The Role of AP-1 and Other Transcription Factors in Canc

本研究的总体目标是识别和表征在肿瘤促进和肿瘤进展过程中推动限速步骤的基因调控事件。AP-1转录因子是Jun和Fos家族蛋白的异源二聚体，其与某些基因的转录启动子上的特定序列结合并驱动其转录。我们对小鼠JB 6细胞和工程小鼠模型的观察（伯恩斯坦和科尔本，科学，1989，Dong等PNAS，1994，Young等，PNAS，1999）确定了AP-1活化是皮肤肿瘤发生和肿瘤进展所必需的。转基因小鼠中显性负性Jun的角质形成细胞特异性表达抑制诱导的AP-1和肿瘤发生，而不抑制与人类致癌相关的多种小鼠模型中的生长、分化或增生。其中包括皮肤肿瘤促进反应因人乳头瘤病毒E7的表达而升高的小鼠（Young等Molec Carc 2002）和通过反复暴露于UVB而诱导形成鳞状细胞癌的小鼠（库珀等Molec Cancer Res 2003）。为了进一步研究，在Powel Brown和Jay Tischlaar的实验室中，四环素调节的TAM 67表达被定向到乳腺和肺上皮。当转基因K14-TAM 67或四环素调节的TAM 67构建体在更进展的人细胞系中表达时，肿瘤细胞表型被抑制，所述人细胞系是致瘤性或锚定非依赖性的，并且显示出升高的AP-1和NF κ B活性（Li et al.，Oncogene，1998和Li等人，Molec Carcinog 2000）。转录因子NF κ B与AP-1协同调节，表明两种因子在转化中的可能重要性（Li et，Cancer Res 1997）。最近的观察已经将NFkB无应答性鉴定为JB 6模型中转化无应答性的解释（Hsu等，Cancer Res 2001，Hu等Carcinogenesis 2004）。转化抗性细胞的转化无应答性归因于不能激活NF κ B p65蛋白。在S536处的p65磷酸化对于DNA结合和抑制剂IkappaB α的泛素化和降解都是重要的（Hu et al Molec Carcinog 2005）。因此，靶向AP-1和NFkB升高防止肿瘤促进和进展的观察结果已经从小鼠JB 6模型扩展到小鼠和人角质形成细胞进展模型，以及转基因小鼠模型。对关键分子相互作用的新认识正在出现。表达AP-1/ NFkB抑制剂TAM 67的转基因小鼠提供了鉴定AP-1或NFkB靶基因的宝贵机会，所述靶基因的表达对于肿瘤转化是关键的。表达微阵列分析揭示了潜在的TAM 67靶基因，是目前研究的主题。这样的靶基因可能是用于癌症预防的有希望的新分子靶标（Young等Trends in Molec Medicine 2003）。最近的研究已经排除了iNOS（达尔等人，Mol Cancer Ther 2003），并且其他研究已经确定了染色质结构蛋白HMGA 1（达尔等人，Oncogene 2004）作为功能上重要的TAM 67靶标的重要性。针对鉴定活化AP-1所需的MAP激酶ERK依赖性分子事件的研究已经鉴定Fos家族蛋白Fra-1的活化为关键事件（Young等，Molec Cell Biol 2002）。Erk或Fra-1蛋白缺陷的JB 6变体是AP-1和转化无响应的。分别通过Erk或Fra-1的表达恢复反应性。AP-1依赖性基因启动子的一个子集预期需要Fra-1用于转录激活，并且这些可以作为癌症预防的特别有效的靶标。

The overall aim of this research is to identify and characterize gene regulation events that propel rate-limiting steps during tumor promotion and tumor progression. The AP-1 transcription factor is a heterodimer of Jun and Fos family proteins that binds to a specific sequence on the transcriptional promoter of certain genes and drives their transcription. Our observations with mouse JB6 cells and with an engineered mouse model (Bernstein and Colburn, Science, 1989, Dong et al PNAS, 1994, Young et al., PNAS, 1999) established that AP-1 activation is required for skin tumorigenesis and tumor progression. Keratinocyte-specific expression of Dominant Negative Jun in transgenic mice inhibits induced AP-1 and tumorigenesis without inhibiting growth, differentiation or hyperplasia in multiple mouse models relevant to human carcinogenesis. Among these are mice whose skin tumor promotion response is elevated by expression of Human Papilloma Virus E7 (Young et al Molec Carc 2002) and mice induced to form squamous carcinomas by repeated exposure to UVB (Cooper et al Molec Cancer Res 2003). For further studies, tetracycline regulated expression of TAM 67 is being directed to mammary and lung epithelia in the laboratories of collaborators Powel Brown and Jay Tischlaar. When the transgene K14-TAM67 or a tetracycline regulated TAM67 construct was expressed in the more progressed human cell lines that are tumorigenic or anchorage independent and show elevated AP-1 and NFkB activities, tumor cell phenotype was suppressed (Li et al., Oncogene, 1998 and Li et al., Molec Carcinog 2000). The transcription factor NFkappa B is coordinately regulated with AP-1, suggesting the possible importance of both factors in transformation (Li et, Cancer Res 1997). Recent observations have identified NFkB non-responsiveness as an explanation for transformation non-responsiveness in the JB6 model (Hsu et al, Cancer Res 2001, Hu et al Carcinogenesis 2004). Transformation resistant cells owe their transformation nonresponsiveness to an inability to activate NFkappa B p65 protein. p65 phosphorylation at S536 is important both for DNA binding and for ubiquitination and degradation of inhibitor IkappaB alpha (Hu et al Molec Carcinog 2005). Thus the observation that targeting AP-1 and NFkB elevation prevents tumor promotion and progression has been extended from the mouse JB6 model to mouse and human keratinocyte progression models, and to transgenic mouse models. New understanding of critical molecular interactions is emerging. Transgenic mice expressing AP-1/ NFkB inhibitor TAM 67 present a valuable opportunity to identify AP-1 or NFkB target genes whose expression is critical to neoplastic transformation. Expression microarray analysis has revealed potential TAM67 target genes that are subjects of current research. Such target genes may be promising new molecular targets for cancer prevention (Young et al Trends in Molec Medicine 2003). Recent studies have excluded iNOS (Dhar et al Mol Cancer Ther 2003)and other studies have established the importance of chromatin architectural protein HMGA1(Dhar et al Oncogene 2004) as a functionally significant TAM67 target. Research directed to identifying MAP kinase ERK dependent molecular events required for activating AP-1 has identified the activation of Fos family protein Fra-1 as a pivotal event (Young et al, Molec Cell Biol 2002). JB6 variants deficient in either Erk or Fra-1 proteins are AP-1 and transformation nonresponsive. Responsiveness is restored by expression of Erk or Fra-1 respectively. A subset of AP-1 dependent gene promoters is expected to require Fra-1 for transcriptional activation and these may serve as particularly effective targets for cancer prevention.