Glycogen synthase 2 knock-out zebrafish exhibit enhanced glucose metabolism in the liver

The liver is one of the major organs for glycogen storage and is essential for maintaining glucose homeostasis in the whole body. Glycogen synthesis in liver is regulated by the rate-limiting enzyme, glycogen synthase 2 (Gys2). In fish, however, the detailed roles of glycogen and Gys2 are poorly understood. In this study, we generated, for the first time, gys2 knock-out (gys2 -/-) zebrafish and evaluated the effects of impaired glycogen synthesis on physiological parameters of this fish. No effects on development or maturation were observed in gys2 -/- strain lacking Gys2. Periodic acid Schiff (PAS) staining, immunofluorescence staining with anti-glycogen antibody, and transmission electron microscopy (TEM) observations confirmed that glycogen synthesis does not occur in the gys2 -/- liver. Liquid chromatography-mass spectrometry (LC-MS/MS) metabolite analysis and isotope tracing revealed a marked increase in the activity of glucose metabolism and levels of related metabolites in the livers of adult gys2 -/-. Surprisingly, glucoamylase treatment released higher amount of glucose from gys2 -/- liver extract than that of the wild-type (WT) counterpart, suggesting the existence of putative compensatory or abberant glucose polymer, which might be caused by the elevated glucose metabolism in the gys2 -/- liver. Meanwhile, RNA-Seq analysis did not show increased expression levels of glucose metabolism-related genes consistent with the metabolic activity in gys2 -/- liver. These results suggest that liver Gys2 is dispensable in zebrafish, and enhanced glucose metabolism and putative backup mechanism to accumulate extra glucose may support the fish lacking Gys2.