Glucose Limitation Induces GCN4 Translation by Activation of Gcn2 Protein Kinase

Abstract

Phosphorylation of the α subunit of eukaryotic initiation factor 2 (eIF-2α) is a well-characterized mechanism regulating protein synthesis in response to environmental stresses. In the yeastSaccharomyces cerevisiae, starvation for amino acids induces phosphorylation of eIF-2α by Gcn2 protein kinase, leading to elevated translation of GCN4, a transcriptional activator of more than 50 genes. Uncharged tRNA that accumulates during amino acid limitation is proposed to activate Gcn2p by associating with Gcn2p sequences homologous to histidyl-tRNA synthetase (HisRS) enzymes. Given that eIF-2α phosphorylation in mammals is induced in response to both carbohydrate and amino acid limitations, we addressed whether activation of Gcn2p in yeast is also controlled by different nutrient deprivations. We found that starvation for glucose induces Gcn2p phosphorylation of eIF-2α and stimulates GCN4translation. Induction of eIF-2α phosphorylation by Gcn2p during glucose limitation requires the function of the HisRS-related domain but is largely independent of the ribosome binding sequences of Gcn2p. Furthermore, Gcn20p, a factor required for Gcn2 protein kinase stimulation of GCN4 expression in response to amino acid starvation, is not essential for GCN4 translational control in response to limitation for carbohydrates. These results indicate there are differences between the mechanisms regulating Gcn2p activity in response to amino acid and carbohydrate deficiency. Gcn2p induction of GCN4 translation during carbohydrate limitation enhances storage of amino acids in the vacuoles and facilitates entry into exponential growth during a shift from low-glucose to high-glucose medium. Gcn2p function also contributes to maintenance of glycogen levels during prolonged glucose starvation, suggesting a linkage between amino acid control and glycogen metabolism.

ACKNOWLEDGMENTS

We thank Mark Goebl, Peter Roach, Janice Blum, Anna DePaoli-Roach, Wayne Wilson, Shuhao Zhu, Krishna Vattem, and members of the Wek laboratory for advice during the course of this work and comments on the manuscript. Additional thanks go to Alan Hinnebusch for helpful discussions and generously providing plasmids, strains and reagents, Gary Krause for eIF-2α antibody, and Robert Harris and Jinnie Garret for advice on amino acid measurements.

This work was supported in part by Public Health Service grant GM49164 from the National Institutes of Health and by American Cancer Society grant RPG MBC-87806 (R.C.W.).