CHARACTERIZATION OF A NOVEL MAP/ERK KINASE

丝裂原活化蛋白激酶（mitogen-activated protein kinases，MAPKs）是一个高度同源的丝氨酸/苏氨酸蛋白激酶超家族，参与细胞核的生长和分化促进信号的转导以及应激反应。酵母中至少有六种MAP激酶，这表明哺乳动物中可能有更多。为了检测另外的MAP激酶，我们使用简并聚合酶链反应（PCR）筛选了大鼠脑文库，并鉴定了称为ERK 7的新型MAPK，其编码61 kD、546个氨基酸长的蛋白质ERK 7，其含有其它ERK的Thr-Glu-Tyr（TEY）活化基序特征，但具有许多独特的性质。ERK 7具有离散的C-末端结构域，其含有SH 3结合基序。与其他ERK不同，ERK 7具有显著的组成型激酶活性，并且该活性依赖于C-末端结构域。此外，ERK 7的C-末端结构域而不是其激酶活性是核定位及其作为DNA合成抑制剂的功能所必需的。最后，ERK 7是第一个被发现与激活氯离子转运的蛋白CLIC 3特异性相关的MAP激酶，并且C-末端结构域足以与CLIC 3结合。MAPK与C-末端SH 3结合结构域的鉴定和C-末端在ERK 7功能中的重要性表明，ERK 7代表MAPK的一个亚家族，其具有在生长和发育中促进信号传导特异性的接头结构域。在本申请中，我们建议进一步表征这种新型激酶的调节和功能。具体而言，我们计划使用分子和细胞方法1）确定ERK 7激活的机制; 2）确定ERK 7的关键功能结构域，结合伴侣和潜在底物; 3）研究ERK 7在细胞和组织中的功能。这些研究的结果将验证ERK 7与其他ERK一样在细胞生长和肿瘤进展的调控中发挥关键作用的假设，并将增加我们对MAPK家族这一新成员的理解。

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.