Gene Expression and Signal Transduction in Transformation and Differentiation

转化和分化中的基因表达和信号转导

• 批准号: 7592581
• 负责人: J F MUSHINSKI
• 金额: $ 115.2万
• 依托单位: DIVISION OF BASIC SCIENCES - NCI
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/7592581
• 关键词: Actins; Adherence; Affect; Alien; Amino Acids; Animals; B-Cell Lymphomas; Binding; Biological Process; Burkitt Lymphoma; Cancer Patient; Catalytic Domain; Cell Proliferation; Cell Shape; Cells; Clinical; Cluster Analysis; Cytoskeleton; Data; Development; Elements; Environment; Event; Experimental Models; Experimental Neoplasms; Family; Gene Expression; Generations; Genes; Genomic Instability; Goals; Growth; Human; In Vitro; Inbred BALB C Mice; Isoenzymes; Learning; Lesion; Lymphocyte; Malignant - descriptor; Malignant Neoplasms; Measures; Mediating; Membrane; MicroRNAs; Molecular; Molecular Genetics; Molecular Probes; Molecular Profiling; Multiple Myeloma; Mus; Mutation; NIH 3T3 Cells; Nature; Neoplasm Metastasis; Neoplastic Cell Transformation; Neoplastic Processes; Non-Hodgkin' s Lymphoma; Nuclear; Oils; Oncogenes; Patients; Pattern; Phorbol Esters; Phosphorylation; Phosphotransferases; Physiological; Plasma Cell Neoplasm; Plasma Cells; Plasmacytoma; Premalignant; Preventive; Protein Isoforms; Protein Kinase C; Protein Overexpression; Protein-Serine-Threonine Kinases; Proteins; Publishing; Research; Role; Signal Transduction; Structure; Surface; System; Threonine; Transgenes; Tumor Promoters; Tumor Suppressor Genes; Tyrosine; Tyrosine Phosphorylation; Up-Regulation; Viral Oncogene; Work; base; c-myc Genes; c-myc Proto-Oncogenes; cell growth; cell motility; cell transformation; comparative genomic hybridization; in vivo; malignant breast neoplasm; man; neoplastic cell; novel; outcome forecast; paxillin; protein kinase C epsilon; protein kinase C gamma; protein kinase C-delta; tissue culture; tumor; tumor progression; Actins; Adherence; Affect; Alien; Amino Acids; Animals; B-Cell Lymphomas; Binding; Biological Process; Burkitt Lymphoma; Cancer Patient; Catalytic Domain; Cell Proliferation; Cell Shape; Cells; Clinical; Cluster Analysis; Cytoskeleton; Data; Development; Elements; Environment; Event; Experimental Models; Experimental Neoplasms; Family; Gene Expression; Generations; Genes; Genomic Instability; Goals; Growth; Human; In Vitro; Inbred BALB C Mice; Isoenzymes; Learning; Lesion; Lymphocyte; Malignant - descriptor; Malignant Neoplasms; Measures; Mediating; Membrane; MicroRNAs; Molecular; Molecular Genetics; Molecular Probes; Molecular Profiling; Multiple Myeloma; Mus; Mutation; NIH 3T3 Cells; Nature; Neoplasm Metastasis; Neoplastic Cell Transformation; Neoplastic Processes; Non-Hodgkin' s Lymphoma; Nuclear; Oils; Oncogenes; Patients; Pattern; Phorbol Esters; Phosphorylation; Phosphotransferases; Physiological; Plasma Cell Neoplasm; Plasma Cells; Plasmacytoma; Premalignant; Preventive; Protein Isoforms; Protein Kinase C; Protein Overexpression; Protein-Serine-Threonine Kinases; Proteins; Publishing; Research; Role; Signal Transduction; Structure; Surface; System; Threonine; Transgenes; Tumor Promoters; Tumor Suppressor Genes; Tyrosine; Tyrosine Phosphorylation; Up-Regulation; Viral Oncogene; Work; base; c-myc Genes; c-myc Proto-Oncogenes; cell growth; cell motility; cell transformation; comparative genomic hybridization; in vivo; malignant breast neoplasm; man; neoplastic cell; novel; outcome forecast; paxillin; protein kinase C epsilon; protein kinase C gamma; protein kinase C-delta; tissue culture; tumor; tumor progression

The chief objective of our research is to understand the molecular and genetic mechanisms responsible for differentiation, cell growth, and neoplastic transformation. We study the oncogenes, tumor-suppressor genes and signal-transducing proteins in mouse and human experimental tumor systems, including BALB/c mouse plasmacytomas, B-cell lymphomas, and NIH 3T3 cells, among others. These are valuable experimental models, because they can be used to devise more specific therapy and preventive measures for human multiple myeloma, non-Hodgkin's lymphomas, and other human malignancies. BALB/c plasmacytomas, like human Burkitt lymphomas, are characterized by constitutive expression of the proto-oncogene, c-Myc. To determine which additional genetic alterations are required for complete transformation, we are using microarray hybridization studies of global gene expression and array-based Comparative Genomic Hybridization to follow genetic changes and changes in gene expression during progression from pre-malignant to fully malignant plasma cell tumors. We are also using microarray hybridization studies to probe the molecular mechanisms at work in development of plasma cell tumors in mice and the mechanisms whereby certain transgenes and viral oncogenes accelerate this neoplastic process. Global gene expression studies are also underway to determine the physiological changes necessary for these tumors to adapt to growth in tissue culture. It is our hypothesis that such adaptive changes in gene expression that enable tumor cells to grow in the foreign environment of culture vessels might be analogous to those needed for human tumors to grow in alien environments following invasion or metastasis. As an extension of this project, we observed marked differences in gene expression profiles between mouse plasma cell tumors growing in tissue culture and the sme tumors growing in intact animals. We wre able to show that these gene expression differences offered a means of predicting liklihood of survival for patients with several different forms of cancer. Our analysis focused mainly on published data from patients with breast cancer. Our studies that compared global gene expression patterns of B-cell lymphomas in mouse and man with expression patterns of plasma cell neoplasms of murine (plasma cell tumors) and human multiple myeloma showed significant and characteriztic differences not only between B-cell lymphomas and plasma cell neoplasms, but also among different subtypes of human and mouse plasma cell neoplasms. Unsupervised hierarchical cluster analysis of expression patterns of the human and murine plasma cell neoplasms showed a similarity between rapid-appearing mouse plasma cell tumors and MM3 and MM4 multiple myelomas, which had particularly poor clinical prognoses. In the study of signal transduction in differentiation and neoplastic transformation, we are investigating the isoform-specific features of the protein kinase C (PKC) family of serine/threonine kinases. We have been focusing on the PKC delta and epsilon isoenzymes, which have opposing effects on cell proliferation. We have shown that most of the isoenzyme-specific determinants are located in the catalytic half (the carboxyl-terminal domain) of these PKCs by creating reciprocal chimeric cDNAs that encode molecules that are half PKC-delta and half PKC-epsilon. We are further dissecting the structure of the catalytic domain to determine which sub-domains determine PKC isoform- specific functions, focusing on the carboxy-terminal 50 amino acids, the "V5 domain." We are studying how alterations in PKC's V5 region affect its kinase activity and its isoform-specific biological function. We are also studying the nature of PKC's ability to cooperate with neoplastic transformation by the c-Myc proto-oncogene. We have shown that phorbol ester-activation of overexpressed PKC-delta disrupts the actin cytoskeleton in human and mouse lymphocytes, leading to the loss of membrane ruffling, a surface alteration needed for cell movement, and the loss of the typical elongated shape of these cells. We have demonstrated that this effect is due to PKC-mediated changes in phosphorylation of key tyrosine residues in the adaptor molecule, paxillin. Whereas the PKC-mediated effects on loss of tyrosine phosphorylation are indirect, we also have learned that PKC-delta can directly bind paxillin and phosphorylate a specific threonine, leading to homotypic aggregation. We have also shown that Myc and one of the PKC isoforms, PKC-gamma, can cooperate to transform NIH3T3 cells in vitro and in vivo, apparently not requiring intra-nuclear Myc. We are trying to understand the mechanism whereby this is accomplished

我们研究的主要目的是了解负责分化，细胞生长和肿瘤转化的分子和遗传机制。我们研究了小鼠和人类实验性肿瘤系统中的肿瘤基因，肿瘤抑制基因和信号转推蛋白，包括BALB/C小鼠浆细胞瘤，B-细胞淋巴瘤和NIH 3T3细胞等。这些是有价值的实验模型，因为它们可用于为人类多发性骨髓瘤，非霍奇金淋巴瘤和其他人类恶性肿瘤制定更具体的疗法和预防措施。 BALB/C浆细胞瘤，例如人伯基特淋巴瘤，其特征在于原始癌基因c-myc的本构表达。为了确定完全转化需要哪些其他遗传改变，我们使用的是对全球基因表达和基于阵列的比较基因组杂交的微阵列杂交研究，以遵循从遗传变化和从恶性到完全恶性血浆细胞肿瘤进展过程中基因表达的变化。我们还使用微阵列杂交研究来探测小鼠浆细胞肿瘤发育中起作用的分子机制以及某些转基因和病毒性致癌基因加速这种肿瘤过程的机制。还在进行全球基因表达研究，以确定这些肿瘤适应组织培养的生长所需的生理变化。我们的假设是，基因表达的这种适应性变化，使肿瘤细胞能够在培养血管的外国环境中生长可能类似于在入侵或转移后人类肿瘤在外星环境中生长所需的肿瘤细胞。作为该项目的扩展，我们观察到在组织培养中生长的小鼠浆细胞肿瘤与完整动物中生长的中小企业肿瘤之间的基因表达谱图显着差异。我们能够证明这些基因表达差异提供了一种预测几种不同形式癌症患者生存比例的方法。我们的分析主要集中在乳腺癌患者的已发表数据上。我们的研究比较了小鼠和人体中B细胞淋巴瘤的全球基因表达模式，其表达模式具有鼠（浆细胞肿瘤）的血浆细胞肿瘤（血浆细胞肿瘤）和人多发性骨髓瘤的显着和特征性差异，不仅在B-cell淋巴瘤和血浆细胞肿瘤之间，而且在不同的子瘤中，而且在不同的子类细胞型中，还显示了人类和小鼠Neeoplasm和小鼠Neeoplasms。人和鼠血浆细胞肿瘤表达模式的无监督分层聚类分析显示，快速出现的小鼠浆细胞肿瘤与MM3和MM4多发性骨髓瘤之间的相似性，这些骨髓瘤特别差。在分化和肿瘤转化中信号转导的研究中，我们正在研究丝氨酸/苏氨酸激酶的蛋白激酶C（PKC）家族的同工型特异性。我们一直专注于PKC三角洲和Epsilon同工酶，它们对细胞增殖具有相反的作用。我们已经表明，大多数同工酶特异性决定因素位于这些PKC的催化半（羧基末端结构域）中，通过创建编码半pkc-delta和一半pkc-epsilon的相互嵌合cDNA。我们正在进一步剖析催化结构域的结构，以确定哪些子域确定PKC同工型 - 特异性功能，重点是羧基末端50氨基酸，即“ V5结构域”。我们正在研究PKC V5区域的变化如何影响其激酶活性及其同工型特异性生物学功能。我们还研究了PKC与C-Myc原癌基因合作与肿瘤转化的能力的性质。我们已经表明，过表达PKC-DELTA的佛波酯激活破坏了人和小鼠淋巴细胞中肌动蛋白的细胞骨架，导致膜皱纹的丧失，细胞运动所需的表面改变以及这些细胞典型的细胞形状的丧失。我们已经证明了这种作用是由于PKC介导的衔接子分子Paxillin中关键酪氨酸残基磷酸化的变化。尽管PKC介导的对酪氨酸磷酸化损失的影响是间接的，但我们还了解到，PKC-DELTA可以直接结合paxillin并磷酸化特定的苏氨酸，从而导致同型聚集。我们还表明，MYC和PKC同工型PKC-Gamma可以合作以在体外和体内转化NIH3T3细胞，显然不需要核内myc。我们正在尝试了解实现这一目标的机制