Occurrence and Characteristics of the Mitochondrial Permeability Transition in Plants

Abstract

The behavior of purified potato mitochondria toward the main effectors of the animal mitochondrial permeability transition has been studied by light scattering, fluorescence, SDS-polyacrylamide gel electrophoresis, and immunoblotting techniques. The addition of Ca 2؉ induces a phosphate-dependent swelling that is fully inhibited by cyclosporin A if dithioerythritol is present. Mg 2؉ cannot be substituted for Ca 2؉ but competes with it. Disruption of the outer membrane and release of several proteins, including cytochrome c, occur upon completion of swelling. Ca 2؉ -induced swelling is delayed and its rate is decreased when pH is shifted from 7.4 to 6.6. It is accelerated by diamide, phenylarsine oxide, and linolenic acid. In the absence of Ca 2؉ , however, linolenic acid (<20 M) rapidly dissipates the succinate-driven membrane potential while having no effect on mitochondrial volume. Anoxic conditions favor in vitro swelling and the concomitant release of cytochrome c and of other proteins in a pH-dependent way. These data indicate that the classical mitochondrial permeability transition occurs also in plants. This may have important implications for our understanding of cell stress and death processes.Since the late 1970s, it has been known that animal mitochondria can experience a sudden increase in the permeability of their inner membrane to low and medium molecular weight compounds via the opening of a pore (1-3). This mitochondrial permeability transition pore (PTP) 1 is viewed as a multiprotein complex composed at least of the voltage-dependent anion channel, the adenine nucleotide translocator (AdNT), and cyclophilin-D, at the contact sites between outer and inner membranes (4). When the pore opens, solutes up to about 1.5 kDa can pass through the inner membrane, a process known as the mitochondrial permeability transition (MPT). Subsequently, the membrane potential (⌬⌿) decays, oxidative phosphorylation is uncoupled from electron flow, intramitochondrial ions and metabolites are released, and a large amplitude swelling can occur, disrupting the outer membrane and releasing intermembrane compounds.Although pore opening primarily requires the accumulation of Ca 2ϩ in the mitochondrial matrix, it is also modulated by numerous factors. For instance, P i , low ⌬⌿, thiol-oxidizing reagents, low ATP level, fatty acids, anoxia, and reaeration stress all favor pore opening, whereas thiol-reducing agents, low pH, high ⌬⌿, and divalent cations other than Ca 2ϩ counteract it (5). Inhibition of MPT is readily achieved with submicromolar concentrations of cyclosporin A (CsA) (6, 7). This highly specific effect has decisively contributed to the acceptance of the pore theory (6, 7) and is used today as the primary diagnostic trait of the classical MPT (5). The implication of mitochondria and PTP in mammalian cell death gave a new impetus to the research. For instance, cytochrome c has been shown to be released from the mitochondrial intermembrane space into the cytosol (8, 9), where it can trigger apoptosis (10). How ...

Identification and Characterization of Cannabinoids That Induce Cell Death through Mitochondrial Permeability Transition in Cannabis Leaf Cells

“…The system that regulates MPT in a Ca 2ϩ -independent manner has not been identified in plants to date. In contrast, Ca 2ϩ -mediated MPT has been reported in the mitochondria of several plants: swelling is induced in wheat mitochondria and potato tuber mitochondria by treatment with high concentrations (2.5-5.0 mM) of Ca 2ϩ in the presence of inorganic phosphate (15,16). However, we found that the properties of the swelling described in those studies are different from those of cannabinoid-treated mitochondrial swelling.…”

“…These results suggested that CBCA and THCA mediate cell death though MPT in Cannabis leaves. Mitochondrial Swelling and ⌬⌿m Reduction by Cannabinoids-Like animal MPT, plant MPT is also considered to be triggered by opening of MPT pores, leading to mitochondrial swelling and loss of mitochondrial membrane potential (⌬⌿m) (16). Therefore, we examined whether CBCA and THCA induce such responses in Cannabis mitochondria.…”

“…Cyclosporin A (CsA) clearly suppressed these cell death responses in cannabinoid-treated cells. Because CsA inhibits the MPT in plants as well as animals (10,16,36), CBCA and THCA were considered to cause cell death through MPT.…”

“…cytochrome c) through collapse of the mitochondrial outer membrane (12,13,15,16,38,39). To examine whether CBCA and THCA induce such collapse, we analyzed proteins released from Cannabis mitochondria treated with these agents.…”

“…Indeed, wheat mitochondria (15) and potato tuber mitochondria (16) have been shown to undergo high amplitude swelling through MPT in response to treatment with high concentrations of Ca 2ϩ , leading to release of cyto-chrome c. However, it has remained unknown whether Ca 2ϩ -mediated MPT, which causes necrosis in animal cells (13,14), induces apoptosis or necrosis in plant cells, because the reported MPT was induced in isolated mitochondria and was not examined in wheat cells or potato tuber cells. Furthermore, it has been reported that Ca 2ϩ induces little swelling in mitochondria isolated from oats (17) and Arabidopsis thaliana (18), suggesting that, in some plants, opening of MPT pores is regulated by molecules other than Ca 2ϩ .…”