Salinity-induced activation of the myo-inositol biosynthesis pathway in tilapia gill epithelium

Abstract

Summary The myo-inositol biosynthesis (MIB) pathway converts glucose-6-phosphate to the compatible osmolyte myo-inositol (MI) that protects cells from osmotic stress. Using proteomics, the enzymes that constitute the MIB pathway, myo-inositol phosphate synthase (MIPS) and inositol monophosphatase 1 (IMPA1), are identified in tilapia (Oreochromis mossambicus) gill epithelium. Targeted, quantitative, label-free proteomics reveals that they are both up-regulated during salinity stress. Up-regulation is stronger when fish are exposed to severe (34 ppt acute and 90 ppt gradual) relative to moderate (70 ppt gradual) salinity stress. IMPA1 always responds more strongly than MIPS suggesting that MIPS is more stable during salinity stress. MIPS is amino-terminally acetylated and the corresponding peptide increases proportionally to MIPS protein while non-acetylated amino-terminal peptide is not detectable indicating that MIPS acetylation is constitutive and may serve to stabilize the protein. Hyperosmotic induction of MIPS and IMPA1 is confirmed using Western blot and real-time qPCR and much higher at mRNA than protein level. Two distinct MIPS mRNA variants are expressed in gill, but one is more strongly regulated by salinity than the other. A single MIPS gene is encoded in the tilapia genome whereas the zebrafish genome lacks MIPS entirely. The genome of euryhaline tilapia contains 4 IMPA genes, two of which are expressed but only one is salinity-regulated in gill epithelium. The genome of stenohaline zebrafish contains a single IMPA gene. We conclude that the MIB pathway represents a major salinity stress coping mechanism that is regulated at multiple levels in euryhaline fish but absent in stenohaline zebrafish.