Tim23–Tim50 pair coordinates functions of translocators and motor proteins in mitochondrial protein import

Department of Chemistry, Graduate School of Science, Nagoya University, Nagoya, Japan.
The Journal of Cell Biology (Impact Factor: 9.83). 02/2009; 184(1):129-41. DOI: 10.1083/jcb.200808068
Source: PubMed


Mitochondrial protein traffic requires coordinated operation of protein translocator complexes in the mitochondrial membrane. The TIM23 complex translocates and inserts proteins into the mitochondrial inner membrane. Here we analyze the intermembrane space (IMS) domains of Tim23 and Tim50, which are essential subunits of the TIM23 complex, in these functions. We find that interactions of Tim23 and Tim50 in the IMS facilitate transfer of precursor proteins from the TOM40 complex, a general protein translocator in the outer membrane, to the TIM23 complex. Tim23-Tim50 interactions also facilitate a late step of protein translocation across the inner membrane by promoting motor functions of mitochondrial Hsp70 in the matrix. Therefore, the Tim23-Tim50 pair coordinates the actions of the TOM40 and TIM23 complexes together with motor proteins for mitochondrial protein import.

    • "''pulls'' the DHFR moiety so close to the TOM complex that proteinase K cannot access the more flexible b 2 portion of the preprotein (Schwarz et al., 1993; Frazier et al., 2004; van der Laan et al., 2005; Krayl et al., 2007; Tamura et al., 2009, Chacinska et al., 2010; Schulz and Rehling, 2014). Unexpectedly, this ''pulling assay'' yielded a split result for mitochondria lacking Mgr2. "
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    ABSTRACT: The majority of preproteins destined for mitochondria carry N-terminal presequences. The presequence translocase of the inner mitochondrial membrane (TIM23 complex) plays a central role in protein sorting. Preproteins are either translocated through the TIM23 complex into the matrix or are laterally released into the inner membrane. We report that the small hydrophobic protein Mgr2 controls the lateral release of preproteins. Mgr2 interacts with preproteins in transit through the TIM23 complex. Overexpression of Mgr2 delays preprotein release, whereas a lack of Mgr2 promotes preprotein sorting into the inner membrane. Preproteins with a defective inner membrane sorting signal are translocated into the matrix in wild-type mitochondria but are released into the inner membrane in Mgr2-deficient mitochondria. We conclude that Mgr2 functions as a lateral gatekeeper of the mitochondrial presequence translocase, providing quality control for the membrane sorting of preproteins. Copyright © 2014 Elsevier Inc. All rights reserved.
    No preview · Article · Nov 2014 · Molecular cell
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    • "HA -mitochondria (Figure 1D, white circle). Therefore, we assigned these crosslinks to Tim23–Tim50-adducts previously identified in other studies (Alder et al, 2008; Tamura et al, 2009). The 80-kDa Tim50-adduct was not affected in mitochondria with tagged versions of Tim23, but was selectively shifted in Tim21 ProtA -mitochondria (Figure 1D, black circle). "
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    ABSTRACT: The mitochondrial presequence translocase interacts with presequence-containing precursors at the intermembrane space (IMS) side of the inner membrane to mediate their translocation into the matrix. Little is known as too how these matrix-targeting signals activate the translocase in order to initiate precursor transport. Therefore, we analysed how signal recognition by the presequence translocase initiates reorganization among Tim-proteins during import. Our analyses revealed that the presequence receptor Tim50 interacts with Tim21 in a signal-sensitive manner in a process that involves the IMS-domain of the Tim23 channel. The signal-driven release of Tim21 from Tim50 promotes recruitment of Pam17 and thus triggers formation of the motor-associated form of the TIM23 complex required for matrix transport.
    Full-text · Article · Feb 2013 · The EMBO Journal
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    • "Pam18, a 168–amino acid protein, has, in addition to a C-terminal matrix-localized J-domain, a single membrane-spanning region and an N-terminal domain in the intermembrane space (IMS) (Figure 1A). This IMS domain interacts with Tim17, as well as Tim23, a core component of the translocon itself, and stabilizes Pam18's association with the translocon (D'Silva et al., 2003, 2008; Truscott et al., 2003; Chacinska et al., 2005; Tamura et al., 2009). In addition, Pam18's J-domain interacts with Pam16 (also called Tim16), a 149–amino acid protein whose N-terminus associates with the translocon , probably indirectly, via interaction with Tim44 (Figure 1A; Frazier et al., 2004; Kozany et al., 2004; Mokranjac et al., 2007; D'Silva et al., 2008; Hutu et al., 2008). "
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    ABSTRACT: The heat-shock protein 70 (Hsp70)-based import motor, associated with the translocon on the matrix side of the mitochondrial inner membrane, drives translocation of proteins via cycles of binding and release. Stimulation of Hsp70's ATPase activity by the translocon-associated J-protein Pam18 is critical for this process. Pam18 forms a heterodimer with the structurally related protein Pam16, via their J-type domains. This interaction has been proposed to perform a critical regulatory function, inhibiting the ATPase stimulatory activity of Pam18. Using biochemical and genetic assays, we tested this hypothesis by assessing the in vivo function of Pam18 variants having altered abilities to stimulate Hsp70's ATPase activity. The observed pattern of genetic interactions was opposite from that predicted if the heterodimer serves an inhibitory function; instead the pattern was consistent with that of mutations known to cause reduction in the stability of the heterodimer. Analysis of a previously uncharacterized region of Pam16 revealed its requirement for formation of an active Pam18:Pam16 complex able to stimulate Hsp70's ATPase activity. Together, our data are consistent with the idea that Pam18 and Pam16 form a stable heterodimer and that the critical role of the Pam18:Pam16 interaction is the physical tethering of Pam18 to the translocon via its interaction with Pam16.
    Full-text · Article · Dec 2011 · Molecular biology of the cell
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