STRUCTURE AND FUNCTION OF ONCOGENES AND ANTI-ONCOGENES

癌基因和抗癌基因的结构和功能

• 批准号: 2468451
• 负责人: J F MUSHINSKI
• 金额: --
• 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/2468451
• 关键词: B lymphocyte; Retroviridae; antibody formation; antitumor antibody; apoptosis; cell cell interaction; cell differentiation; chimeric proteins; chromosome translocation; cyclins; gene expression; helper T lymphocyte; human tissue; immunoglobulin genes; interleukin 6; laboratory mouse; molecular cloning; monoclonal antibody; neoplasm /cancer genetics; neoplastic transformation; oncogenes; plasma cell neoplasm; protein kinase C; tissue /cell culture

Our research goal is to understand the molecular structure and function of the genes that play critical roles in normal growth and differentiation, neoplastic transformation, and apoptosis in mouse and human tissues and tumors of the hematopoietic system. We study oncogenes, c-myc, v-raf and v-abl; anti-oncogenes, esp. the bcl-2 family; cell cycle-regulating proteins (cyclins) and their inhibitors, p21 (waf) and p16; as well as molecules that transduce signals within the cell, e.g., protein kinase C (PKC). We and others have shown that the deregulated expression of c-myc secondary to chromosomal translocations in the c-myc region is an essential element in the series of genetic alterations that are involved in plasmacytomagenesis in BALB/c mice and in Burkitt and AIDS-associated lymphomas in man. We have also shown that c-myc can also be dysregulated in these tumor cells by retroviral insertion of strong enhancers in myc's upstream flank. It is not known why BALB/c mice are particularly susceptible to these genetic insults, but we have a candidate mechanism. We have found that the BALB/c mouse has an unusual defect in a special form of excision repair of DNA damage. This form of excision repair is unusual in that it is not coupled to RNA transcription, which is usual for DNA excision repair. Furthermore, the defect is only manifest in repair of DNA damage in the c-myc, Pvt1, switch Ig alpha and Ig kappa genes, namely the sites of recurrent chromosomal translocation in B-lymphocytic neoplasms. This is a plausible mechanism for the production of the gene-specific, strain-specific genomic instability that predisposes to the chromosome translocations that lead to constitutive expression of c-myc. This overexpression of c-myc, in turn, leads to an extension of gene-specific genetic instability to a new subset of genes, including cyclin D2. This gene becomes amplified and overexpressed in the face of c-myc overexpression, contributing to cell proliferation. We have produced a recombinant retrovirus that expresses v-abl and c-myc (ABL-MYC). This virus rapidly induces plasmacytomas in vitro and in vivo in BALB/c, nude and other strains of mice. This technology has been used to produce monoclonal antibodies to parasites, particulate, protein and peptide antigens, and it offers an alternative to hybridoma technology. We have shown that this combination of oncogenes is unique in that it permits differentiation of B cells into plasma cells in the absence of T-cell help and without ip pristane. Because T cells are not necessary in the induction of plasmacytomas with this retrovirus, we were able to show that normal T cells actually retard the emergence of these tumors. We have cloned eight PKC isozymes into expression vectors and produced cell lines that overexpress each of these isoforms in a variety of hematopoietic cell lines. Plasmacytomas are usually IL-6-dependent in vivo and in vitro, and withdrawal of IL-6 leads to their death by apoptosis. This process of apoptosis can be delayed by activating PKC-d with a specific phorbol ester. PKC-delta is also able to mediate macrophage differentiation in promyelocytes and cytoskeleton- mediated changes in the shape of B lymphocytes. We wished to determine which portion of the PKC-delta protein determines its isozyme-specific functions by constructing expression vectors that overexpress chimeric molecules that are half PKC-delta and half PKC-epsilon. The overexpression of these chimeras in different cell types showed that the C-terminal half, containing the catalytic domain, appears to bear most of the isoform-specific determinants of myeloid differentiation and transformation.

我们的研究目标是了解分子结构和功能 在正常生长中起关键作用的基因 小鼠和小鼠的分化、肿瘤转化和细胞凋亡 人体组织和造血系统肿瘤。我们学习 癌基因、c-myc、v-raf 和 v-abl；抗癌基因，特别是。 bcl-2 家庭;细胞周期调节蛋白（细胞周期蛋白）及其抑制剂， p21 (waf) 和 p16；以及在其中转导信号的分子 细胞，例如蛋白激酶 C (PKC)。我们和其他人已经证明 继发于染色体的 c-myc 表达失调 c-myc 区域的易位是 与浆细胞瘤发生有关的一系列遗传改变 在 BALB/c 小鼠以及人类伯基特和艾滋病相关淋巴瘤中。我们 还表明 c-myc 在这些肿瘤中也可能失调 通过逆转录病毒在 myc 上游插入强增强子来增强细胞 侧翼。 目前尚不清楚为什么 BALB/c 小鼠特别容易受到这些影响 基因侮辱，但我们有一个候选机制。我们发现 BALB/c 小鼠在特殊形式的切除中存在不寻常的缺陷 DNA损伤的修复。这种形式的切除修复很不寻常，因为 它不与 RNA 转录偶联，而 RNA 转录通常用于 DNA 切除 维修。此外，缺陷仅表现在DNA修复上 c-myc、Pvt1、开关 Ig α 和 Ig kappa 基因受损，即 B淋巴细胞中反复染色体易位的位点 肿瘤。这是一种合理的生产机制 基因特异性、品系特异性的基因组不稳定性容易导致 染色体易位导致组成型表达 c-myc。 c-myc 的这种过度表达反过来导致 新基因子集的基因特异性遗传不稳定性，包括 细胞周期蛋白D2。该基因在面部被放大并过度表达 c-myc 过度表达，有助于细胞增殖。 我们已经生产了表达 v-abl 和 c-myc (ABL-MYC)。该病毒在体外迅速诱导浆细胞瘤 BALB/c、裸鼠和其他品系小鼠的体内实验。这项技术有 用于生产针对寄生虫、颗粒物的单克隆抗体， 蛋白质和肽抗原，它提供了杂交瘤的替代方案 技术。我们已经证明这种癌基因的组合是独特的 因为它允许 B 细胞分化为浆细胞 缺乏 T 细胞的帮助并且没有 ip pristane。因为T细胞不是 用这种逆转录病毒诱导浆细胞瘤是必要的，我们 我们能够证明正常 T 细胞实际上延缓了 这些肿瘤。 我们已将 8 个 PKC 同工酶克隆到表达载体中并产生 在多种细胞中过表达这些亚型的细胞系 造血细胞系。浆细胞瘤通常具有 IL-6 依赖性 体内和体外，IL-6 的撤除会导致它们死亡 细胞凋亡。激活 PKC-d 可以延迟细胞凋亡的过程 与特定的佛波酯。 PKC-delta 也能够调解 早幼粒细胞中的巨噬细胞分化和细胞骨架介导 B淋巴细胞的形状发生变化。我们希望确定哪一个 PKC-delta 蛋白的部分决定了其同工酶特异性 通过构建过表达嵌合体的表达载体来发挥功能 一半 PKC-δ 和一半 PKC-ε 的分子。这 这些嵌合体在不同细胞类型中的过度表达表明 C 末端一半包含催化结构域，似乎带有 骨髓分化的大多数异构体特异性决定因素 和转型。