MULTIPLE MECHANISMS OF SKN-1 FUNCTION IN VIVO

描述（由申请人提供）：C。线虫转录因子SKN-1 指形成消化系统的早期胚胎细胞，但也有 胚胎后的功能尚未阐明。SKN-1通过一种 不寻常的机制，但与两种脊椎动物NRF bZIP蛋白有关， 特别是在一个叫做DIDLID的独特反式激活元件中。脊椎动物 NRF蛋白是抵抗氧化应激所必需的，并且直接 激活抗氧化酶基因。SKN-1同样是氧化性 抗应激，并直接激活肠道表达的一个关键 抗氧化酶这表明，SKN-1和NRF蛋白保持了 平行的功能和作用机制，以及对 SKN-1在C.线虫将适用于脊椎动物的抗氧化剂 的防御合作.我们的初步发现提出了三个相关的假设， 现在将在体内进行测试：SKN-1具有多种功能， SKN-1通过不同的机制作用于不同的靶基因 依赖或独立于cbp-1的C.线虫p300/CBP 组蛋白乙酰转移酶（HAT），并且保守的DIDLID元件是 对于其CBP-1独立功能至关重要。为了测试这些模型，我们将 研究SKN-1在抗氧化防御中的参与， 解毒基因激活我们wifi也使用定点突变来 在体内测试包括DIDLID及其CBP-1结合的SKN-1元件是否 区域需要履行其抗氧化和胚胎功能， 激活相关的靶基因。最后，我们将调查 SKN-1通过决定如何与CBP-1结合而发挥作用的机制 以及其功能是否涉及其他HAT，并通过识别生理 DIDLID和其他CBP-1非依赖性SKN-1元件的蛋白靶点。这项工作 将阐明保守的转录调控相互作用， 在对氧化应激的转录反应中保守， 消化系统功能，C.优雅是最初的基础 消化系统的规格。它还将应用独特的优势， 氏梭elegans系统来阐明体内后生动物转录 因子通过不同的机制调节细胞类型特异性基因， 不同的监管背景。

DESCRIPTION (provided by applicant): The C. elegans transcription factor SKN-1 specifies early embryonic cells which form the digestive system, but also has postembryonic functions that have not been elucidated. SKN-1 binds DNA by an unusual mechanism, but is related to the two vertebrate NRF bZIP proteins, especially within a unique transactivation element called DIDLID. Vertebrate NRF proteins are required for resistance to oxidative stresses, and directly activate antioxidant enzyme genes. SKN-1 is similarly required for oxidative stress resistance, and directly activates intestinal expression of a critical antioxidant enzyme. This suggests that SKN-1 and NRF proteins have maintained parallel functions, and mechanisms of action, and that insights obtained into SKN-1 functions in C. elegans will be applicable to vertebrate antioxidant defenses. Our preliminary findings suggest three related hypotheses that we will now test in vivo: that SKN-1 has multiple functions which parallel those of NRF proteins, that SKN-1 acts at different target genes through mechanisms that are dependent upon or independent of cbp-i, the C. elegans p300/CBP histone acetyl transferase (HAT), and that the conserved DIDLID element is critical for its cbp-1-independent functions. To test these models, we will investigate the involvement of SKN-1 in antioxidant defenses, and in direct detoxification gene activation. We wifi also use site-directed mutagenesis to test in vivo whether SKN-1 elements that include DIDLID and its CBP-1--binding region are required to fulfil its antioxidant and embryonic functions, and to activate the respective target genes involved. Finally, we will investigate the mechanisms through which SKN-1 functions by determining how it binds to CBP-1 and whether its functions involve other HATs, and by identifying physiological protein targets of DIDLID and other CBP-1-independent SKN-1 elements. This work will elucidate conserved transcriptional regulatory interactions that have been conserved in transcriptional responses to oxidative stress, an ancestral digestive system function, and that in C. elegans are fundamental to initial specification of the digestive system. It will also apply the unique advantages of the C. elegans system to elucidate in vivo how a metazoan transcription factor regulates cell-type-specific genes through distinct mechanisms in different regulatory contexts.