Cooperation of translocase complexes in mitochondrial protein import

Most mitochondrial proteins are synthesized in the cytosol and imported into one of the four mitochondrial compartments: outer membrane, intermembrane space, inner membrane, and matrix. Each compartment contains protein complexes that interact with precursor proteins and promote their transport. These translocase complexes do not act as independent units but cooperate with each other and further membrane complexes in a dynamic manner. We propose that a regulated coupling of translocases is important for the coordination of preprotein translocation and efficient sorting to intramitochondrial compartments

Bacterial porin disrupts mitochondrial membrane potential and sensitizes host cells to apoptosis

The bacterial PorB porin, an ATP-binding beta-barrel protein of pathogenic Neisseria gonorrhoeae, triggers host cell apoptosis by an unknown mechanism. PorB is targeted to and imported by host cell mitochondria, causing the breakdown of the mitochondrial membrane potential (delta psi m). Here, we show that PorB induces the condensation of the mitochondrial matrix and the loss of cristae structures, sensitizing cells to the induction of apoptosis via signaling pathways activated by BH3-only proteins. PorB is imported into mitochondria through the general translocase TOM but, unexpectedly, is not recognized by the SAM sorting machinery, usually required for the assembly of beta-barrel proteins in the mitochondrial outer membrane. PorB integrates into the mitochondrial inner membrane, leading to the breakdown of delta psi m. The PorB channel is regulated by nucleotides and an isogenic PorB mutant defective in ATP-binding failed to induce delta psi m loss and apoptosis, demonstrating that dissipation of delta psi m is a requirement for cell death caused by neisserial infection

Eukaryote-wide sequence analysis of mitochondrial β-barrel outer membrane proteins

<p>Abstract</p> <p>Background</p> <p>The outer membranes of mitochondria are thought to be homologous to the outer membranes of Gram negative bacteria, which contain 100's of distinct families of <it>β</it>-barrel membrane proteins (BOMPs) often forming channels for transport of nutrients or drugs. However, only four families of mitochondrial BOMPs (MBOMPs) have been confirmed to date. Although estimates as high as 100 have been made in the past, the number of yet undiscovered MBOMPs is an open question. Fortunately, the recent discovery of a membrane integration signal (the <it>β</it>-signal) for MBOMPs gave us an opportunity to look for undiscovered MBOMPs.</p> <p>Results</p> <p>We present the results of a comprehensive survey of eukaryotic protein sequences intended to identify new MBOMPs. Our search employs recent results on <it>β</it>-signals as well as structural information and a novel BOMP predictor trained on both bacterial and mitochondrial BOMPs. Our principal finding is circumstantial evidence suggesting that few MBOMPs remain to be discovered, if one assumes that, like known MBOMPs, novel MBOMPs will be monomeric and <it>β</it>-signal dependent. In addition to this, our analysis of MBOMP homologs reveals some exceptions to the current model of the <it>β</it>-signal, but confirms its consistent presence in the C-terminal region of MBOMP proteins. We also report a <it>β</it>-signal independent search for MBOMPs against the yeast and Arabidopsis proteomes. We find no good candidates MBOMPs in yeast but the Arabidopsis results are less conclusive.</p> <p>Conclusions</p> <p>Our results suggest there are no remaining MBOMPs left to discover in yeast; and if one assumes all MBOMPs are <it>β</it>-signal dependent, few MBOMP families remain undiscovered in any sequenced organism.</p

The Essentials of Protein Import in the Degenerate Mitochondrion of Entamoeba histolytica

Several essential biochemical processes are situated in mitochondria. The metabolic transformation of mitochondria in distinct lineages of eukaryotes created proteomes ranging from thousands of proteins to what appear to be a much simpler scenario. In the case of Entamoeba histolytica, tiny mitochondria known as mitosomes have undergone extreme reduction. Only recently a single complete metabolic pathway of sulfate activation has been identified in these organelles. The E. histolytica mitosomes do not produce ATP needed for the sulfate activation pathway and for three molecular chaperones, Cpn60, Cpn10 and mtHsp70. The already characterized ADP/ATP carrier would thus be essential to provide cytosolic ATP for these processes, but how the equilibrium of inorganic phosphate could be maintained was unknown. Finally, how the mitosomal proteins are translocated to the mitosomes had remained unclear. We used a hidden Markov model (HMM) based search of the E. histolytica genome sequence to discover candidate (i) mitosomal phosphate carrier complementing the activity of the ADP/ATP carrier and (ii) membrane-located components of the protein import machinery that includes the outer membrane translocation channel Tom40 and membrane assembly protein Sam50. Using in vitro and in vivo systems we show that E. histolytica contains a minimalist set up of the core import components in order to accommodate a handful of mitosomal proteins. The anaerobic and parasitic lifestyle of E. histolytica has produced one of the simplest known mitochondrial compartments of all eukaryotes. Comparisons with mitochondria of another amoeba, Dictystelium discoideum, emphasize just how dramatic the reduction of the protein import apparatus was after the loss of archetypal mitochondrial functions in the mitosomes of E. histolytica

Possible Existence of Lysosome-Like Organella within Mitochondria and Its Role in Mitochondrial Quality Control

The accumulation of unhealthy mitochondria results in mitochondrial dysfunction, which has been implicated in aging, cancer, and a variety of degenerative diseases. However, the mechanism by which mitochondrial quality is regulated remains unclear. Here, we show that Mieap, a novel p53-inducible protein, induces intramitochondrial lysosome-like organella that plays a critical role in mitochondrial quality control. Mieap expression is directly regulated by p53 and is frequently lost in human cancer as result of DNA methylation. Mieap dramatically induces the accumulation of lysosomal proteins within mitochondria and mitochondrial acidic condition without destroying the mitochondrial structure (designated MALM, for Mieap-induced accumulation of lysosome-like organelles within mitochondria) in response to mitochondrial damage. MALM was not related to canonical autophagy. MALM is involved in the degradation of oxidized mitochondrial proteins, leading to increased ATP synthesis and decreased reactive oxygen species generation. These results suggest that Mieap induces intramitochondrial lysosome-like organella that plays a critical role in mitochondrial quality control by eliminating oxidized mitochondrial proteins. Cancer cells might accumulate unhealthy mitochondria due to p53 mutations and/or Mieap methylation, representing a potential cause of the Warburg effect

SARS-Coronavirus Replication Is Supported by a Reticulovesicular Network of Modified Endoplasmic Reticulum

Positive-strand RNA viruses, a large group including human pathogens such as SARS-coronavirus (SARS-CoV), replicate in the cytoplasm of infected host cells. Their replication complexes are commonly associated with modified host cell membranes. Membrane structures supporting viral RNA synthesis range from distinct spherular membrane invaginations to more elaborate webs of packed membranes and vesicles. Generally, their ultrastructure, morphogenesis, and exact role in viral replication remain to be defined. Poorly characterized double-membrane vesicles (DMVs) were previously implicated in SARS-CoV RNA synthesis. We have now applied electron tomography of cryofixed infected cells for the three-dimensional imaging of coronavirus-induced membrane alterations at high resolution. Our analysis defines a unique reticulovesicular network of modified endoplasmic reticulum that integrates convoluted membranes, numerous interconnected DMVs (diameter 200–300 nm), and “vesicle packets” apparently arising from DMV merger. The convoluted membranes were most abundantly immunolabeled for viral replicase subunits. However, double-stranded RNA, presumably revealing the site of viral RNA synthesis, mainly localized to the DMV interior. Since we could not discern a connection between DMV interior and cytosol, our analysis raises several questions about the mechanism of DMV formation and the actual site of SARS-CoV RNA synthesis. Our data document the extensive virus-induced reorganization of host cell membranes into a network that is used to organize viral replication and possibly hide replicating RNA from antiviral defense mechanisms. Together with biochemical studies of the viral enzyme complex, our ultrastructural description of this “replication network” will aid to further dissect the early stages of the coronavirus life cycle and its virus-host interactions

Legionella pneumophila Secretes a Mitochondrial Carrier Protein during Infection

The Mitochondrial Carrier Family (MCF) is a signature group of integral membrane proteins that transport metabolites across the mitochondrial inner membrane in eukaryotes. MCF proteins are characterized by six transmembrane segments that assemble to form a highly-selective channel for metabolite transport. We discovered a novel MCF member, termed Legionella nucleotide carrier Protein (LncP), encoded in the genome of Legionella pneumophila, the causative agent of Legionnaire's disease. LncP was secreted via the bacterial Dot/Icm type IV secretion system into macrophages and assembled in the mitochondrial inner membrane. In a yeast cellular system, LncP induced a dominant-negative phenotype that was rescued by deleting an endogenous ATP carrier. Substrate transport studies on purified LncP reconstituted in liposomes revealed that it catalyzes unidirectional transport and exchange of ATP transport across membranes, thereby supporting a role for LncP as an ATP transporter. A hidden Markov model revealed further MCF proteins in the intracellular pathogens, Legionella longbeachae and Neorickettsia sennetsu, thereby challenging the notion that MCF proteins exist exclusively in eukaryotic organisms