Mitochondria are surrounded by a double membrane system that forms four intra-organelle compartments: the outer membrane, inner membrane, intermembrane space, and matrix. Each of the two membranes contains a unique set of proteins defining specific functions of that membrane. The vast majority of mitochondrial proteins including those of the mitochondrial membranes are nuclear encoded and synthesized as precursor proteins in the cytosol. Subsequently, they are targeted to the mitochondria and become sorted to the correct submitochondrial destination. A small portion of the mitochondrial inner membrane proteins is encoded by the mitochondrial genome. These proteins are synthesized on mitochondrial ribosomes and are inserted by dedicated machinery into the inner membrane. This chapter summarizes our current knowledge of the signals that target mitochondrial membrane proteins to their correct intracellular location, and describes the mechanisms by which mitochondrial translocation machineries recognize precursor proteins and mediate their insertion into mitochondrial membranes.