Signal presequences increase mitochondrial permeability and open the multiple conductance channel

We have reported that the signal presequence of cytochrome oxidase subunit IV from Neurospora crassa increases the permeability of isolated rat liver mitochondria [P. M. Sokolove and K. W. Kinnally (1996) Arch. Biochem. Biophys. 336, 69] and regulates the behavior of the mutiple conductance channel (MCC) of yeast inner mitochondrial membrane [T. A. Lohret and K. W. Kinnally (1995) J. Biol. Chem. 270, 15950]. Here we examine in greater detail the action of a number of mitochondrial presequences from various sources and of several control peptides on the permeability of isolated rat liver mitochondria and on MCC activity monitored via patch-clamp techniques in both mammalian mitoplasts and a reconstituted yeast system. The data indicate that the ability to alter mitochondrial permeability is a property of most, but not all, signal peptides. Furthermore, it is clear that, although signal peptides are characterized by positive charge and the ability to form amphiphilic alpha helices, these two characteristics are not sufficient to guarantee mitochondrial effects. Finally, the results reveal a strong correlation between peptide effects on the permeability of isolated mitochondria and on MCC activity: peptides that induced swelling of mouse and rat mitochondria also activated the quiescent MCC of mouse mitoplasts and induced flickering of active MCC reconstituted from yeast mitochondrial membranes. Moreover, relative peptide efficacies were very similar for mitochondrial swelling and both types of patch-clamp experiments. We propose that patch-clamp recordings of MCC activity and the high-amplitude swelling induced by signal peptides reflect the opening of a single channel. Based on the selective responsiveness of that channel to signal peptides and the dependence of its opening in isolated mitochondria on membrane potential, we further suggest that the channel is involved in the mitochondrial protein import process.