = 6.7±1.7μM）和c-Fos–c-Jun异二聚体（约10至100 nM）。成像荧光互相关光谱（SPIM-FCCS）和分子动力学模型证实，c-Fos同二聚体稳定缔合并可以与染色质结合。我们的结果将c-Fos同型二聚体确定为AP-1复合物的一种新型形式，它可能是c-Fos过表达组织中的自主转录因子，并可能有助于肿瘤的发展。

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