Catabolism of stored triacylglycerol (TAG) from cytoplasmic lipid droplets is critical for providing energy substrates, membrane building blocks, and signaling lipids in most cells of the body. However, the lipolytic machinery dictating TAG hydrolysis varies greatly among different cell types. Within the adipocyte, TAG hydrolysis is dynamically regulated by hormones to ensure appropriate metabolic adaptation to nutritional and physiologic cues. In other cell types such as hepatocytes, myocytes, and macrophages, mobilization of stored TAG is regulated quite differently. Within the last decade, mutations in two key genes involved in TAG hydrolysis, alpha-beta hydrolase domain 5 (ABHD5/CGI-58) and adipose triglyceride lipase (ATGL/PNPLA2), were found to cause two distinct neutral lipid storage diseases (NLSD) in humans. These genetic links, along with supporting evidence in mouse models, have prompted a number of studies surrounding the biochemical function(s) of these proteins. Although both CGI-58 and ATGL have been clearly implicated in TAG hydrolysis in multiple tissues and have even been shown to physically interact with each other, recent evidence suggests that they may also have distinct roles. The purpose of this review is to summarize the most recent insights into how CGI-58 and ATGL regulate lipid metabolism and signaling.