GCN-2 dependent inhibition of protein synthesis activates osmosensitive gene transcription via WNK and Ste20 kinase signaling

Abstract

Lee EC, Strange K. GCN-2 dependent inhibition of protein synthesis activates osmosensitive gene transcription via WNK and Ste20 kinase signaling. Am J Physiol Cell Physiol 303: C1269 -C1277, 2012. First published October 17, 2012; doi:10.1152/ajpcell.00294.2012.-Increased gpdh-1 transcription is required for accumulation of the organic osmolyte glycerol and survival of Caenorhabditis elegans during hypertonic stress. Our previous work has shown that regulators of gpdh-1 (rgpd) gene knockdown constitutively activates gpdh-1 expression. Fifty-five rgpd genes play essential roles in translation suggesting that inhibition of protein synthesis is an important signal for regulating osmoprotective gene transcription. We demonstrate here that translation is reduced dramatically by hypertonic stress or knockdown of rgpd genes encoding aminoacyl-tRNA synthetases and eukaryotic translation initiation factors (eIFs). Toxin-induced inhibition of translation also activates gpdh-1 expression. Hypertonicityinduced translation inhibition is mediated by general control nonderepressible (GCN)-2 kinase signaling and eIF-2␣ phosphoryation. Loss of gcn-1 or gcn-2 function prevents eIF-2␣ phosphorylation, completely blocks reductions in translation, and inhibits gpdh-1 transcription. gpdh-1 expression is regulated by the highly conserved with-no-lysine kinase (WNK) and Ste20 kinases WNK-1 and GCK-3, which function in the GCN-2 signaling pathway downstream from eIF-2␣ phosphorylation. Our previous work has shown that hypertonic stress causes rapid and dramatic protein damage in C. elegans and that inhibition of translation reduces this damage. The current studies demonstrate that reduced translation also serves as an essential signal for activation of WNK-1/GCK-3 kinase signaling and subsequent transcription of gpdh-1 and possibly other osmoprotective genes.C. elegans; organic osmolytes; hypertonic stress; osmoregulation MAINTENANCE OF CELLULAR AND extracellular water balance is essential for life. All cells are exposed to osmotic stress from changes in intracellular solute levels that occur during normal fluctuations in metabolism and membrane solute flux. Countless diverse organisms live in osmotically unstable environments and are exposed to extreme changes in extracellular osmolality. Mammalian cells are largely protected from extracellular osmotic perturbations by the kidney, which tightly regulates blood ionic and osmotic concentrations. However, cells in the renal medulla and gut are subjected normally to large osmotic challenges due to the renal concentrating mechanism and changes in fluid and food intake. In addition, several disorders, such as renal failure, diabetes, syndrome of inappropriate antidiuretic hormone secretion, and hypernatremia, disrupt plasma osmolality, and hypertonic solutions are used widely to treat diverse clinical problems such as intracranial hypertension and hemorrhagic shock. Cellular osmotic homeostasis requires the regulated gain or loss of solutes and activation of mechanisms that repair or remove ...

Physiological and molecular mechanisms of salt and water homeostasis in the nematodeCaenorhabditis elegans

“…The authors first confirmed that silencing of multiple genes previously found to repress a gpdh-1 reporter (61) reduce protein synthesis and increase expression of endogenous gpdh-1 mRNA (62). They also demonstrated that protein synthesis is reduced nearly 50%, and gpdh-1 mRNA is induced Ͼ50-fold, within an hour of transfer to hypertonic agar (Fig.…”

“…Mechanisms used to regulate protein synthesis during hypertonic stress and the downstream signals that link protein homeostasis to osmosensitive gene expression were investigated in a recent study (62). The authors first confirmed that silencing of multiple genes previously found to repress a gpdh-1 reporter (61) reduce protein synthesis and increase expression of endogenous gpdh-1 mRNA (62).…”

“…WNK1 and PASK/OSR1, the mammalian homologs of WNK-1 and GCK-3, function together to regulate volume-sensitive ion transporters, renal salt transporters, and blood pressure (41), indicating that the function of this signaling cascade in salt and water homeostasis was highly conserved. It remains to be seen whether WNK1 and PASK/OSR1 also play a role in regulating osmotic responsive gene expression as was recently shown in C. elegans (62). Protein confirmation is determined by fundamental chemical and biophysical principles.…”

“…Interestingly, while investigating downstream signaling mechanisms in an RNAi screen for genes required for gpdh-1 expression, the authors recovered wnk-1, the protein kinase previously found to be required for volume recovery (18). Induction of gpdh-1, but not eIF phosphorylation, requires wnk-1 and gck-3, suggesting that the WNK/GCK-VI cascade may function to activate osmosensitive gene expression downstream from changes in protein synthesis (62). Importantly, these findings are the first to link regulation of ion transport by the conserved WNK/GCK-VI cascade with protein synthesis and osmosensitive gene expression in any species (Fig.…”

“…GPDH-1 catalyzes the rate-limiting step of glycerol synthesis. The gpdh-1 gene is expressed at low levels in worms on standard agar but is induced several-fold within the first hour of exposure to nonlethal high osmolarity (59,(61)(62) (Fig. 3).…”