Dominant-Negative Lox-1 Blocks Homodimerization of Wild-Type Lox-1–Induced Cell Proliferation Through Extracellular Signal Regulated Kinase 1/2 Activation
Dominant-Negative Lox-1 Blocks Homodimerization of Wild-Type Lox-1–Induced Cell Proliferation Through Extracellular Signal Regulated Kinase 1/2 Activation
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DOI：
10.1161/01.hyp.0000229825.98545.5e
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发表时间：
2006-08
期刊：
影响因子：
8.3
通讯作者：
H. Tanigawa;S. Miura;Yoshino Matsuo;M. Fujino;T. Sawamura;K. Saku
中科院分区：
文献类型：
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作者：
H. Tanigawa;S. Miura;Yoshino Matsuo;M. Fujino;T. Sawamura;K. Saku
C型凝集素样氧化低密度脂蛋白（Ox-LDL）受体-1（Lox-1）与自然杀伤细胞受体Ly 49 A和CD 94属于同一家族，并在功能上经历二聚化。尽管牛（B）Lox-1 C端的Lys 262和Lys 263在Ox-LDL的摄取中起重要作用，但这些残基的突变尚未被认为是显性负性的潜在来源。我们假设显性阴性人（h）Lox-1与Lox-1-野生型（WT）形成异源二聚体并阻断Lox-1-WT诱导的细胞信号传导。基于使用分子成像技术与激光扫描共聚焦显微镜和免疫沉淀在hLox-1表达的中国仓鼠卵巢细胞系统中，hLox-1-WT的同源二聚体定位于细胞膜，Ox-LDL激活细胞外信号调节激酶（ERK）1/2没有hLox-1-WT的易位。hLox-1的Lys 266和Lys 267对应于bLox-1的Lys 262和Lys 263，被突变（hLox 1-K266 A/K267 A），并且突变的受体抑制hLox-1-WT诱导的胸苷掺入和ERK 1/2激活。尽管Ox-LDL与显性负性突变体受体结合并被细胞质吸收，但ERK 1/2激活被细胞膜中突变体受体和hLox-1-WT的异源二聚体化阻断。此外，在表达hLox-1-WT的人冠状动脉平滑肌细胞中，我们证实了ERK 1/2和[3 H]-胸苷掺入的激活是由Ox-LDL的加入引起的，并且这些作用被hLox 1-K266 A/K267 A阻断。总之，目前的研究结果构成了第一个证据表明，旨在阻断细胞增殖途径在受体水平的策略可能是有用的，为削弱Lox-1诱导的细胞增殖。
C-type lectin-like oxidized low-density lipoprotein (Ox-LDL) receptor-1 (Lox-1) belongs to the same family as natural killer cell receptors Ly49A and CD94 and functionally undergoes dimerization. Although Lys262 and Lys263 in the C terminus of bovine (b)Lox-1 play an important role in the uptake of Ox-LDL, mutation of these residues has not been suggested to be a potential source of the dominant-negative property. We hypothesize that dominant-negative human (h)Lox-1 forms a heterodimer with Lox-1–wild-type (WT) and blocks Lox-1–WT–induced cell signaling. Based on the use of molecular imaging techniques with laser scanning confocal microscopy and immunoprecipitation in an hLox-1–expressing Chinese hamster ovary cell system, homodimerization of hLox-1–WT was localized in the cell membrane, and Ox-LDL activated extracellular signal regulated kinase (ERK)1/2 without the translocation of hLox-1-WT. Lys266 and Lys267 of hLox-1, corresponding with Lys262 and Lys263 of bLox-1, were mutated (hLox1-K266A/K267A), and the mutant receptor inhibited hLox-1–WT–induced thymidine incorporation and ERK1/2 activation. Although Ox-LDL binds to the dominant-negative mutant receptor and is taken up by cytoplasm, ERK1/2 activation was blocked by heterodimerization with the mutant receptor and hLox-1–WT in the cell membrane. In addition, in human coronary artery smooth muscle cells, which express hLox-1–WT, we confirmed that the activation of ERK1/2 and [3H]-thymidine incorporation was caused by the addition of Ox-LDL, and these actions were blocked by hLox1-K266A/K267A. In conclusion, the present findings constitute the first evidence that strategies aimed at blocking cell-proliferative pathways at the receptor level could be useful for impairing Lox-1–induced cell proliferation.