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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 tim23-1 mutation does not affect the peptide-sensitive channel, PSC, of the mitochondrial outer membrane. (A)  Sample current traces for PSC from proteoliposomes containing  outer membranes from wild-type or tim23-1 strains are shown in  the presence and absence (control) of 50 μM yCOX-IV1-13 peptide in the bath. (B) The flicker rates of PSC of wild-type and  tim23-1 strains were determined from current traces represented  by those in A above.
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Figure 9: The tim23-1 mutation does not affect the peptide-sensitive channel, PSC, of the mitochondrial outer membrane. (A) Sample current traces for PSC from proteoliposomes containing outer membranes from wild-type or tim23-1 strains are shown in the presence and absence (control) of 50 μM yCOX-IV1-13 peptide in the bath. (B) The flicker rates of PSC of wild-type and tim23-1 strains were determined from current traces represented by those in A above.

Mentions: To show that the tim23-1 mutation specifically affected MCC activity, we examined the activity of the outer membrane peptide-sensitive channel, PSC. Like MCC in the inner membrane, the conductance of PSC is transiently blocked by presequence peptides (Henry et al., 1996; Fèvre et al., 1994; Theiffry et al., 1992; Juin et al., 1995). Outer membranes were isolated from mitochondria of wild-type cells and the tim23-1 mutant, and then reconstituted into proteoliposomes. When membrane patches were examined, we found that the PSC activities from both preparations were comparable in terms of conductance, selectivity, and voltage dependence. Furthermore, addition of the yCOX-IV1-13 peptide increased the flicker rate of PSC from both wild-type and tim23-1 strains by ∼10fold as shown by the current traces and histograms of Fig. 9. This inhibition was both dose- and voltage-dependent (data not shown). In addition, we previously found deletion of VDAC, or the adenine nucleotide translocator had no effect on MCC activity in proteoliposomes (Lohret and Kinnally, 1995a; Lohret et al., 1996). Our results indicate the tim23-1 mutation specifically alters the activity of the inner membrane MCC, and has no effect on the outer membrane PSC activity. The Tim23 antibody and the tim23-1 mutation allow discrimination between MCC and PSC activities.


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 tim23-1 mutation does not affect the peptide-sensitive channel, PSC, of the mitochondrial outer membrane. (A)  Sample current traces for PSC from proteoliposomes containing  outer membranes from wild-type or tim23-1 strains are shown in  the presence and absence (control) of 50 μM yCOX-IV1-13 peptide in the bath. (B) The flicker rates of PSC of wild-type and  tim23-1 strains were determined from current traces represented  by those in A above.
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Related In: Results  -  Collection

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Figure 9: The tim23-1 mutation does not affect the peptide-sensitive channel, PSC, of the mitochondrial outer membrane. (A) Sample current traces for PSC from proteoliposomes containing outer membranes from wild-type or tim23-1 strains are shown in the presence and absence (control) of 50 μM yCOX-IV1-13 peptide in the bath. (B) The flicker rates of PSC of wild-type and tim23-1 strains were determined from current traces represented by those in A above.
Mentions: To show that the tim23-1 mutation specifically affected MCC activity, we examined the activity of the outer membrane peptide-sensitive channel, PSC. Like MCC in the inner membrane, the conductance of PSC is transiently blocked by presequence peptides (Henry et al., 1996; Fèvre et al., 1994; Theiffry et al., 1992; Juin et al., 1995). Outer membranes were isolated from mitochondria of wild-type cells and the tim23-1 mutant, and then reconstituted into proteoliposomes. When membrane patches were examined, we found that the PSC activities from both preparations were comparable in terms of conductance, selectivity, and voltage dependence. Furthermore, addition of the yCOX-IV1-13 peptide increased the flicker rate of PSC from both wild-type and tim23-1 strains by ∼10fold as shown by the current traces and histograms of Fig. 9. This inhibition was both dose- and voltage-dependent (data not shown). In addition, we previously found deletion of VDAC, or the adenine nucleotide translocator had no effect on MCC activity in proteoliposomes (Lohret and Kinnally, 1995a; Lohret et al., 1996). Our results indicate the tim23-1 mutation specifically alters the activity of the inner membrane MCC, and has no effect on the outer membrane PSC activity. The Tim23 antibody and the tim23-1 mutation allow discrimination between MCC and PSC activities.

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