Cell-penetrating peptides are excluded from the mitochondrial matrix.

CPPs (cell-penetrating peptides) facilitate cellular uptake of covalently attached macromolecules, through an as yet controversial mechanism that either involves direct membrane passage or a type of endocytosis. We investigated the potential of the CPPs penetratin and Tat to act as mitochondria-targeting vectors by testing whether they were internalized by isolated mitochondria, and by mitochondria within cells in culture. We also tested peptides conjugated to the mitochondria-targeting moiety triphenylphosphonium. We found no evidence for mitochondrial uptake by penetratin, Tat or their triphenylphosphonium conjugates. This result suggests that CPPs are unsuitable as mitochondria-targeting vectors, and implies an endocytic mode of cellular uptake for CPPs.