Molecular basis of arsenic-induced cell transformation

描述(申请人提供)：砷是一种有充分证据的人类致癌物质。我们的目标是解决这一中心假设，即MAP激酶、P90 S6激酶和组蛋白H3是砷诱导的信号转导和细胞转化的媒介。特异目的1，确定砷诱导的细胞转化是否需要p38激酶；特异性目的2，确定c-jun氨基末端激酶(JNKs)在砷诱导的细胞转化和凋亡中的作用；特异性目的3，确定p90S6激酶/MAPKAP-K1(P90RSK)的激活是否参与砷诱导的信号转导和细胞转化；特异性目的4，确定p38激酶、JNKs和p90RSK在砷诱导的组蛋白H3磷酸化中的作用以及组蛋白H3磷酸化在细胞转化中的作用。针对特定目的1的策略是使用p38激酶抑制剂或显性负突变体或siRNA来抑制p38激酶的激活。针对具体目标2，我们将(1)测试JNK磷酸化失活是否对砷耐药表型的形成是必需的；(2)测试抑制JNK磷酸化/激活是否会导致抑制亚砷酸盐诱导的细胞转化或凋亡；(3)测试将激活的JNK突变体引入砷耐药细胞是否会拯救抗凋亡表型。为了达到特定的目的3，我们将使用表达p90RSK的显性负性突变体或siRNA的细胞系和p90RSK缺陷的细胞系来抑制p90RSK。针对具体目的4，我们将(1)阻断特定的抑制剂、显性负性突变体、siRNA或基因敲除细胞系对JNK、p38和p90RSK的激活；(2)使用组蛋白H3、显性负性突变体(组蛋白H3 S10突变为A10、H3 S28突变为A28)和组蛋白H3野生型表达siRNA的细胞系来研究组蛋白H3在砷诱导的细胞转化中的作用。这些知识将有助于设计更有效、更具体、副作用更少的砷诱导癌症的化学预防策略。

DESCRIPTION (provided by applicant): Arsenic is a well-documented human carcinogen. Our goal is to address the central hypothesis that MAP kinases, p90 S6 kinases, and histone H3 are mediators of arsenic-induced signal transduction and cell transformation. The Specific Aims are: Specific Aim 1, to determine whether arsenic-induced p38 kinase is required for arsenic-induced cell transformation; Specific Aim 2, to determine the role of c-Jun N-terminal kinases (JNKs) in arsenic-induced cell transformation and apoptosis; Specific Aim 3, to determine whether activation of p90 S6 kinase/MAPKAP-K1 (p90RSK) is involved in arsenic-induced signal transduction and cell transformation; and Specific Aim 4, to determine the role of p38 kinase, JNKs, and p90RSK in arsenic-induced phosphorylation of histone H3 and the role of phosphorylation of histone H3 in cell transformation. The strategy for Specific Aim 1 is to use p38 kinase inhibitors or dominant negative mutants or siRNA to inhibit p38 kinase activation. For Specific Aim 2, we will (1) test whether inactivation of JNK phosphorylation is required for development of resistant phenotype to arsenic; (2) test whether inhibition of JNK phosphorylation/activation will cause inhibition of arsenite-induced cell transformation or apoptosis; (3) test whether introduction of an active JNK mutant into arsenic resistant cells will cause the rescue of the apoptosis-resistant phenotype. For Specific Aim 3, we will inhibit p90RSK using cell lines expressing a dominant negative mutant or siRNA of p90RSK and cell lines which are deficient in p90RSK. For Specific Aim 4, we will (1) block the activation of JNK, p38, and p90RSK by specific inhibitors, dominant negative mutants, siRNA, or gene knockout cell lines; (2) use cell lines expressing siRNA of histone H3, dominant negative mutants (histone H3 S10 mutated to A10, H3 S28 mutated to A28) and histone H3 wild-type to study the role of histone H3 in arsenic-induced cell transformation. Such knowledge will facilitate the design of more effective and specific strategies with fewer side effects for chemoprevention of arsenic-induced cancer.