Morven Graham

Mitochondria maintain tight regulation of inner mitochondrial membrane (IMM) permeability to sustain ATP production. Stressful events cause cellular calcium (Ca(2+)) dysregulation followed by rapid loss of IMM potential known as permeability transition (PT), which produces osmotic shifts, metabolic dysfunction, and cell death. The molecular identity of the mitochondrial PT pore (mPTP) was previously unknown. We show that the purified reconstituted c-subunit ring of the FO of the F1FO ATP synthase forms a voltage-sensitive channel, the persistent opening of which leads to rapid and uncontrolled depolarization of the IMM in cells. Prolonged high matrix Ca(2+) enlarges the c-subunit ring and unhooks it from cyclophilin D/cyclosporine A binding sites in the ATP synthase F1, providing a mechanism for mPTP opening. In contrast, recombinant F1 beta-subunit applied exogenously to the purified c-subunit enhances the probability of pore closure. Depletion of the c-subunit attenuates Ca(2+)-induced IMM depolarization and inhibits Ca(2+) and reactive oxygen species-induced cell death whereas increasing the expression or single-channel conductance of the c-subunit sensitizes to death. We conclude that a highly regulated c-subunit leak channel is a candidate for the mPTP. Beyond cell death, these findings also imply that increasing the probability of c-subunit channel closure in a healthy cell will enhance IMM coupling and increase cellular metabolic efficiency.

and human cells, we show here that seipin, an ER protein implicated in LD biology, mediates a discrete step in LD formation-the conversion of small, nascent LDs to larger, mature LDs. Seipin forms discrete and dynamic foci in the ER that interact with nascent LDs to enable their growth. In the absence of seipin, numerous small, nascent LDs accumulate near the ER and most often fail to grow. Those that do grow prematurely acquire lipid synthesis enzymes and undergo expansion, eventually leading to the giant LDs characteristic of seipin deficiency. Our studies identify a discrete step of LD formation, namely the conversion of nascent LDs to mature LDs, and define a molecular role for seipin in this process, most likely by acting at ER-LD contact sites to enable lipid transfer to nascent LDs.