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Science (New York, N.Y.) Sep 2021β barrel outer membrane proteins (β-OMPs) play vital roles in mitochondria, chloroplasts, and Gram-negative bacteria. Evolutionarily conserved complexes such as the...
β barrel outer membrane proteins (β-OMPs) play vital roles in mitochondria, chloroplasts, and Gram-negative bacteria. Evolutionarily conserved complexes such as the mitochondrial sorting and assembly machinery (SAM) mediate the assembly of β-OMPs. We investigated the SAM-mediated assembly of the translocase of the outer membrane (TOM) core complex. Cryo–electron microscopy structures of SAM–fully folded Tom40 and the SAM-Tom40/Tom5/Tom6 complexes at ~3-angstrom resolution reveal that Sam37 stabilizes the mature Tom40 mainly through electrostatic interactions, thus facilitating subsequent TOM assembly. These results support the β barrel switching model and provide structural insights into the assembly and release of β barrel complexes.
Topics: Carrier Proteins; Cell Line; Cryoelectron Microscopy; Humans; Membrane Proteins; Mitochondrial Membrane Transport Proteins; Mitochondrial Membranes; Mitochondrial Precursor Protein Import Complex Proteins; Models, Molecular; Multiprotein Complexes; Protein Conformation; Protein Folding; Protein Structure, Secondary; Protein Subunits; Protein Transport; Saccharomyces cerevisiae Proteins; Static Electricity
PubMed: 34446444
DOI: 10.1126/science.abh0704 -
Methods in Molecular Biology (Clifton,... 2024Mitochondrial β-barrel proteins fulfill crucial roles in the biogenesis and function of the cell organelle. They mediate the import and membrane insertion of proteins...
Mitochondrial β-barrel proteins fulfill crucial roles in the biogenesis and function of the cell organelle. They mediate the import and membrane insertion of proteins and transport of small metabolites and ions. All β-barrel proteins are made as precursors on cytosolic ribosomes and are imported into mitochondria. The β-barrel proteins fold and assemble with partner proteins in the outer membrane. The in vitro import of radiolabelled proteins into isolated mitochondria is a powerful tool to investigate the import of β-barrel proteins, the folding of the β-barrel proteins, and their assembly into protein complexes. Altogether, the in vitro import assay is a versatile and crucial assay to analyze the mechanisms of the biogenesis of mitochondrial β-barrel proteins.
Topics: Mitochondrial Proteins; Saccharomyces cerevisiae Proteins; Saccharomyces cerevisiae; Mitochondria; Protein Transport; Mitochondrial Membrane Transport Proteins
PubMed: 38478280
DOI: 10.1007/978-1-0716-3734-0_13 -
Trends in Cell Biology Sep 2019Despite the progress in understanding the molecular responses to mitochondrial damage, responses to aberrant accumulation of mitochondrial precursor proteins and... (Review)
Review
Despite the progress in understanding the molecular responses to mitochondrial damage, responses to aberrant accumulation of mitochondrial precursor proteins and mitochondrial import defects remain poorly understood. Recent work (Mårtensson et al., Nature, 2019) has unveiled a pathway similar to endoplasmic-reticulum-associated degradation (ERAD) in fine-tuning the fidelity of translocase of the outer mitochondrial membrane (TOM) complex-mediated mitochondrial import.
Topics: Carrier Proteins; Endoplasmic Reticulum-Associated Degradation; Mitochondrial Membrane Transport Proteins; Mitochondrial Membranes; Mitochondrial Precursor Protein Import Complex Proteins; Mitochondrial Proteins; Protein Transport; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins
PubMed: 31358413
DOI: 10.1016/j.tcb.2019.07.003 -
Pharmacological Research Oct 2020SAM50, a 7-8 nm diameter β-barrel channel of the mitochondrial outer membrane, is the central channel of the sorting and assembly machinery (SAM) complex involved in... (Review)
Review
SAM50, a 7-8 nm diameter β-barrel channel of the mitochondrial outer membrane, is the central channel of the sorting and assembly machinery (SAM) complex involved in the biogenesis of β-barrel proteins. Interestingly, SAM50 is not known to have channel translocase activity; however, we have recently found that this channel is necessary and sufficient for mitochondrial entry of cytotoxic proteases. Cytotoxic lymphocytes eliminate cells that pose potential hazards, such as virus- and bacteria-infected cells as well as cancer cells. They induce cell death following the delivery of granzyme cytotoxic proteases into the cytosol of the target cell. Although granzyme A and granzyme B (GA and GB), the best characterized of the five human granzymes, trigger very distinct apoptotic cascades, they share the ability to directly target the mitochondria. GA and GB do not have a mitochondrial targeting signal, yet they enter the target cell mitochondria to disrupt respiratory chain complex I and induce mitochondrial reactive oxygen species (ROS)-dependent cell death. We found that granzyme mitochondrial entry requires SAM50 and the translocase of the inner membrane 22 (TIM22). Preventing granzymes' mitochondrial entry compromises their cytotoxicity, indicating that this event is unexpectedly an important step for cell death. Although mitochondria are best known for their roles in cell metabolism and energy conversion, these double-membrane organelles are also involved in Ca homeostasis, metabolite transport, cell cycle regulation, cell signaling, differentiation, stress response, redox homeostasis, aging, and cell death. This multiplicity of functions is matched with the complexity and plasticity of the mitochondrial proteome as well as the organelle's morphological and structural versatility. Indeed, mitochondria are extremely dynamic and undergo fusion and fission events in response to diverse cellular cues. In humans, there are 1500 different mitochondrial proteins, the vast majority of which are encoded in the nuclear genome and translated by cytosolic ribosomes, after which they must be imported and properly addressed to the right mitochondrial compartment. To this end, mitochondria are equipped with a very sophisticated and highly specific protein import machinery. The latter is centered on translocase complexes embedded in the outer and inner mitochondrial membranes working along five different import pathways. We will briefly describe these import pathways to put into perspective our finding regarding the ability of granzymes to enter the mitochondria.
Topics: Animals; Humans; Membrane Proteins; Mitochondria; Mitochondrial Membranes; Mitochondrial Precursor Protein Import Complex Proteins; Mitochondrial Proteins; Peptide Hydrolases; T-Lymphocytes, Cytotoxic
PubMed: 32919042
DOI: 10.1016/j.phrs.2020.105196 -
Journal of Cell Science Sep 2023Chloroplasts conduct photosynthesis and numerous metabolic and signalling processes that enable plant growth and development. Most of the ∼3000 proteins in...
Chloroplasts conduct photosynthesis and numerous metabolic and signalling processes that enable plant growth and development. Most of the ∼3000 proteins in chloroplasts are nucleus encoded and must be imported from the cytosol. Thus, the protein import machinery of the organelle (the TOC-TIC apparatus) is of fundamental importance for chloroplast biogenesis and operation. Cytosolic factors target chloroplast precursor proteins to the TOC-TIC apparatus, which drives protein import across the envelope membranes into the organelle, before various internal systems mediate downstream routing to different suborganellar compartments. The protein import system is proteolytically regulated by the ubiquitin-proteasome system (UPS), enabling centralized control over the organellar proteome. In addition, the UPS targets a range of chloroplast proteins directly. In this Cell Science at a Glance article and the accompanying poster, we present mechanistic details of these different chloroplast protein targeting and translocation events, and of the UPS systems that regulate chloroplast proteins.
Topics: Ubiquitin; Chloroplasts; Photosynthesis; Proteasome Endopeptidase Complex; Chloroplast Proteins; Protein Transport
PubMed: 37732520
DOI: 10.1242/jcs.241125 -
FEBS Letters Sep 2019Protein transport into the mammalian endoplasmic reticulum (ER) used to be seen as strictly cotranslational, that is temporarily and mechanistically coupled to protein... (Review)
Review
Protein transport into the mammalian endoplasmic reticulum (ER) used to be seen as strictly cotranslational, that is temporarily and mechanistically coupled to protein synthesis. In the course of the last decades, however, several classes of precursors of soluble and membrane proteins were found to be post-translationally imported into the ER, without any involvement of the ribosome. The first such class to be identified were the small presecretory proteins; tail-anchored membrane proteins followed next. In both classes, the inherent address tag is released from the translating ribosome before the initiation of ER import, as part of the fully synthesized precursor. In small presecretory proteins, the information for ER targeting and -translocation via the polypeptide-conducting Sec61-channel is encoded by a classical N-terminal signal peptide, which is released from the ribsosome before targeting due to the small size of the full-length precursor. Here, we discuss the current state of research on targeting and translocation of small presecretory proteins into the mammalian ER. In closing, we present a unifying hypothesis for ER protein translocation in terms of an energy diagram for Sec61-channel gating.
Topics: Endoplasmic Reticulum; Humans; Membrane Proteins; Protein Transport
PubMed: 31325177
DOI: 10.1002/1873-3468.13542 -
Trends in Cell Biology May 2024Peroxisomes are vital metabolic organelles that import their lumenal (matrix) enzymes from the cytosol using mobile receptors. Surprisingly, the receptors can even... (Review)
Review
Peroxisomes are vital metabolic organelles that import their lumenal (matrix) enzymes from the cytosol using mobile receptors. Surprisingly, the receptors can even import folded proteins, but the underlying mechanism has been a mystery. Recent results reveal how import receptors shuttle cargo into peroxisomes. The cargo-bound receptors move from the cytosol across the peroxisomal membrane completely into the matrix by a mechanism that resembles transport through the nuclear pore. The receptors then return to the cytosol through a separate retrotranslocation channel, leaving the cargo inside the organelle. This cycle concentrates imported proteins within peroxisomes, and the energy for cargo import is supplied by receptor export. Peroxisomal protein import thus fundamentally differs from other previously known mechanisms for translocating proteins across membranes.
Topics: Peroxisomes; Protein Transport; Humans; Animals; Cytosol; Receptors, Cytoplasmic and Nuclear
PubMed: 37743160
DOI: 10.1016/j.tcb.2023.08.005 -
Proceedings of the National Academy of... May 2024Targeting proteins to specific subcellular destinations is essential in prokaryotes, eukaryotes, and the viruses that infect them. Chimalliviridae phages encapsulate...
Targeting proteins to specific subcellular destinations is essential in prokaryotes, eukaryotes, and the viruses that infect them. Chimalliviridae phages encapsulate their genomes in a nucleus-like replication compartment composed of the protein chimallin (ChmA) that excludes ribosomes and decouples transcription from translation. These phages selectively partition proteins between the phage nucleus and the bacterial cytoplasm. Currently, the genes and signals that govern selective protein import into the phage nucleus are unknown. Here, we identify two components of this protein import pathway: a species-specific surface-exposed region of a phage intranuclear protein required for nuclear entry and a conserved protein, PicA (Protein importer of chimalliviruses A), that facilitates cargo protein trafficking across the phage nuclear shell. We also identify a defective cargo protein that is targeted to PicA on the nuclear periphery but fails to enter the nucleus, providing insight into the mechanism of nuclear protein trafficking. Using CRISPRi-ART protein expression knockdown of PicA, we show that PicA is essential early in the chimallivirus replication cycle. Together, our results allow us to propose a multistep model for the Protein Import Chimallivirus pathway, where proteins are targeted to PicA by amino acids on their surface and then licensed by PicA for nuclear entry. The divergence in the selectivity of this pathway between closely related chimalliviruses implicates its role as a key player in the evolutionary arms race between competing phages and their hosts.
Topics: Viral Proteins; Bacteriophages; Protein Transport; Cell Nucleus; Virus Replication
PubMed: 38687783
DOI: 10.1073/pnas.2321190121 -
Annual Review of Biochemistry Jun 2022Mitochondria are central to energy production, metabolism and signaling, and apoptosis. To make new mitochondria from preexisting mitochondria, the cell needs to import... (Review)
Review
Mitochondria are central to energy production, metabolism and signaling, and apoptosis. To make new mitochondria from preexisting mitochondria, the cell needs to import mitochondrial proteins from the cytosol into the mitochondria with the aid of translocators in the mitochondrial membranes. The translocase of the outer membrane (TOM) complex, an outer membrane translocator, functions as an entry gate for most mitochondrial proteins. Although high-resolution structures of the receptor subunits of the TOM complex were deposited in the early 2000s, those of entire TOM complexes became available only in 2019. The structural details of these TOM complexes, consisting of the dimer of the β-barrel import channel Tom40 and four α-helical membrane proteins, revealed the presence of several distinct paths and exits for the translocation of over 1,000 different mitochondrial precursor proteins. High-resolution structures of TOM complexes now open up a new era of studies on the structures, functions, and dynamics of the mitochondrial import system.
Topics: Carrier Proteins; Mitochondria; Mitochondrial Membrane Transport Proteins; Mitochondrial Precursor Protein Import Complex Proteins; Mitochondrial Proteins; Protein Transport; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins
PubMed: 35287471
DOI: 10.1146/annurev-biochem-032620-104527 -
Biochimica Et Biophysica Acta May 2016Peroxisomes are organelles that play an important role in many cellular tasks. The functionality of peroxisomes depends on the proper import of their matrix proteins.... (Review)
Review
Peroxisomes are organelles that play an important role in many cellular tasks. The functionality of peroxisomes depends on the proper import of their matrix proteins. Peroxisomal matrix proteins are imported posttranslationally in a folded, sometimes even oligomeric state. They harbor a peroxisomal targeting sequence (PTS), which is recognized by dynamic PTS-receptors in the cytosol. The PTS-receptors ferry the cargo to the peroxisomal membrane, where they become part of a transient import pore and then release the cargo into the peroxisomal lumen. Subsequentially, the PTS-receptors are ubiquitinated in order to mark them for the export-machinery, which releases them back to the cytosol. Upon deubiquitination, the PTS-receptors can facilitate further rounds of cargo import. Because the ubiquitination of the receptors is an essential step in the import cycle, it also represents a central regulatory element that governs peroxisomal dynamics. In this review we want to give an introduction to the functional role played by ubiquitination during peroxisomal protein import and highlight the mechanistic concepts that have emerged based on data derived from different species since the discovery of the first ubiquitinated peroxin 15years ago. Moreover, we discuss future tasks and the potential of using advanced technologies for investigating further details of peroxisomal protein transport.
Topics: ATPases Associated with Diverse Cellular Activities; Adenosine Triphosphatases; Animals; Eukaryotic Cells; Gene Expression Regulation; Humans; Membrane Proteins; Peroxisomes; Plants; Protein Isoforms; Protein Structure, Tertiary; Protein Transport; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins; Signal Transduction; Ubiquitin; Ubiquitination
PubMed: 26367801
DOI: 10.1016/j.bbamcr.2015.09.010