Tetracyclines cause cell stress-dependent ATF4 activation and mTOR inhibition

• Tetracyclines cause ATF4 activation, resulting in enhanced expression of ATF4 downstream targets inhibin βE, ATF3, and CHOP.
• Prolonged treatment with tetracyclines leads to mTOR inhibition, resulting in autophagy and cell cycle arrest.
• The effect of tetracyclines on cell viability is magnesium dependent.
• Minocycline exerts a selective steatogenic effect.

Tetracyclines have long been used as valuable broad-spectrum antibiotics. The high antibacterial activity of tetracyclines, combined with their good tolerability, has led to their widespread use in treating various infectious diseases. However, similar to other antibiotics, tetracyclines are also known for their adverse effects on different human tissues, including hepatic steatosis. We observed that tetracyclines, including doxycycline and minocycline, caused enhanced expression of the liver chalone inhibin βE in HepG2 cells, mediated by the cell stress-regulated transcription factor ATF4. ATF4 and its target genes ATF3, CHOP, and inhibin βE are involved in cell cycle control, cell survival, cell metabolism, and modulation of cytokine expression. Furthermore, we observed that long term tetracycline incubation also caused inhibition of the mTOR complex, a central regulator of cell metabolism, further contributing to the observed cell-cycle arrest and autophagy in doxycycline- and minocycline-treated cell lines. ATF4 activation and mTOR inhibition link two crucial regulators of the cellular stress response and cell metabolism to the effects of tetracyclines on eukaryotic cell metabolism, and may help to understand the antibiotic-independent influence of these drugs on human tissues. Since the observed effects of tetracyclines on human cells were also found to be dependent on the magnesium ion concentrations supplied, the data further indicate the importance of magnesium supplementation to reduce or prevent side effects of long term treatment with tetracyclines.