The Molecular Biodiversity of Protein Targeting and Protein Transport Related to the Endoplasmic Reticulum

Looking at the variety of the thousands of different polypeptides that have been focused on in the research on the endoplasmic reticulum from the last five decades taught us one humble lesson: no one size fits all. Cells use an impressive array of components to enable the safe transport of protein cargo from the cytosolic ribosomes to the endoplasmic reticulum. Safety during the transit is warranted by the interplay of cytosolic chaperones, membrane receptors, and protein translocases that together form functional networks and serve as protein targeting and translocation routes. While two targeting routes to the endoplasmic reticulum, SRP (signal recognition particle) and GET (guided entry of tail-anchored proteins), prefer targeting determinants at the N- and C-terminus of the cargo polypeptide, respectively, the recently discovered SND (SRP-independent) route seems to preferentially cater for cargos with non-generic targeting signals that are less hydrophobic or more distant from the termini. With an emphasis on targeting routes and protein translocases, we will discuss those functional networks that drive efficient protein topogenesis and shed light on their redundant and dynamic nature in health and disease.

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NAC functions as a modulator of SRP during the early steps of protein targeting to the endoplasmic reticulum

Molecular Biology of the Cell ◽

10.1091/mbc.e12-02-0112 ◽

2012 ◽

Vol 23 (16) ◽

pp. 3027-3040 ◽

Cited By ~ 41

Author(s):

Ying Zhang ◽

Uta Berndt ◽

Hanna Gölz ◽

Arlette Tais ◽

Stefan Oellerer ◽

...

Keyword(s):

Endoplasmic Reticulum ◽

Protein Targeting ◽

Signal Sequence ◽

Signal Recognition Particle ◽

Signal Recognition ◽

Signal Sequences ◽

Chain Complexes ◽

Ribosomal Tunnel ◽

Nascent Chain ◽

Combined Data

Nascent polypeptide-associated complex (NAC) was initially found to bind to any segment of the nascent chain except signal sequences. In this way, NAC is believed to prevent mistargeting due to binding of signal recognition particle (SRP) to signalless ribosome nascent chain complexes (RNCs). Here we revisit the interplay between NAC and SRP. NAC does not affect SRP function with respect to signalless RNCs; however, NAC does affect SRP function with respect to RNCs targeted to the endoplasmic reticulum (ER). First, early recruitment of SRP to RNCs containing a signal sequence within the ribosomal tunnel is NAC dependent. Second, NAC is able to directly and tightly bind to nascent signal sequences. Third, SRP initially displaces NAC from RNCs; however, when the signal sequence emerges further, trimeric NAC·RNC·SRP complexes form. Fourth, upon docking to the ER membrane NAC remains bound to RNCs, allowing NAC to shield cytosolically exposed nascent chain domains not only before but also during cotranslational translocation. The combined data indicate a functional interplay between NAC and SRP on ER-targeted RNCs, which is based on the ability of the two complexes to bind simultaneously to distinct segments of a single nascent chain.

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Promiscuous targeting of polytopic membrane proteins to SecYEG or YidC by the Escherichia coli signal recognition particle

Molecular Biology of the Cell ◽

10.1091/mbc.e11-07-0590 ◽

2012 ◽

Vol 23 (3) ◽

pp. 464-479 ◽

Cited By ~ 44

Author(s):

Thomas Welte ◽

Renuka Kudva ◽

Patrick Kuhn ◽

Lukas Sturm ◽

David Braig ◽

...

Keyword(s):

Membrane Proteins ◽

Ribosomal Proteins ◽

Protein Transport ◽

Integration Site ◽

Signal Recognition Particle ◽

Cross Linking ◽

Secretory Proteins ◽

Signal Recognition ◽

Protein Insertion ◽

C Terminus

Protein insertion into the bacterial inner membrane is facilitated by SecYEG or YidC. Although SecYEG most likely constitutes the major integration site, small membrane proteins have been shown to integrate via YidC. We show that YidC can also integrate multispanning membrane proteins such as mannitol permease or TatC, which had been considered to be exclusively integrated by SecYEG. Only SecA-dependent multispanning membrane proteins strictly require SecYEG for integration, which suggests that SecA can only interact with the SecYEG translocon, but not with the YidC insertase. Targeting of multispanning membrane proteins to YidC is mediated by signal recognition particle (SRP), and we show by site-directed cross-linking that the C-terminus of YidC is in contact with SRP, the SRP receptor, and ribosomal proteins. These findings indicate that SRP recognizes membrane proteins independent of the downstream integration site and that many membrane proteins can probably use either SecYEG or YidC for integration. Because protein synthesis is much slower than protein transport, the use of YidC as an additional integration site for multispanning membrane proteins may prevent a situation in which the majority of SecYEG complexes are occupied by translating ribosomes during cotranslational insertion, impeding the translocation of secretory proteins.

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SRPassing Co-translational Targeting: The Role of the Signal Recognition Particle in Protein Targeting and mRNA Protection

International Journal of Molecular Sciences ◽

10.3390/ijms22126284 ◽

2021 ◽

Vol 22 (12) ◽

pp. 6284

Author(s):

Morgana K. Kellogg ◽

Sarah C. Miller ◽

Elena B. Tikhonova ◽

Andrey L. Karamyshev

Keyword(s):

Endoplasmic Reticulum ◽

Noncoding Rna ◽

Protein Targeting ◽

Signal Recognition Particle ◽

Structure And Function ◽

Secretory Proteins ◽

Signal Recognition ◽

Domains Of Life ◽

And Function

Signal recognition particle (SRP) is an RNA and protein complex that exists in all domains of life. It consists of one protein and one noncoding RNA in some bacteria. It is more complex in eukaryotes and consists of six proteins and one noncoding RNA in mammals. In the eukaryotic cytoplasm, SRP co-translationally targets proteins to the endoplasmic reticulum and prevents misfolding and aggregation of the secretory proteins in the cytoplasm. It was demonstrated recently that SRP also possesses an earlier unknown function, the protection of mRNAs of secretory proteins from degradation. In this review, we analyze the progress in studies of SRPs from different organisms, SRP biogenesis, its structure, and function in protein targeting and mRNA protection.

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Emerging View on the Molecular Functions of Sec62 and Sec63 in Protein Translocation

International Journal of Molecular Sciences ◽

10.3390/ijms222312757 ◽

2021 ◽

Vol 22 (23) ◽

pp. 12757

Author(s):

Sung-jun Jung ◽

Hyun Kim

Keyword(s):

Endoplasmic Reticulum ◽

Protein Targeting ◽

Protein Translocation ◽

Signal Recognition Particle ◽

Signal Recognition ◽

Conducting Channel ◽

Evolutionarily Conserved ◽

Translocation Pathway ◽

Molecular Functions ◽

Sec61 Channel

Most secreted and membrane proteins are targeted to and translocated across the endoplasmic reticulum (ER) membrane through the Sec61 protein-conducting channel. Evolutionarily conserved Sec62 and Sec63 associate with the Sec61 channel, forming the Sec complex and mediating translocation of a subset of proteins. For the last three decades, it has been thought that ER protein targeting and translocation occur via two distinct pathways: signal recognition particle (SRP)-dependent co-translational or SRP-independent, Sec62/Sec63 dependent post-translational translocation pathway. However, recent studies have suggested that ER protein targeting and translocation through the Sec translocon are more intricate than previously thought. This review summarizes the current understanding of the molecular functions of Sec62/Sec63 in ER protein translocation.

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The beta subunit of the signal recognition particle receptor is a transmembrane GTPase that anchors the alpha subunit, a peripheral membrane GTPase, to the endoplasmic reticulum membrane.

The Journal of Cell Biology ◽

10.1083/jcb.128.3.273 ◽

1995 ◽

Vol 128 (3) ◽

pp. 273-282 ◽

Cited By ~ 76

Author(s):

J D Miller ◽

S Tajima ◽

L Lauffer ◽

P Walter

Keyword(s):

Endoplasmic Reticulum ◽

Protein Targeting ◽

Signal Sequence ◽

Transmembrane Protein ◽

Signal Recognition Particle ◽

Endoplasmic Reticulum Membrane ◽

Protein Chain ◽

Signal Recognition ◽

Signal Recognition Particle Receptor ◽

Er Membrane

The signal recognition particle receptor (SR) is required for the cotranslational targeting of both secretory and membrane proteins to the endoplasmic reticulum (ER) membrane. During targeting, the SR interacts with the signal recognition particle (SRP) which is bound to the signal sequence of the nascent protein chain. This interaction catalyzes the GTP-dependent transfer of the nascent chain from SRP to the protein translocation apparatus in the ER membrane. The SR is a heterodimeric protein comprised of a 69-kD subunit (SR alpha) and a 30-kD subunit (SR beta) which are associated with the ER membrane in an unknown manner. SR alpha and the 54-kD subunits of SRP (SRP54) each contain related GTPase domains which are required for SR and SRP function. Molecular cloning and sequencing of a cDNA encoding SR beta revealed that SR beta is a transmembrane protein and, like SR alpha and SRP54, is a member of the GTPase superfamily. Although SR beta defines its own GTPase subfamily, it is distantly related to ARF and Sar1. Using UV cross-linking, we confirm that SR beta binds GTP specifically. Proteolytic digestion experiments show that SR alpha is required for the interaction of SRP with SR. SR alpha appears to be peripherally associated with the ER membrane, and we suggest that SR beta, as an integral membrane protein, mediates the membrane association of SR alpha. The discovery of its guanine nucleotide-binding domain, however, makes it likely that its role is more complex than that of a passive anchor for SR alpha. These findings suggest that a cascade of three directly interacting GTPases functions during protein targeting to the ER membrane.

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Getting to the membrane: how is co-translational protein targeting to the endoplasmic reticulum regulated?

Biochemical Society Transactions ◽

10.1042/bst0311232 ◽

2003 ◽

Vol 31 (6) ◽

pp. 1232-1237 ◽

Cited By ~ 5

Author(s):

M.R. Pool

Keyword(s):

Endoplasmic Reticulum ◽

Protein Targeting ◽

Signal Recognition Particle ◽

Signal Recognition ◽

Cognate Receptor

Co-translational protein targeting to the endoplasmic reticulum requires the signal-recognition particle, its cognate receptor and the translocation channel. Co-ordination of the targeting process necessitates the interaction of these components with each other and with the ribosome. These interactions are regulated by three GTPases, which act in concert, ensuring the fidelity of the targeting process.

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The role of molecular chaperones in protein transport into the endoplasmic reticulum

Philosophical Transactions of the Royal Society B Biological Sciences ◽

10.1098/rstb.1993.0032 ◽

1993 ◽

Vol 339 (1289) ◽

pp. 335-341 ◽

Cited By ~ 9

Keyword(s):

Endoplasmic Reticulum ◽

Molecular Chaperones ◽

Protein Transport ◽

Protein Translocation ◽

Signal Recognition Particle ◽

Microsomal Membrane ◽

Nucleoside Triphosphate ◽

Secretory Proteins ◽

Hsc 70 ◽

Hydrolysis Of

In eukaryotic cells export of the vast majority of newly synthesized secretory proteins is initiated at the level of the membrane of the endoplasmic reticulum (microsomal membrane). The precursors of secretory proteins are not transported across the microsomal m em brane in their native state. Typically, signal peptides in the precursor proteins are involved in preserving the transport-competent state. Furthermore, there are two alternatively acting mechanisms involved in preserving transport competence in the cytosol. The first mechanism involves two ribonucleoparticles (ribosome and signal recognition particle) and their receptors on the microsomal surface and requires the hydrolysis of GTP. The second mechanism does not involve ribonucleoparticles and their receptors but depends on the hydrolysis of A TP and on cA-acting molecular chaperones, such as heat shock cognate protein 70 (hsc 70). In both mechanisms a translocase in the microsomal membrane mediates protein translocation. This translocase includes a signal peptide receptor on the cu-side of the microsomal membrane and a component that also depends on the hydrolysis of ATP. At least in certain cases, an additional nucleoside triphosphate-requiring step is involved which is related to the trans -acting molecular chaperone BiP.

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Vestiges of the bacterial signal recognition particle-based protein targeting in mitochondria

Molecular Biology and Evolution ◽

10.1093/molbev/msab090 ◽

2021 ◽

Author(s):

Jan Pyrih ◽

Tomáš Pánek ◽

Ignacio Miguel Durante ◽

Vendula Rašková ◽

Kristýna Cimrhanzlová ◽

...

Keyword(s):

Endoplasmic Reticulum ◽

Membrane Protein ◽

Signal Peptide ◽

Protein Targeting ◽

Fluorescent Protein ◽

Mitochondrial Protein ◽

Signal Recognition Particle ◽

Signal Recognition ◽

Mitochondrial Ribosome ◽

Docking Protein

Abstract The main bacterial pathway for inserting proteins into the plasma membrane relies on the signal recognition particle (SRP), composed of the Ffh protein and an associated RNA component, and the SRP-docking protein FtsY. Eukaryotes use an equivalent system of archaeal origin to deliver proteins into the endoplasmic reticulum, whereas a bacteria-derived SRP and FtsY function in the plastid. Here we report on the presence of homologs of the bacterial Ffh and FtsY proteins in various unrelated plastid-lacking unicellular eukaryotes, namely Heterolobosea, Alveida, Goniomonas, and Hemimastigophora. The monophyly of novel eukaryotic Ffh and FtsY groups, predicted mitochondrial localization experimentally confirmed for Naegleria gruberi, and a strong alphaproteobacterial affinity of the Ffh group, collectively suggest they constitute parts of an ancestral mitochondrial signal peptide-based protein-targeting system inherited from the last eukaryotic common ancestor, but lost from the majority of extant eukaryotes. The ability of putative signal peptides, predicted in a subset of mitochondrial-encoded N. gruberi proteins, to target a reporter fluorescent protein into the endoplasmic reticulum of Trypanosoma brucei, likely through their interaction with the cytosolic SRP, provided further support for this notion. We also illustrate that known mitochondrial ribosome-interacting proteins implicated in membrane protein targeting in opisthokonts (Mba1, Mdm38, and Mrx15) are broadly conserved in eukaryotes and non-redundant with the mitochondrial SRP system. Finally, we identified a novel mitochondrial protein (MAP67) present in diverse eukaryotes and related to the signal peptide-binding domain of Ffh, which may well be a hitherto unrecognized component of the mitochondrial membrane protein-targeting machinery.

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Maturation of the Na,K-ATPase in the Endoplasmic Reticulum in Health and Disease

The Journal of Membrane Biology ◽

10.1007/s00232-021-00184-z ◽

2021 ◽

Author(s):

Vitalii Kryvenko ◽

Olga Vagin ◽

Laura A. Dada ◽

Jacob I. Sznajder ◽

István Vadász

Keyword(s):

Endoplasmic Reticulum ◽

Intracellular Signaling ◽

Loss Of Function ◽

Α Subunit ◽

Rate Limiting Step ◽

Atpase Subunits ◽

A Cell ◽

Health And Disease ◽

And Function ◽

Rate Limiting

Abstract The Na,K-ATPase establishes the electrochemical gradient of cells by driving an active exchange of Na+ and K+ ions while consuming ATP. The minimal functional transporter consists of a catalytic α-subunit and a β-subunit with chaperon activity. The Na,K-ATPase also functions as a cell adhesion molecule and participates in various intracellular signaling pathways. The maturation and trafficking of the Na,K-ATPase include co- and post-translational processing of the enzyme in the endoplasmic reticulum (ER) and the Golgi apparatus and subsequent delivery to the plasma membrane (PM). The ER folding of the enzyme is considered as the rate-limiting step in the membrane delivery of the protein. It has been demonstrated that only assembled Na,K-ATPase α:β-complexes may exit the organelle, whereas unassembled, misfolded or unfolded subunits are retained in the ER and are subsequently degraded. Loss of function of the Na,K-ATPase has been associated with lung, heart, kidney and neurological disorders. Recently, it has been shown that ER dysfunction, in particular, alterations in the homeostasis of the organelle, as well as impaired ER-resident chaperone activity may impede folding of Na,K-ATPase subunits, thus decreasing the abundance and function of the enzyme at the PM. Here, we summarize our current understanding on maturation and subsequent processing of the Na,K-ATPase in the ER under physiological and pathophysiological conditions. Graphic Abstract

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Alternative redox forms of ASNA-1 separate insulin signaling from tail-anchored protein targeting and cisplatin resistance in C. elegans

Scientific Reports ◽

10.1038/s41598-021-88085-y ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Dorota Raj ◽

Ola Billing ◽

Agnieszka Podraza-Farhanieh ◽

Bashar Kraish ◽

Oskar Hemmingsson ◽

...

Keyword(s):

Endoplasmic Reticulum ◽

Insulin Secretion ◽

Protein Targeting ◽

Cisplatin Resistance ◽

Acquired Resistance ◽

Endoplasmic Reticulum Membrane ◽

C Elegans ◽

Cisplatin Sensitivity ◽

Tumor Environment ◽

The One

AbstractCisplatin is a frontline cancer therapeutic, but intrinsic or acquired resistance is common. We previously showed that cisplatin sensitivity can be achieved by inactivation of ASNA-1/TRC40 in mammalian cancer cells and in Caenorhabditis elegans. ASNA-1 has two more conserved functions: in promoting tail-anchored protein (TAP) targeting to the endoplasmic reticulum membrane and in promoting insulin secretion. However, the relation between its different functions has remained unknown. Here, we show that ASNA-1 exists in two redox states that promote TAP-targeting and insulin secretion separately. The reduced state is the one required for cisplatin resistance: an ASNA-1 point mutant, in which the protein preferentially was found in the oxidized state, was sensitive to cisplatin and defective for TAP targeting but had no insulin secretion defect. The same was true for mutants in wrb-1, which we identify as the C. elegans homolog of WRB, the ASNA1/TRC40 receptor. Finally, we uncover a previously unknown action of cisplatin induced reactive oxygen species: cisplatin induced ROS drives ASNA-1 into the oxidized form, and selectively prevents an ASNA-1-dependent TAP substrate from reaching the endoplasmic reticulum. Our work suggests that ASNA-1 acts as a redox-sensitive target for cisplatin cytotoxicity and that cisplatin resistance is likely mediated by ASNA-1-dependent TAP substrates. Treatments that promote an oxidizing tumor environment should be explored as possible means to combat cisplatin resistance.

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