RNA interference of argininosuccinate synthetase restores sensitivity to recombinant arginine deiminase (rADI) in resistant cancer cells
Journal
Journal of Biomedical Science
Journal Volume
18
Journal Issue
1
Date Issued
2011
Author(s)
Abstract
Background: Sensitivity of cancer cells to recombinant arginine deiminase (rADI) depends on expression of argininosuccinate synthetase (AS), a rate-limiting enzyme in synthesis of arginine from citrulline. To understand the efficiency of RNA interfering of AS in sensitizing the resistant cancer cells to rADI, the down regulation of AS transiently and permanently were performed in vitro, respectively. Methods. We studied the use of down-regulation of this enzyme by RNA interference in three human cancer cell lines (A375, HeLa, and MCF-7) as a way to restore sensitivity to rADI in resistant cells. The expression of AS at levels of mRNA and protein was determined to understand the effect of RNA interference. Cell viability, cell cycle, and possible mechanism of the restore sensitivity of AS RNA interference in rADI treated cancer cells were evaluated. Results: AS DNA was present in all cancer cell lines studied, however, the expression of this enzyme at the mRNA and protein level was different. In two rADI-resistant cell lines, one with endogenous AS expression (MCF-7 cells) and one with induced AS expression (HeLa cells), AS small interference RNA (siRNA) inhibited 37-46% of the expression of AS in MCF-7 cells. ASsiRNA did not affect cell viability in MCF-7 which may be due to the certain amount of residual AS protein. In contrast, ASsiRNA down-regulated almost all AS expression in HeLa cells and caused cell death after rADI treatment. Permanently down-regulated AS expression by short hairpin RNA (shRNA) made MCF-7 cells become sensitive to rADI via the inhibition of 4E-BP1-regulated mTOR signaling pathway. Conclusions: Our results demonstrate that rADI-resistance can be altered via AS RNA interference. Although transient enzyme down-regulation (siRNA) did not affect cell viability in MCF-7 cells, permanent down-regulation (shRNA) overcame the problem of rADI-resistance due to the more efficiency in AS silencing. ? 2011 Wu et al; licensee BioMed Central Ltd.
Subjects
arginine deiminase; argininosuccinate synthetase; resistance; RNA interference
SDGs
Other Subjects
arginine deiminase; argininosuccinate synthase; DNA; initiation factor 4E binding protein 1; mammalian target of rapamycin; messenger RNA; recombinant arginine deiminase; recombinant enzyme; short hairpin RNA; small interfering RNA; unclassified drug; antineoplastic agent; arginine; arginine deiminase; argininosuccinate synthase; citrulline; hydrolase; recombinant protein; small interfering RNA; article; cancer cell; cancer cell culture; cell cycle; cell death; cell strain MCF 7; cell viability; down regulation; drug mechanism; drug sensitivity; gene expression; HeLa cell; human; human cell; priority journal; protein expression; RNA interference; sensitization; cell survival; drug resistance; enzymology; genetics; metabolism; neoplasm; tumor cell line; Antineoplastic Agents; Arginine; Argininosuccinate Synthase; Cell Line, Tumor; Cell Survival; Citrulline; Drug Resistance, Neoplasm; Gene Expression; Hela Cells; Humans; Hydrolases; Neoplasms; Recombinant Proteins; RNA Interference; RNA, Small Interfering
Type
journal article