Analyses of Maltipotential Functions of a Novel Signaling Molecule, Cas

p130cas是一种对接蛋白，包含一个SH3结构域和多个酪氨酸残基。p130cas位于局灶黏附处，在整合素刺激下酪氨酸磷酸化，并被认为通过c-Crk和其他蛋白质将信号传递给肌动蛋白应激纤维的重塑和细胞运动。在Far Western筛选p130cas SH3结构域配体的过程中，我们克隆了一个名为CIZ的新蛋白。CIZ由一个假定的亮氨酸拉链、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C2H2锌指和谷氨酰胺-丙氨酸重复序列组成。CIZ与细胞中的Cas结合，位于细胞核和局灶黏附处。我们利用瞬时种间异核体形成实验证明了CIZ是一种核-胞质穿梭蛋白。为了在细胞核中寻找CIZ的靶点，我们通过cast分析确定了CIZ的DNA结合一致性为（G/C）AAAAA(A)。在基质金属蛋白酶（降解细胞外基质蛋白的酶）的几个启动子中发现了共识样序列。CIZ与MMP-1（胶原酶）启动子中的共识样序列结合。CIZ的过表达上调了MMP-1、MMP-3（基质溶素）和MMP-7（基质溶素）启动子的转录，并且在Cas的存在下，这种转录激活得到增强。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是一种结合Cas的新型锌指蛋白，是一种核-胞质穿梭蛋白，可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.