A mitochondrial signal peptide from Neurospora crassa increases the permeability of isolated rat liver mitochondria

Patricia M. Sokolove, Kathleen W. Kinnally

Research output: Contribution to journalArticle

Abstract

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 μM) 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 μM), 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.

Original languageEnglish (US)
Pages (from-to)69-76
Number of pages8
JournalArchives of Biochemistry and Biophysics
Volume336
Issue number1
DOIs
StatePublished - Dec 1 1996

Fingerprint

Neurospora crassa
Mitochondria
Liver Mitochondrion
Protein Sorting Signals
Liver
Swelling
Rats
Permeability
Peptides
Mitochondrial Swelling
Voltage-Dependent Anion Channels
Wasp Venoms
Dibucaine
Membranes
Trifluoperazine
Membrane structures
Egtazic Acid
Electron Transport Complex IV
Propranolol
Mitochondrial Membranes

Keywords

  • mitochondria
  • permeability transition
  • protein import
  • signal peptide

ASJC Scopus subject areas

  • Biochemistry
  • Biophysics
  • Molecular Biology

Cite this

A mitochondrial signal peptide from Neurospora crassa increases the permeability of isolated rat liver mitochondria. / Sokolove, Patricia M.; Kinnally, Kathleen W.

In: Archives of Biochemistry and Biophysics, Vol. 336, No. 1, 01.12.1996, p. 69-76.

Research output: Contribution to journalArticle

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abstract = "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 μM) 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 μM), 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.",
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