The signature mitochondrial phospholipid cardiolipin plays an important role in mitochondrial function, and alterations in cardiolipin metabolism are associated with human disease. Topologically, cardiolipin biosynthesis and remodeling are complex. Precursor phospholipids must be transported from the ER, across the mitochondrial outer membrane to the matrix-facing leaflet of the inner membrane, where cardiolipin biosynthesis commences. Post-synthesis, cardiolipin undergoes acyl chain remodeling, requiring additional trafficking steps, before it achieves its final distribution within both mitochondrial membranes. This process is regulated at several points via multiple independent mechanisms. Here, we review the regulation and topology of cardiolipin biosynthesis and remodeling in the yeast Saccharomyces cerevisiae. Although cardiolipin metabolism is more complicated in mammals, yeast have been an invaluable model for dissecting the steps required for this process. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.