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Tim23, a protein import component of the mitochondrial inner membrane, is required for normal activity of the multiple conductance channel, MCC.

Lohret TA, Jensen RE, Kinnally KW - J. Cell Biol. (1997)

Bottom Line: We also find that MCC activity is altered in mitochondria isolated from yeast carrying the tim23-1 mutation.Tim23 antibodies and the tim23-1 mutation do not, however, alter the activity of PSC, a presequence-peptide sensitive channel in the mitochondrial outer membrane.Our results show that Tim23p is required for normal MCC activity and raise the possibility that precursors are translocated across the inner membrane through the pore of MCC.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Sciences, University at Albany, SUNY, New York 12222, USA.

ABSTRACT
We previously showed that the conductance of a mitochondrial inner membrane channel, called MCC, was specifically blocked by peptides corresponding to mitochondrial import signals. To determine if MCC plays a role in protein import, we examined the relationship between MCC and Tim23p, a component of the protein import complex of the mitochondrial inner membrane. We find that antibodies against Tim23p, previously shown to inhibit mitochondrial protein import, inhibit MCC activity. We also find that MCC activity is altered in mitochondria isolated from yeast carrying the tim23-1 mutation. In contrast to wild-type MCC, we find that the conductance of MCC from the tim23-1 mutant is not significantly blocked by mitochondrial presequence peptides. Tim23 antibodies and the tim23-1 mutation do not, however, alter the activity of PSC, a presequence-peptide sensitive channel in the mitochondrial outer membrane. Our results show that Tim23p is required for normal MCC activity and raise the possibility that precursors are translocated across the inner membrane through the pore of MCC.

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The conductance of MCC isolated from the tim23-1  mutant is not blocked by presequence peptides in the bath. Proteoliposomes were prepared from inner membranes from wildtype cells (A) or the tim23-1 mutant (B), and current traces of  MCC activity were recorded from patches in the presence and absence of 50 μM yCOX-IV1-13 peptide in the bath solution. (C)  The flicker rate (events/sec) from the open state to lower conductance states of MCC at 20 mV from wild-type and tim23-1 strains  was determined in the absence (control) and presence of 50 μM  of either yCOX-IV1-13 or fCOX-IV3-22. Error bars indicate standard deviations from a minimum of four determinations.
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Figure 5: The conductance of MCC isolated from the tim23-1 mutant is not blocked by presequence peptides in the bath. Proteoliposomes were prepared from inner membranes from wildtype cells (A) or the tim23-1 mutant (B), and current traces of MCC activity were recorded from patches in the presence and absence of 50 μM yCOX-IV1-13 peptide in the bath solution. (C) The flicker rate (events/sec) from the open state to lower conductance states of MCC at 20 mV from wild-type and tim23-1 strains was determined in the absence (control) and presence of 50 μM of either yCOX-IV1-13 or fCOX-IV3-22. Error bars indicate standard deviations from a minimum of four determinations.

Mentions: We found that the electrical properties of MCC isolated from wild-type and tim23-1 strains were virtually identical (compare control current traces of Fig. 5 A with 5 B and Fig. 6 A with 6 B). In particular, MCC from both strains had the same peak conductance, predominant transition size, mean open time, and cation selectivity. Furthermore, permeability ratios for K+/Cl− were ∼6 for MCC from wildtype and tim23-1 patches with a 150:30 mM KCl gradient. Conductance through MCC from both strains was voltage dependent, i.e., MCC is predominantly open at low (e.g., 20 mV) but not high potentials of either polarity. The V0 (voltage where the probability of opening and closing are the same) at positive potentials was less than V0 at negative potentials for MCC from both wild-type and tim23-1 strains.


Tim23, a protein import component of the mitochondrial inner membrane, is required for normal activity of the multiple conductance channel, MCC.

Lohret TA, Jensen RE, Kinnally KW - J. Cell Biol. (1997)

The conductance of MCC isolated from the tim23-1  mutant is not blocked by presequence peptides in the bath. Proteoliposomes were prepared from inner membranes from wildtype cells (A) or the tim23-1 mutant (B), and current traces of  MCC activity were recorded from patches in the presence and absence of 50 μM yCOX-IV1-13 peptide in the bath solution. (C)  The flicker rate (events/sec) from the open state to lower conductance states of MCC at 20 mV from wild-type and tim23-1 strains  was determined in the absence (control) and presence of 50 μM  of either yCOX-IV1-13 or fCOX-IV3-22. Error bars indicate standard deviations from a minimum of four determinations.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2139772&req=5

Figure 5: The conductance of MCC isolated from the tim23-1 mutant is not blocked by presequence peptides in the bath. Proteoliposomes were prepared from inner membranes from wildtype cells (A) or the tim23-1 mutant (B), and current traces of MCC activity were recorded from patches in the presence and absence of 50 μM yCOX-IV1-13 peptide in the bath solution. (C) The flicker rate (events/sec) from the open state to lower conductance states of MCC at 20 mV from wild-type and tim23-1 strains was determined in the absence (control) and presence of 50 μM of either yCOX-IV1-13 or fCOX-IV3-22. Error bars indicate standard deviations from a minimum of four determinations.
Mentions: We found that the electrical properties of MCC isolated from wild-type and tim23-1 strains were virtually identical (compare control current traces of Fig. 5 A with 5 B and Fig. 6 A with 6 B). In particular, MCC from both strains had the same peak conductance, predominant transition size, mean open time, and cation selectivity. Furthermore, permeability ratios for K+/Cl− were ∼6 for MCC from wildtype and tim23-1 patches with a 150:30 mM KCl gradient. Conductance through MCC from both strains was voltage dependent, i.e., MCC is predominantly open at low (e.g., 20 mV) but not high potentials of either polarity. The V0 (voltage where the probability of opening and closing are the same) at positive potentials was less than V0 at negative potentials for MCC from both wild-type and tim23-1 strains.

Bottom Line: We also find that MCC activity is altered in mitochondria isolated from yeast carrying the tim23-1 mutation.Tim23 antibodies and the tim23-1 mutation do not, however, alter the activity of PSC, a presequence-peptide sensitive channel in the mitochondrial outer membrane.Our results show that Tim23p is required for normal MCC activity and raise the possibility that precursors are translocated across the inner membrane through the pore of MCC.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Sciences, University at Albany, SUNY, New York 12222, USA.

ABSTRACT
We previously showed that the conductance of a mitochondrial inner membrane channel, called MCC, was specifically blocked by peptides corresponding to mitochondrial import signals. To determine if MCC plays a role in protein import, we examined the relationship between MCC and Tim23p, a component of the protein import complex of the mitochondrial inner membrane. We find that antibodies against Tim23p, previously shown to inhibit mitochondrial protein import, inhibit MCC activity. We also find that MCC activity is altered in mitochondria isolated from yeast carrying the tim23-1 mutation. In contrast to wild-type MCC, we find that the conductance of MCC from the tim23-1 mutant is not significantly blocked by mitochondrial presequence peptides. Tim23 antibodies and the tim23-1 mutation do not, however, alter the activity of PSC, a presequence-peptide sensitive channel in the mitochondrial outer membrane. Our results show that Tim23p is required for normal MCC activity and raise the possibility that precursors are translocated across the inner membrane through the pore of MCC.

Show MeSH
Related in: MedlinePlus