Transcriptomics of varying strength and timing of specific activation of the Integrated Stress Response

The integrated stress response (ISR) is a central signaling pathway induced by a variety of insults, but how its outputs contribute to downstream physiological effects across diverse cellular contexts remains unclear. Using a synthetic tool, we specifically and tunably activated the ISR and performed multi-omics profiling to define the core modules elicited by this response in the absence of co-activation of parallel pathways commonly induced by pleiotropic stressors. We found that the ISR can elicit time- and dose-dependent gene expression changes that cluster into four modules with ATF4 driving only a small but fast and sensitive module that includes many amino acid metabolic enzymes. We showed that ATF4 was required to reroute carbon utilization towards amino acid synthesis derived both from glucose and reductive carboxylation of glutamine and away from the tricarboxylic acid cycle and fatty acid biogenesis revealing a new role for ATF4 in modulating cellular energetics. We also discovered an ATF4-independent reorganization of cellular lipids that promotes triglycerides synthesis and accumulation of lipid droplets that was essential for cell survival. Together, we demonstrate that a minimal ISR-inducing system is sufficient to trigger formation of two distinct cellular structures, stress granules and lipid droplets, and a previously unappreciated metabolic state.