Salinity is an important factor that impacts the physiology and ecological competitiveness of dinoflagellates in marine environments. However, little is known about its regulatory mechanisms through the encoded gene and metabolite network. In this study, the physiological response, transcriptome and broadly targeted metabolic profile of Karenia mikimotoi from simulated oceanic conditions (salinity of 33) to acclimate to the optimal growth conditions (salinity of 25) were investigated. The result revealed various biological pathways and metabolic processes as key influences of salinity change. The combined transcriptome and metabolome analyses identified several metabolites, including S-adenosyl-l-methionine (SAM) and N-acetylputrescine, that act as nodes in the regulation network. In low salinity culture, cells showed faster growth, higher photosynthetic efficiency, lower ROS generation, and lower enzymatic scavenging than those in higher salinity culture. Meanwhile, gene superoxide dismutase was downregulated in low salinity culture. While the accumulation of nonenzymatic antioxidants such as polyamine and cysteine were enhanced under low salinity conditions. Furthermore, genes related to carbon metabolism, biosynthesis of amino acids, sulfur metabolism and glycolysis were up-regulated. The up-regulated expression of cysteine and methionine metabolism-related genes may also be the acclimated mechanisms to increase carbon metabolism processes under low salinity. The enhanced glyoxylate shunt was also observed under low salinity to bypass the decarboxylation steps in the TCA cycle, to modulate carbon balance, and cause an increased flow of carbon to glucose. These findings shed light on a comprehensive sophisticated strategies of K. mikimotoi to respond to low salinity conditions.