Analyses of Maltipotential Functions of a Novel Signaling Molecule, Cas

p130 cas是一种含有SH 3结构域和多个酪氨酸残基的对接蛋白。p130 cas位于粘着斑，对整联蛋白刺激作出反应而被酪氨酸磷酸化，并被认为通过c-Crk和其它蛋白质将信号传递给肌动蛋白应力纤维的重塑和细胞运动。在通过Far Western筛选寻找p130 cas的SH 3结构域配体的过程中，我们克隆了一个新的蛋白质CIZ，它由一个亮氨酸拉链、一个富含丝氨酸/苏氨酸的区域、一个富含脯氨酸的序列、5、6或8个Kruppel型C2 H2锌指和谷氨酰胺-丙氨酸重复序列组成。CIZ结合细胞中的Cas并且位于细胞核和粘着斑处。我们发现，CIZ是一个核质穿梭蛋白，使用瞬时种间异核体形成试验。为了寻找CIZ在细胞核内的作用靶点，我们通过CASTing分析确定了CIZ的DNA结合共有序列为（G/C）AAAAA（A）。基质金属蛋白酶是降解细胞外基质蛋白的酶，其启动子中存在类似共有序列。CIZ与MMP-1（胶原酶）启动子中的共识样序列结合。CIZ的过表达上调MMP-1、MMP-3（基质溶解素）和MMP-7（基质溶解素）启动子的转录，并且这种转录激活在Cas存在时增强。此外，CIZ的稳定过表达促进了培养基中MMP-7的产生。综上所述，CIZ是一种新型锌指蛋白，结合Cas，是一种核质穿梭蛋白，并调节基质金属蛋白酶的表达。

p130cas is a docking protein that contains an SH3 domain and multiple tyrosine residues. p130cas is located at focal adhesions, is tyrosine phosphorylated in response to integrin stimulation, and is thought to transmit signals, via c-Crk and other proteins, to the remodeling of actin stress fibers and to the cell movement. In the search for the ligands of the SH3 domain of p130cas by Far Western screening, we cloned a novel protein named CIZ.CIZ consists of a putative leucine zipper, serine/threonine rich region, a proline rich sequence, 5, 6 or 8 Kruppel-type C2H2 zinc fingers, and the glutamine-alanine repeat. CIZ binds Cas in cells and is located in nucleus and at focal adhesions. We showed that CIZ is a nucleo-cytoplasmic shuttling protein, using the transient interspecies heterokaryon formation assay. In order to search for the targets of CIZ in nucleus, we determined the DNA binding consensus of CIZ as (G/C)AAAAA(A) by CASTing analysis. The consensus-like sequences are found in several promoters of matrix metalloproteinases, which are the enzymes to degrade the extracellular matrix proteins. CIZ binds to a consensus-like sequence in MMP-1 (collagenase) promoter. Overexpression of CIZ upregulates the transcriptions from MMP-1, MMP-3 (stromelysin), and MMP-7 (matrilysin) promoters, and this transactivation was enhanced in the presense of Cas. Furthermore, the stable overexpression of CIZ promoted the production of MMP-7 in culture medium. In summary, CIZ, a novel zinc finger protein, binds Cas, is a nucleo-cytoplasmic shuttling protein, and regulates the expression of matrix metalloproteinases.