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Studies of folate and branched-chain...

Wu, Keqiang.

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Studies of folate and branched-chain amino acid biosynthesis in plants.

紀錄類型:	書目-電子資源 : Monograph/item
正題名/作者:	Studies of folate and branched-chain amino acid biosynthesis in plants./
作者:	Wu, Keqiang.
出版者:	Ann Arbor : ProQuest Dissertations & Theses, : 1994,
面頁冊數:	297 p.
附註:	Source: Dissertations Abstracts International, Volume: 76-02, Section: B.
Contained By:	Dissertations Abstracts International76-02B.
標題:	Botany. -
電子資源:	https://pqdd.sinica.edu.tw/twdaoapp/servlet/advanced?query=NN00211
ISBN:	9780612002111

Studies of folate and branched-chain amino acid biosynthesis in plants.
Wu, Keqiang.

Studies of folate and branched-chain amino acid biosynthesis in plants. - Ann Arbor : ProQuest Dissertations & Theses, 1994 - 297 p.

Source: Dissertations Abstracts International, Volume: 76-02, Section: B.

Thesis (Ph.D.)--The University of Saskatchewan (Canada), 1994.

This item must not be sold to any third party vendors.

The overall objective of this research was to investigate the value of using resistant mutants and variants to study the nature and control of folate and branched-chain amino acid biosynthetic pathways. A wild-type Datura innoxia cell line (Px4) was used to select for methotrexate-resistant cell variants through a stepwise procedure. Two independently selected cell lines, MTX161 and MTX132, were stable and shown to be 5- to 15-times more resistant to methotrexate than wild type. These methotrexate-resistant cells were similar to the wild-type cells in levels and kinetic properties of dihydrofolate reductase, the sensitivity of dihydrofolate reductase to methotrexate, and the binding of ($\\sp3$H) methotrexate to soluble proteins. However, greatly reduced uptake of methotrexate in the resistant cell lines was detected compared with the wild type. High performance liquid chromatographic analyses indicated that methotrexate could be converted into polyglutamylated forms in Datura cells. Decreased MTX polyglutamylation in MTX-resistant cells was found compared with the wild type. In vivo analyses of folylpolyglutamate pools of the wild-type and methotrexate-resistant Datura cell lines indicated that hexaglutamates were the predominant form in these cell lines. Although in vivo and in vitro folylpolyglutamate synthesis was found to be similar in these Datura cell lines, about a 4-fold increase in specific activity of gamma-glutamyl hydrolase was detected in the MTX161 cell line. The increase in this enzyme in one of the methotrexate-resistant cell lines suggested that breakdown of polyglutamylated forms of methotrexate may play a role in acquired methotrexate resistance. Two independently isolated FUdR-resistant mutants of Arabidopsis (FUD-1 and FUD-2) were identified from ethylmethane sulfonate mutagenized seeds. The resistance was found to be due to single recessive nuclear mutations. Genetic complementation tests indicated that these mutations were in the same gene locus, which was designated fur1 and mapped to linkage group 4 of Arabidopsis. Biochemical studies demonstrated that the FUdR resistance in these two mutants was due to decreased drug uptake. A valine-resistant mutant line of Arabidopsis thaliana, VAL-2, was identified by screening M2 populations of ethylmethane sulfonate-mutagenized seeds. The resistance was due to a single, dominant, nuclear gene mutation. Assay of acetolactate synthase indicated that the valine resistance in this mutant was caused by decreased sensitivity of acetolactate synthase to valine. The mutant gene, var1, maps, or is very closely linked, to CSR1, the gene encoding acetolactate synthase in Arabidopsis.

ISBN: 9780612002111Subjects--Topical Terms:

516217
Botany.
Subjects--Index Terms:

Arabidopsis thaliana

Studies of folate and branched-chain amino acid biosynthesis in plants.
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520 $a The overall objective of this research was to investigate the value of using resistant mutants and variants to study the nature and control of folate and branched-chain amino acid biosynthetic pathways. A wild-type Datura innoxia cell line (Px4) was used to select for methotrexate-resistant cell variants through a stepwise procedure. Two independently selected cell lines, MTX161 and MTX132, were stable and shown to be 5- to 15-times more resistant to methotrexate than wild type. These methotrexate-resistant cells were similar to the wild-type cells in levels and kinetic properties of dihydrofolate reductase, the sensitivity of dihydrofolate reductase to methotrexate, and the binding of ($\\sp3$H) methotrexate to soluble proteins. However, greatly reduced uptake of methotrexate in the resistant cell lines was detected compared with the wild type. High performance liquid chromatographic analyses indicated that methotrexate could be converted into polyglutamylated forms in Datura cells. Decreased MTX polyglutamylation in MTX-resistant cells was found compared with the wild type. In vivo analyses of folylpolyglutamate pools of the wild-type and methotrexate-resistant Datura cell lines indicated that hexaglutamates were the predominant form in these cell lines. Although in vivo and in vitro folylpolyglutamate synthesis was found to be similar in these Datura cell lines, about a 4-fold increase in specific activity of gamma-glutamyl hydrolase was detected in the MTX161 cell line. The increase in this enzyme in one of the methotrexate-resistant cell lines suggested that breakdown of polyglutamylated forms of methotrexate may play a role in acquired methotrexate resistance. Two independently isolated FUdR-resistant mutants of Arabidopsis (FUD-1 and FUD-2) were identified from ethylmethane sulfonate mutagenized seeds. The resistance was found to be due to single recessive nuclear mutations. Genetic complementation tests indicated that these mutations were in the same gene locus, which was designated fur1 and mapped to linkage group 4 of Arabidopsis. Biochemical studies demonstrated that the FUdR resistance in these two mutants was due to decreased drug uptake. A valine-resistant mutant line of Arabidopsis thaliana, VAL-2, was identified by screening M2 populations of ethylmethane sulfonate-mutagenized seeds. The resistance was due to a single, dominant, nuclear gene mutation. Assay of acetolactate synthase indicated that the valine resistance in this mutant was caused by decreased sensitivity of acetolactate synthase to valine. The mutant gene, var1, maps, or is very closely linked, to CSR1, the gene encoding acetolactate synthase in Arabidopsis.
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