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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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Presequence peptides specifically induce a transient  blockade of MCC conductance. The conductance of a patch excised from a proteoliposome-containing mitochondrial inner  membranes from the wild-type strain was measured at 20 mV.  The single channel current traces in the absence of peptide (control) and in the presence of 50 μM synB2 or yCOX-IV1-13 peptide  in the bath solution were band-width limited to 2 kHz. Total current amplitude diagrams and current traces show the occupancy  of open (O), substate (S) and closed (C) conductance levels. The  probability of occupying the open state was 0.9, 0.8, and 0.4 in  the absence of peptide (control), in the presence of synB2, and in  the presence of yCOX-IV1-13, respectively.
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Figure 3: Presequence peptides specifically induce a transient blockade of MCC conductance. The conductance of a patch excised from a proteoliposome-containing mitochondrial inner membranes from the wild-type strain was measured at 20 mV. The single channel current traces in the absence of peptide (control) and in the presence of 50 μM synB2 or yCOX-IV1-13 peptide in the bath solution were band-width limited to 2 kHz. Total current amplitude diagrams and current traces show the occupancy of open (O), substate (S) and closed (C) conductance levels. The probability of occupying the open state was 0.9, 0.8, and 0.4 in the absence of peptide (control), in the presence of synB2, and in the presence of yCOX-IV1-13, respectively.

Mentions: As shown previously, MCC in the absence of peptide had a predominant transition size of ∼500 pS, a peak conductance of ∼1 nS, a mean open time of ∼25 ms at 20 mV, and a cation selectivity (Lohret and Kinnally, 1995a; see also Figs. 2 and 3). When a peptide based on the first 13 residues of the Saccharomyces cerevisiae cytochrome oxidase subunit IV presequence (yCOX-IV1-13) was added to the bath solution, a transient blockade of MCC conductance was induced during perfusion of the chamber as indicated by the decrease in mean current (Fig. 2 A). While transitions to lower conductance levels were relatively infrequent in the absence of presequence peptide (seen as downward deflections in the current trace of Fig. 2 B), large amplitude, rapid flickering between the open and lower conductance states developed in the current trace during the introduction of yCOX-IV1-13 (Fig. 2 C) and persisted after the perfusion was complete (Fig. 2 D).


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)

Presequence peptides specifically induce a transient  blockade of MCC conductance. The conductance of a patch excised from a proteoliposome-containing mitochondrial inner  membranes from the wild-type strain was measured at 20 mV.  The single channel current traces in the absence of peptide (control) and in the presence of 50 μM synB2 or yCOX-IV1-13 peptide  in the bath solution were band-width limited to 2 kHz. Total current amplitude diagrams and current traces show the occupancy  of open (O), substate (S) and closed (C) conductance levels. The  probability of occupying the open state was 0.9, 0.8, and 0.4 in  the absence of peptide (control), in the presence of synB2, and in  the presence of yCOX-IV1-13, respectively.
© Copyright Policy
Related In: Results  -  Collection

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

Figure 3: Presequence peptides specifically induce a transient blockade of MCC conductance. The conductance of a patch excised from a proteoliposome-containing mitochondrial inner membranes from the wild-type strain was measured at 20 mV. The single channel current traces in the absence of peptide (control) and in the presence of 50 μM synB2 or yCOX-IV1-13 peptide in the bath solution were band-width limited to 2 kHz. Total current amplitude diagrams and current traces show the occupancy of open (O), substate (S) and closed (C) conductance levels. The probability of occupying the open state was 0.9, 0.8, and 0.4 in the absence of peptide (control), in the presence of synB2, and in the presence of yCOX-IV1-13, respectively.
Mentions: As shown previously, MCC in the absence of peptide had a predominant transition size of ∼500 pS, a peak conductance of ∼1 nS, a mean open time of ∼25 ms at 20 mV, and a cation selectivity (Lohret and Kinnally, 1995a; see also Figs. 2 and 3). When a peptide based on the first 13 residues of the Saccharomyces cerevisiae cytochrome oxidase subunit IV presequence (yCOX-IV1-13) was added to the bath solution, a transient blockade of MCC conductance was induced during perfusion of the chamber as indicated by the decrease in mean current (Fig. 2 A). While transitions to lower conductance levels were relatively infrequent in the absence of presequence peptide (seen as downward deflections in the current trace of Fig. 2 B), large amplitude, rapid flickering between the open and lower conductance states developed in the current trace during the introduction of yCOX-IV1-13 (Fig. 2 C) and persisted after the perfusion was complete (Fig. 2 D).

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