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Transcriptional up-regulation of gamma-glutamylcysteine synthetase gene expression in melphalan-resistant human prostate carcinoma cells.
Mol Pharmacol. 1994 Nov; 46(5):909-14.MP

Abstract

Tumor cell resistance to many chemotherapeutic agents, including alkylating agents, cisplatin, and doxorubicin, is frequently associated with increased intracellular levels of the nonprotein sulfhydryl glutathione (GSH). Recent evidence has demonstrated that increased GSH levels can be accompanied by an increase in the activity of gamma-glutamylcysteine synthetase (GCS), which catalyzes the rate-limiting step in de novo synthesis of GSH, and by an increase in the steady state level of mRNA for the catalytic subunit of GCS. Using melphalan-resistant DU 145/M4.5 human prostate carcinoma cells, which express elevated GSH levels, GCS enzyme activity, and GCS mRNA levels, we sought to determine the mechanism(s) responsible for the increased GCS mRNA expression. As determined by Northern analyses and RNase protection assays, the steady state level of GCS message in the resistant cells was increased 10-20-fold, in comparison with the drug-sensitive parent DU 145 cells. No significant difference in gene copy number or evidence of rearrangement was detected in the resistant cell line by Southern analyses. The GCS-specific mRNA isolated from the resistant cells was less stable than that isolated from the drug-sensitive cells (half-lives of 6 hr and 9 hr, respectively), indicating that this difference does not contribute to the increased steady state levels in the resistant cells. Nuclear run-on experiments revealed that the GCS transcription rate in the DU 145/M4.5 cells was increased approximately 12-fold, in comparison with that detected in the DU 145 cells. This difference in transcription rate was comparable in magnitude to the difference in steady state mRNA levels detectable in the two cell populations. Similar correlations between steady state GCS mRNA levels and transcription rates were also observed in other DU 145 lines expressing intermediate degrees of resistance to melphalan and correspondingly intermediate GCS mRNA elevations. These data suggest that GCS expression is transcriptionally regulated in these melphalan-resistant tumor cells.

Authors+Show Affiliations

Department of Human Oncology, University of Wisconsin School of Medicine, Madison 53792.No affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, U.S. Gov't, P.H.S.

Language

eng

PubMed ID

7969079

Citation

Mulcahy, R T., et al. "Transcriptional Up-regulation of Gamma-glutamylcysteine Synthetase Gene Expression in Melphalan-resistant Human Prostate Carcinoma Cells." Molecular Pharmacology, vol. 46, no. 5, 1994, pp. 909-14.
Mulcahy RT, Untawale S, Gipp JJ. Transcriptional up-regulation of gamma-glutamylcysteine synthetase gene expression in melphalan-resistant human prostate carcinoma cells. Mol Pharmacol. 1994;46(5):909-14.
Mulcahy, R. T., Untawale, S., & Gipp, J. J. (1994). Transcriptional up-regulation of gamma-glutamylcysteine synthetase gene expression in melphalan-resistant human prostate carcinoma cells. Molecular Pharmacology, 46(5), 909-14.
Mulcahy RT, Untawale S, Gipp JJ. Transcriptional Up-regulation of Gamma-glutamylcysteine Synthetase Gene Expression in Melphalan-resistant Human Prostate Carcinoma Cells. Mol Pharmacol. 1994;46(5):909-14. PubMed PMID: 7969079.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Transcriptional up-regulation of gamma-glutamylcysteine synthetase gene expression in melphalan-resistant human prostate carcinoma cells. AU - Mulcahy,R T, AU - Untawale,S, AU - Gipp,J J, PY - 1994/11/1/pubmed PY - 1994/11/1/medline PY - 1994/11/1/entrez SP - 909 EP - 14 JF - Molecular pharmacology JO - Mol. Pharmacol. VL - 46 IS - 5 N2 - Tumor cell resistance to many chemotherapeutic agents, including alkylating agents, cisplatin, and doxorubicin, is frequently associated with increased intracellular levels of the nonprotein sulfhydryl glutathione (GSH). Recent evidence has demonstrated that increased GSH levels can be accompanied by an increase in the activity of gamma-glutamylcysteine synthetase (GCS), which catalyzes the rate-limiting step in de novo synthesis of GSH, and by an increase in the steady state level of mRNA for the catalytic subunit of GCS. Using melphalan-resistant DU 145/M4.5 human prostate carcinoma cells, which express elevated GSH levels, GCS enzyme activity, and GCS mRNA levels, we sought to determine the mechanism(s) responsible for the increased GCS mRNA expression. As determined by Northern analyses and RNase protection assays, the steady state level of GCS message in the resistant cells was increased 10-20-fold, in comparison with the drug-sensitive parent DU 145 cells. No significant difference in gene copy number or evidence of rearrangement was detected in the resistant cell line by Southern analyses. The GCS-specific mRNA isolated from the resistant cells was less stable than that isolated from the drug-sensitive cells (half-lives of 6 hr and 9 hr, respectively), indicating that this difference does not contribute to the increased steady state levels in the resistant cells. Nuclear run-on experiments revealed that the GCS transcription rate in the DU 145/M4.5 cells was increased approximately 12-fold, in comparison with that detected in the DU 145 cells. This difference in transcription rate was comparable in magnitude to the difference in steady state mRNA levels detectable in the two cell populations. Similar correlations between steady state GCS mRNA levels and transcription rates were also observed in other DU 145 lines expressing intermediate degrees of resistance to melphalan and correspondingly intermediate GCS mRNA elevations. These data suggest that GCS expression is transcriptionally regulated in these melphalan-resistant tumor cells. SN - 0026-895X UR - https://www.unboundmedicine.com/medline/citation/7969079/Transcriptional_up_regulation_of_gamma_glutamylcysteine_synthetase_gene_expression_in_melphalan_resistant_human_prostate_carcinoma_cells_ L2 - http://molpharm.aspetjournals.org/cgi/pmidlookup?view=long&pmid=7969079 DB - PRIME DP - Unbound Medicine ER -