CHARACTERIZATION OF A NOVEL MAP/ERK KINASE

丝裂原活化蛋白激酶（MAPKs）是一个高度同源的脯氨酸定向丝氨酸/苏氨酸激酶超家族，参与生长和分化促进信号的转导，以及对细胞核的应激反应。酵母中至少有六种MAP激酶的存在表明哺乳动物中可能有更多。为了检测更多的MAP激酶，我们使用退化聚合酶链反应（PCR）筛选了一个大鼠脑文库，并鉴定了一个名为ERK7的新MAPK，它编码一个61 kD， 546个氨基酸的长蛋白ERK7，它包含其他erk的Thr-Glu-Tyr （TEY）激活基序特征，但具有许多独特的特性。ERK7具有包含SH3结合基序的离散c端结构域。与其他ERKs不同，ERK7具有显著的组成激酶活性，并且这种活性依赖于c端结构域。此外，ERK7的c端结构域而不是其激酶活性是核定位和其作为DNA合成抑制剂的功能所必需的。最后，ERK7是第一个被发现与一种激活氯离子转运蛋白CLIC3特异性结合的MAP激酶，其c端结构域足以与CLIC3结合。具有c端sh3结合结构域的MAPK的鉴定以及c端在ERK7功能中的重要性表明，ERK7代表了具有适配器结构域的MAPK亚家族，这些结构域有助于生长和发育中的信号特异性。在这个应用中，我们建议进一步表征这种新激酶的调控和功能。具体来说，我们计划使用分子和细胞方法来确定ERK7被激活的机制；2)确定ERK7的关键功能域、结合伙伴和潜在底物；3)研究ERK7在细胞和组织中的功能。这些研究的结果将验证ERK7和其他erk一样，在细胞生长和肿瘤进展的调节中起关键作用的假设，并将增加我们对MAPK家族新成员的理解。

The mitogen-activated protein kinases (MAPKs) are a superfamily of highly homologous proline-directed serine/threonine kinases that participate in the transduction of growth and differentiation-promoting signals, as well as stress responses to the cell nucleus. The presence of at least six MAP kinases in yeast suggests that there are likely to more in mammals. In order to detect additional MAP kinases, we screened a rat brain library using degenerate polymerase chain reaction (PCR) and identified a novel MAPK termed ERK7 that encodes a 61 kD, 546 amino- acid long protein ERK7 contains the Thr-Glu-Tyr (TEY) activation motif characteristics of other ERKs, but has a number of properties that are unique. ERK7 has a discrete C-terminal domain that contains SH3 binding motifs. Unlike other ERKs, ERK7 has significant constitutive kinase activity, and this activity is dependent upon the C-terminal domain. Furthermore, ERK7's C-terminal domain rather than its kinase activity is required for nuclear localization and its function as an inhibitor of DNA synthesis. Finally, ERK7 is the first MAP kinase that has been found to specifically associate with a protein that activates chloride ion transport, CLIC3, and the C-terminal domain is sufficient for CLIC3 binding. The identification of a MAPK with C-terminal SH3-binding domains and the importance of the C-terminus in ERK7 function suggests that ERK7 represents a subfamily of MAPKs with adaptor domains that contribute to signaling specificity in growth and development. In this application, we propose to further characterize the regulation and function of this novel kinase. Specifically, we plan to use molecular and cellular approaches to 1) Determine the mechanism by which ERK7 is activated; 2) Identify key functional domains, binding partners and potential substrates of ERK7; and 3) Investigate the function of ERK7 in cell and tissues. The results of these studies will test the hypothesis that ERK7, like other ERKs, play a key role in the regulation of cell growth and tumor progression, and will increase our understanding of this new member of the MAPK family.