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首页> 外文学位 >Somatic cell and molecular genetic analysis of various Chinese hamster ovary (CHO) mutant cells defective in sterol-dependent regulation of cholesterol biosynthesis and LDL receptor expression.
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Somatic cell and molecular genetic analysis of various Chinese hamster ovary (CHO) mutant cells defective in sterol-dependent regulation of cholesterol biosynthesis and LDL receptor expression.

机译：各种中国仓鼠卵巢（CHO）突变细胞的体细胞和分子遗传学分析，它们在胆固醇依赖的胆固醇生物合成和LDL受体表达调控中存在缺陷。

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摘要

Wild-type or normal Chinese hamster ovary cells show coordinate regulation of cholesterol biosynthesis and LDL receptor expression in response to sterol in culture medium. In order to identify genes which play an important role in sterol-mediated regulation of of cholesterol metabolism, we selected mutant cell clones which were abnormal in regulating cholesterol metabolism. So far, we have isolated a total of three novel mutants, each one due to a single gene defect. The major focus of my thesis research has been the cloning of these genes.;Using the amphotericin B enrichment procedure, we have isolated several non-leaky mutant CHO cell clones (M4, M19 and M21) which belong to the same genetic complementation group as the mutant M1 cells previously reported from this laboratory (Limanek, J. S., Chin, J., and Chang, T. Y. 1978, PNAS 75:452-456). All of the 4 mutant cell clones were defective in sterol-dependent coordinate regulation of cholesterol biosynthesis and LDL receptor expression. In addition, we have isolated a novel type of 25-hydroxycholesterol resistant mutant CHO cell clone, designated as W1 cell, using M1, M4, M19, or M21 cells as parental cells. A different mutant cell clone was isolated in this laboratory, mutant 25RA cell, which was shown to be resistant to 25-hydroxycholesterol (Chang, T. Y., and Limanek, J. S. 1980 JBC 225:7787-7795). 25RA and W1 cells grow in medium without cholesterol. Unlike wild-type CHO cells, 25RA and W1 cells have elevated level of cholesterol biosynthesis rate and LDL receptor expression when they are grown in medium containing cholesterol. We have found that the mutations in W1 cell and 25RA cell are different. We also found that the M1 and 25RA genes are different, and that a functional M1 gene is required for expression of the 25RA cell phenotype.;When total human genomic DNA was transfected into mutant M19 cells, it gave transfectant cell clones with wild-type cell phenotype. On the other hand, M19 cells acquired the W1 cell phenotype when they were transfected with total W1 genomic DNA. Using gene transfer experiments, we have identified and cloned unique DNA fragments which are closely linked to the human M1 and the W1 genes. Presently, we are using these linked DNA fragments to do a chromosome walk.
机译：野生型或正常中国仓鼠卵巢细胞在培养基中响应甾醇对胆固醇的生物合成和LDL受体表达具有协调性调节作用。为了鉴定在胆固醇介导的胆固醇代谢调节中起重要作用的基因，我们选择了在调节胆固醇代谢中异常的突变细胞克隆。到目前为止，我们已经分离出三个新的突变体，每个突变体都是由于单个基因缺陷造成的。本文的主要研究重点是克隆这些基因。通过两性霉素B富集程序，我们分离了几个非泄漏的CHO细胞克隆（M4，M19和M21），它们与该突变体M1细胞先前是从该实验室报道的（Limanek，JS，Chin，J。和Chang，TY 1978，PNAS 75：452-456）。所有这4个突变细胞克隆在胆固醇依赖的胆固醇生物合成和LDL受体表达的协调调控方面均存在缺陷。此外，我们使用M1，M4，M19或M21细胞作为亲代细胞，分离出了一种新型的25-羟基胆固醇抗性突变CHO细胞克隆，称为W1细胞。在该实验室中分离出不同的突变细胞克隆，即突变的25RA细胞，其被证明对25-羟基胆固醇具有抗性（Chang，T.Y。和Limanek，J.S.1980 JBC 225：7787-7795）。 25RA和W1细胞在不含胆固醇的培养基中生长。与野生型CHO细胞不同，当25RA和W1细胞在含有胆固醇的培养基中生长时，它们的胆固醇生物合成速率和LDL受体表达水平升高。我们发现W1细胞和25RA细胞中的突变是不同的。我们还发现M1和25RA基因不同，并且表达25RA细胞表型需要功能性M1基因。当将人类全基因组DNA转染到突变M19细胞中时，得到了具有野生型的转染细胞克隆细胞表型。另一方面，当用总的W1基因组DNA转染M19细胞时，它们获得了W1细胞表型。使用基因转移实验，我们已经鉴定并克隆了与人类M1和W1基因紧密相连的独特DNA片段。目前，我们正在使用这些链接的DNA片段进行染色体漫步。

著录项

  • 作者

    Hasan, Mazahir Tahir.;

  • 作者单位

    Dartmouth College.;

  • 授予单位 Dartmouth College.;
  • 学科 Biology Molecular.;Chemistry Biochemistry.;Biology Genetics.
  • 学位 Ph.D.
  • 年度 1993
  • 页码 172 p.
  • 总页数 172
  • 原文格式 PDF
  • 正文语种 eng
  • 中图分类
  • 关键词

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