Abstract
BackgroundWe investigated mTOR regulation of gene expression by studying rapamycin effect in two hepatic cell lines, the non-tumorigenic WB-F344 cells and the tumorigenic WB311 cells. The latter are resistant to the growth inhibitory effects of rapamycin, thus providing us with an opportunity to study the gene expression effects of rapamycin without confounding effects on cell proliferation.Methodology/Principal FindingsThe hepatic cells were exposed to rapamycin for 24 hr. Microarray analysis on total RNA preparations identified genes that were affected by rapamycin in both cell lines and, therefore, modulated independent of growth arrest. Further studies showed that the promoter regions of these genes included E-box-containing transcription factor binding sites at higher than expected rates. Based on this, we tested the hypothesis that c-Myc is involved in regulation of gene expression by mTOR by comparing genes altered by rapamycin in the hepatic cells and by c-Myc induction in fibroblasts engineered to express c-myc in an inducible manner. Results showed enrichment for c-Myc targets among rapamycin sensitive genes in both hepatic cell lines. However, microarray analyses on wild type and c-myc null fibroblasts showed similar rapamycin effect, with the set of rapamycin-sensitive genes being enriched for c-Myc targets in both cases.Conclusions/SignificanceThere is considerable overlap in the regulation of gene expression by mTOR and c-Myc. However, regulation of gene expression through mTOR is c-Myc-independent and cannot be attributed to the involvement of specific transcription factors regulated by the rapamycin-sensitive mTOR Complex 1.
Highlights
The mammalian Target of Rapamycin is a central regulator of many biological processes that are essential for cell growth, including cell cycle progression, protein translation, ribosomal biogenesis, autophagy and metabolism [1,2]
A cross comparison of expression patterns (Figure 1A) showed that 791 gene features were exclusively altered in WB-F344 cells after rapamycin treatment while 484 gene features were exclusively altered in the WB311 cells
Examination of the 106 gene features (Figure 1B) showed that the expression of 26 of the genes was affected in opposite directions in the WB-F344 and WB311 cells
Summary
The mammalian Target of Rapamycin (mTOR) is a central regulator of many biological processes that are essential for cell growth, including cell cycle progression, protein translation, ribosomal biogenesis, autophagy and metabolism [1,2]. Among the effects of mTOR signaling, regulation of gene expression and transcription is well characterized for several specific genes, including DNA polymerase I (Pol I), Insulin Growth Factor II (IGF II), ribosomal DNA [6,7,8] and regulators of mitochondrial oxidative function [9]. We investigated mTOR regulation of gene expression by studying rapamycin effect in two hepatic cell lines, the non-tumorigenic WB-F344 cells and the tumorigenic WB311 cells. The latter are resistant to the growth inhibitory effects of rapamycin, providing us with an opportunity to study the gene expression effects of rapamycin without confounding effects on cell proliferation
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