MAPK in Dunaliella tertiolecta regulates glycerol production in response to osmotic shock

Abstract

Glycerol production is modulated in some halo-tolerant organisms in response to extracellular osmotic stress. Glycerol-3-phosphate dehydrogenase (GPDH) is a rate-limiting enzyme of glycerol synthesis in both yeast and the green microalgae Dunaliella. In yeast, a High-Osmolarity-Glycerol (HOG) pathway, which is a mitogen-activated protein kinase (MAPK) cascade, has been suggested to activate the expression of GPDH and thus the accumulation of glycerol under osmotic stress. In Dunaliella tertiolecta, however, the osmo-regulatory mechanisms for glycerol synthesis are not well understood. In this study, the homologues of MAPK and GPDH in D. tertiolecta were cloned using Rapid Amplification of cDNA Ends (RACE) to investigate the molecular basis of the osmo-regulatory mechanisms. The isolated cDNA sequences were named DtMAPK (GenBank: KJ930518) and DtGPDH (GenBank: KJ930370), respectively. It was found that after osmotic shock DtMAPK and DtGPDH expression increased within 0.5 h and 1 h, respectively. In addition, glycerol production and DtGPDH expression level paralleled the expression of DtMAPK under various osmotic stress conditions. This suggests a close correlation between the expression of these two genes and glycerol production in D. tertiolecta. Moreover, suppressed transcription of DtGPDH and delayed accumulation of intracellular glycerol were observed in DtMAPK knock-down cells upon hyper-osmotic shock, providing further evidence that DtMAPK is involved in the regulation of DtGPDH expression and thus glycerol synthesis. Our findings demonstrate that a MAPK-mediated signalling pathway similar to the yeast HOG pathway may exist in D. tertiolecta in response to osmotic stress.