Genomic Instability and Evolution of Drug Resistance

基因组不稳定性和耐药性的演变

• 批准号: 7201643
• 负责人: MACUS T KUO
• 金额: $ 29.35万
• 依托单位: UNIVERSITY OF TX MD ANDERSON CAN CTR
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-04-01 至 2009-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7201643
• 关键词: 1q31; Affect; Amino Acids; Animals; Bladder; Cell Aging; Cell Death; Cell Nucleus; Cells; Chromosome Breakage; Chromosome Fragile Sites; Chromosome abnormality; Colon; Complementary DNA; Cultured Cells; Cytoplasm; DNA; DNA amplification; Development; Differentiation and Growth; Drug resistance; Epithelial; Epithelial Cells; Epithelium; Event; Evolution; Gene Amplification; Genes; Genomic Instability; Genus Cola; Human; Knock-out; Learning; Length; Localized; Malignant - descriptor; Malignant Epithelial Cell; Mammalian Cell; Mammary gland; Messenger RNA; Mitotic; Molecular; Mouse Strains; Multi-Drug Resistance; Neoplastic Cell Transformation; Open Reading Frames; Organ; Ovary; Pattern; Physiology; Play; Process; Prostate; Proteins; Recombinants; Role; Sequence Analysis; Source; Staging; Structure; System; Testing; Tissues; Transfection; Variant; cancer cell; cell growth; cell growth regulation; cell type; chemotherapy; drug sensitivity; embryonic stem cell; expression vector; gene function; insight; novel; programs; senescence

DESCRIPTION (provided by applicant): DNA amplification is frequently observed in cancer cells and in drug-resistant cells. One important mechanism involved in DNA amplification is the breakage-fusion-bridge (BFB) process. We previously demonstrated that chromosomal fragile site breakage triggers the BFB cycle. In many CHO multidrug-resistant cells, amplification of mdr1 is associated with the breakage of chromosomal fragile site 1q31. To learn the molecular mechanism underlying the initial events of mdrl amplification, we have cloned 1q31 fragile site DNA. Strikingly, we found that this fragile site contains a novel gene, designated as fragile site associated (fsa) gene, Fas encodes a mRNA of greater than14 kb. Full-length human FSA cDNA has been cloned. Sequence analyses revealed that its mRNA is bicistronic and contains two evolutionarily conserved nonoverlapping open reading frames (orf1 and orf2). While basal levels of FSA mRNA seem to express in many cell types, immunohistochemical analyses revealed a co-expression pattern of the two off-encoded proteins in the post-mitotic, well-differentiated epithelial compartments of many organs; including colon, mammary glands, ovary, prostate and bladder. These findings suggest that FSA plays important roles in, regulating mammalian epithelial growth and differentiation. Moreover, in normal mature epithelia, the FSA protein seems to be localized in nuclei, whereas in the malignant cells of epithelial origins, FSA seems to be mainly cytoplasmic. These results suggest a nucleus-cytoplasm shuffling during epithelial transformation. We hypothesize that elevated FSA expression may induce premature senescence and affect drug sensitivities to chemotherapy of epithelial cells. We propose three specific aims to further elucidate the structure/function of this gene. In Aim I, we propose to critically re-investigate the intracellular localizations of FSA-orf1 and FSA-orf2, and their expression in normal and malignant epithelial cells from many tissue sources. In Aim II, we propose to investigate the function of FSA-orf1 and FSA-ort2 in cultured cells by transfection and to test the above-mentioned hypothesis. And in Aim III, we propose to investigate the function of FSA-orf1 and FSA-orf2 using the knock out strategies. We anticipate from these studies to gain important insights into the function of FSA in the regulation of cell growth and differentiation in normal and malignant epithelial cells.

描述（由申请人提供）： DNA 扩增经常在癌细胞和耐药细胞中观察到。 DNA 扩增涉及的一项重要机制是断裂-融合-桥 (BFB) 过程。我们之前证明染色体脆弱位点断裂会触发 BFB 循环。在许多 CHO 多重耐药细胞中，mdr1 的扩增与染色体脆弱位点 1q31 的断裂有关。为了了解 mdrl 扩增初始事件背后的分子机制，我们克隆了 1q31 脆弱位点 DNA。引人注目的是，我们发现这个脆弱位点包含一个新的基因，称为脆弱位点相关（fsa）基因，Fas编码大于14 kb的mRNA。全长人类 FSA cDNA 已被克隆。序列分析显示其 mRNA 是双顺反子，并包含两个进化保守的非重叠开放阅读框（orf1 和 orf2）。虽然 FSA mRNA 的基础水平似乎在许多细胞类型中表达，但免疫组织化学分析揭示了这两种脱编码蛋白在许多器官的有丝分裂后、分化良好的上皮区室中的共表达模式；包括结肠、乳腺、卵巢、前列腺和膀胱。这些发现表明 FSA 在调节哺乳动物上皮生长和分化中发挥重要作用。此外，在正常成熟上皮细胞中，FSA蛋白似乎位于细胞核中，而在上皮来源的恶性细胞中，FSA似乎主要位于细胞质中。这些结果表明上皮转化过程中存在核-细胞质改组。我们假设 FSA 表达升高可能会诱导上皮细胞过早衰老并影响对化疗的药物敏感性。我们提出了三个具体目标来进一步阐明该基因的结构/功能。在目标 I 中，我们建议批判性地重新研究 FSA-orf1 和 FSA-orf2 的细胞内定位，以及它们在来自许多组织来源的正常和恶性上皮细胞中的表达。在Aim II中，我们建议通过转染研究FSA-orf1和FSA-ort2在培养细胞中的功能并检验上述假设。在目标 III 中，我们建议使用敲除策略研究 FSA-orf1 和 FSA-orf2 的功能。我们期望从这些研究中获得关于 FSA 在调节正常和恶性上皮细胞的细胞生长和分化中的功能的重要见解。