How finicky is mitochondrial protein import?

Recently it was reported that artificial targeting signals or signals specific for organelles other than mitochondria could direct proteins into mitochondria. Here we discuss findings which suggest that specific steps of mitochondrial protein import can be bypassed. Non-specific targeting signals appear to use this bypass pathway. Such import occurs at very low rates under physiological conditions and therefore does not affect the uniqueness of mitochondrial protein composition

Contact sites between inner and outer membranes

Contact sites between both mitochondrial membranes play a predominant role in the transport of nuclear-coded precursor proteins into mitochondria. The characterization of contact sites was greatly advanced by the reversible accumulation of precursor proteins in transit (translocation intermediates). It was found that the sites are saturable, apparently contain proteinaceous components and mediate extensive unfolding of the polypeptide chain in translocation. Some components of mitochondrial contact sites are currently being identified

A mitochondrial import receptor for the ADP/ATP carrier

We have identified a mitochondrial outer membrane protein of 72 kd (MOM72) that exhibits the properties of an import receptor for the ADP/ATP carrier (AAC), the most abundant mitochondrial protein. Monospecific antibodies and Fab fragments against MOM72 selectively inhibit import of AAC at the level of specific binding to the mitochondria. AAC bound to the mitochondrial surface is coprecipitated with antibodies against MOM72 after lysis of mitochondria with detergent. MOM72 thus has a complementary function to that of MOM19, which acts as an import receptor for the majority of mitochondrial proteins studied so far but not for the AAC. The import pathway of the precursor of MOM72 appears to involve MOM19 as receptor

Precursor proteins in transit through mitochondrial contact sites interact with hsp70 in the matrix

We previously reported that hsp70 in the mitochondrial matrix (mt-hsp70 = Ssc1p) is required for import of precursor proteins destined for the matrix or intermembrane space. Here we show that mt-hsp70 is also needed for the import of mitochondrial inner membrane proteins. In particular, the precursor of ADP/ATP carrier that is known not to interact with hsp60 on its assembly pathway requires functional mt-hsp70 for import, suggesting a general role of mt-hsp70 in membrane translocation of precursors. Moreover, a precursor arrested in contact sites was specifically co-precipitated with antibodies directed against mt-hsp70. We conclude that mt-hsp70 is directly involved in the translocation of many, if not all, precursor proteins that are transported across the inner membrane

Mitochondrial import receptors for precursor proteins

The specific targeting of precursor proteins synthesized in the cytosol to various cell organelles is a central aspect of intracellular protein traffic. Several hundred different proteins are imported from the cytosol into the mitochondria. Recent studies have identified the mitochondrial outer membrane proteins MOM19, MOM72, MOM38 (≈ISP42) and p32 which have a role in initial steps of protein import. The first three components are present in a multi-subunit complex that catalyses recognition and membrane insertion of precursor proteins

Transport of proteins into mitochondria

The transfer of cytoplasmically synthesized precursor proteins into or across the inner mitochondrial membrane is dependent on energization of the membrane. To investigate the role of this energy requirement, a buffer system was developed in which efficient import of ADP/ATP carrier into mitochondria from the receptor-bound state occurred. This import was rapid and was dependent on divalent cations, whereas the binding of precursor proteins to the mitochondrial surface was slow and was independent of added divalent cations. Using this buffer system, the import of ADP/ATP carrier could be driven by a valinomycin-induced potassium diffusion potential. The protonophore carbonylcyanide m-chlorophenyl-hydrazone was not able to abolish this import. Imposition of a delta pH did not stimulate the import. We conclude that the membrane potential delta psi itself and not the total protonmotive force delta p is the required energy source