Sokolove P M, Kinnally K W
Department of Pharmacology & Experimental Therapeutics, University of Maryland School of Medicine, Baltimore 21201, USA.
Arch Biochem Biophys. 1996 Dec 1;336(1):69-76. doi: 10.1006/abbi.1996.0533.
Mitochondria that contain Ca2+ can be induced by a variety of triggering agents and conditions to undergo a permeability transition (PT); the inner membrane becomes nonselectively permeable to small solutes. Mastoparan, an amphipathic peptide from wasp venom, has recently been reported to induce this transition (Pfeiffer et al., 1995, J. Biol. Chem. 270,4923). We have examined the effect on the permeability of isolated rat liver mitochondria of a second amphipathic peptide, the signal sequence of cytochrome oxidase subunit IV from Neurospora crassa (pCoxIV, amino acids 3-22), which targets subunit IV to its mitochondrial location. Permeability increases were visualized via mitochondrial swelling with the following results. (1) pCoxIV (5-100 microM) induced concentration-dependent mitochondrial swelling. Control peptides from the N- and C-termini of the voltage-dependent anion-selective channel had no such effect. (2) Swelling required mitochondrial energization; it was eliminated or halted by the uncoupler carbonyl cyanide p-(trifluoromethoxy)phenylhydrazone. (3) Peptide-induced swelling was slowed by increasing concentrations of KCl. (4) Swelling was enhanced by inorganic phosphate (<1 mM). (5) Trifluoperazine (50 microM), propranolol (0.5 mM), and dibucaine (0.5 mM) were potent inhibitors of peptide-induced swelling, whereas other inhibitors of the classical PT (cyclosporin A, EGTA, and ADP) inhibited only partially. (6) pCoxIV opened a pore rather than disrupting mitochondrial membrane structure, but 50% inhibition of peptide-induced swelling required polyethylene glycol of molecular weight substantially larger than that needed to inhibit the Ca2+-induced PT to the same extent. In summary, pCoxIV opens a pore in isolated mitochondria. The dependence of pore opening on membrane potential and the inhibition of the peptide-induced permeability increase by increasing salt concentration suggest that this effect of the signal peptide is related to its interactions with mitochondria during protein import. The peptide-induced pore appears, however, to be distinct from both the classical permeability transition pore and the mastoparan-induced permeability increase.
含有钙离子的线粒体可被多种触发剂和条件诱导发生通透性转变（PT）；内膜对小溶质变得非选择性通透。蜂毒中的两亲性肽马斯托帕兰最近被报道可诱导这种转变（Pfeiffer等人，1995年，《生物化学杂志》270,4923）。我们研究了另一种两亲性肽——粗糙脉孢菌细胞色素氧化酶亚基IV的信号序列（pCoxIV，氨基酸3 - 22）对分离的大鼠肝线粒体通透性的影响，该信号序列将亚基IV靶向到其线粒体位置。通过线粒体肿胀观察到通透性增加，结果如下：（1）pCoxIV（5 - 100微摩尔）诱导浓度依赖性的线粒体肿胀。电压依赖性阴离子选择性通道的N端和C端的对照肽没有这种作用。（2）肿胀需要线粒体供能；解偶联剂羰基氰化物对-（三氟甲氧基）苯腙可消除或阻止肿胀。（3）肽诱导的肿胀随着氯化钾浓度增加而减缓。（4）无机磷酸盐（<1毫摩尔）增强肿胀。（5）三氟拉嗪（50微摩尔）、普萘洛尔（0.5毫摩尔）和丁卡因（0.5毫摩尔）是肽诱导肿胀的有效抑制剂，而经典PT的其他抑制剂（环孢素A）、乙二醇双乙酸盐（EGTA）和二磷酸腺苷（ADP）仅部分抑制。（6）pCoxIV打开一个孔而不是破坏线粒体膜结构，但对肽诱导肿胀的50%抑制所需的聚乙二醇分子量显著大于在相同程度上抑制钙离子诱导的PT所需的分子量。总之，pCoxIV在分离的线粒体中打开一个孔。孔开放对膜电位的依赖性以及通过增加盐浓度对肽诱导的通透性增加的抑制表明，信号肽的这种作用与其在蛋白质导入过程中与线粒体的相互作用有关。然而，肽诱导的孔似乎与经典的通透性转变孔和马斯托帕兰诱导的通透性增加都不同。
