Orchestrating biological activities of immune cells through metabolic reprogramming reveals a new approach to harnessing immune responses. Increasing evidence reveals that the mitochondrion is a central regulator for modulating metabolic reprogramming and controlling immune cell activation and functions. In addition to supporting bioenergetic demands, the mitochondrion serves as a signaling platform through the generation of reactive oxygen species and metabolites of the tricarboxylic acid cycle to modulate signaling cascades controlling immune cell activation and immune responses. Herein, we discuss the mechanisms through which the mitochondrion acts as a master regulator to fine-tune immune responses elicited by macrophages and T cells.