The genetic mechanisms of osmoregulation accompanied by salinity changes in aquatic crustaceans are still unclear. In this study, two different salinity transition experiments were conducted, and the gene expression profiles of gill tissues between the two salinity transition processes were compared to unveil the genetic basis of osmoregulation and high/low salinity acclimation for Eriocheir sinensis. A total of 910 genes were significantly differentially expressed within the two salinity transition processes. Genes associated with ion transport, signal transduction, immune response, and DNA repair generally showed fluctuated expressions in the two salinity transition processes. Genes associated with the nitrogen compound metabolic process, sexual reproduction system, and ammonium transport, such as AASS, HDAC4, VG, MARF1, VMO1, and RHBG, were upregulated in a high salinity environment. Genes associated with lipid metabolism processes, such as OXCT1, TEX2, DIB, and CHKB, were upregulated in a low salinity environment. Results revealed the common and specific responsive mechanism of osmoregulation and different levels of salinity acclimation within the two salinity transition processes of E. sinensis. This study provides valuable genetic resources and novel insights into the osmoregulation research of crustaceans.