Explore the vast range of titles available.

Proteins to go The type III secretion system (T3SS) is a bacterial organelle that delivers bacterial proteins into eukaryotic cells. First identified in pathogens, genome scanning has revealed these machines in many other bacteria that are symbiotic or pathogenic for animals or plants. Jorge Galán and Hans Wolf-Watz review recent work on the mechanism of T3SS action. Its presence in pathogens makes it a possible target for novel antimicrobial strategies, and these machines might also be harnessed to deliver proteins for therapeutic or vaccine purposes. Bacteria that have sustained long-standing close associations with eukaryotic hosts have evolved specific adaptations to survive and replicate in this environment. Perhaps one of the most remarkable of those adaptations is the type III secretion system (T3SS)—a bacterial organelle that has specifically evolved to deliver bacterial proteins into eukaryotic cells. Although originally identified in a handful of pathogenic bacteria, T3SSs are encoded by a large number of bacterial species that are symbiotic or pathogenic for humans, other animals including insects or nematodes, and plants. The study of these systems is leading to unique insights into not only organelle assembly and protein secretion but also mechanisms of symbiosis and pathogenesis.

Signal peptides (SPs) are short amino acid sequences in the amino terminus of many newly synthesized proteins that target proteins into, or across, membranes. Bioinformatic tools can predict SPs from amino acid sequences, but most cannot distinguish between various types of signal peptides. We present a deep neural network-based approach that improves SP prediction across all domains of life and distinguishes between three types of prokaryotic SPs. SignalP 5.0 improves proteome-wide detection of signal peptides across all organisms and can distinguish between different types of signal peptides in prokaryotes.

Key Points More than one-third of the bacterial proteome is destined for the cell envelope. The general secretory (Sec) pathway is the ubiquitous, central and essential protein export pathway into, and through, the plasma membrane. Nascent exported chains that emerge from the ribosome are scanned by chaperones and export-specific proteins that select them from cytoplasmic residents and direct them for either co-translational or post-translational export through the SecYEG channel. Kinetic competition between the scanning factors that recognize exported protein signals selects for the export route. SecA, the ATPase motor of post-translational export, 'proof-reads' its substrates, which are possibly already at the ribosome, and assembles with SecYEG into the translocase holoenzyme. SecA is allosterically activated by amino-terminal signals that are present on exported proteins and uses its highly regulated quaternary and intraprotomeric dynamics for chemo–mechanical coupling that drives protein translocation. SecYEG is regulated by its cytoplasmic partners, ribosomes and SecA, and is activated by exported proteins for either vectorial or lateral translocation. In this Review, Tsirigotaki et al . discuss recent biochemical, structural and mechanistic insights that have been gained into the consecutive steps of the general secretory (Sec) pathway. They focus on the architecture and dynamics of SecYEG and its regulation by ribosomes and SecA, and present current models of the mechanisms and energetics of the Sec-pathway-dependent secretion process in bacteria. The general secretory (Sec) pathway comprises an essential, ubiquitous and universal export machinery for most proteins that integrate into, or translocate through, the plasma membrane. Sec exportome polypeptides are synthesized as pre-proteins that have cleavable signal peptides fused to the exported mature domains. Recent advances have re-evaluated the interaction networks of pre-proteins with chaperones that are involved in pre-protein targeting from the ribosome to the SecYEG channel and have identified conformational signals as checkpoints for high-fidelity targeting and translocation. The recent structural and mechanistic insights into the channel and its ATPase motor SecA are important steps towards the elucidation of the allosteric crosstalk that mediates secretion. In this Review, we discuss recent biochemical, structural and mechanistic insights into the consecutive steps of the Sec pathway — sorting and targeting, translocation and release — in both co-translational and post-translational modes of export. The architecture and conformational dynamics of the SecYEG channel and its regulation by ribosomes, SecA and pre-proteins are highlighted. Moreover, we present conceptual models of the mechanisms and energetics of the Sec-pathway dependent secretion process in bacteria.

Components of the endoplasmic-reticulum-associated signal peptidase complex is required for infection by numerous flaviviruses, including West Nile, dengue and Zika viruses, but is not required for infection by other types of virus or for host protein synthesis. Flaviviruses infect hundreds of millions of people annually, and no antiviral therapy is available 1 , 2 . We performed a genome-wide CRISPR/Cas9-based screen to identify host genes that, when edited, resulted in reduced flavivirus infection. Here, we validated nine human genes required for flavivirus infectivity, and these were associated with endoplasmic reticulum functions including translocation, protein degradation, and N -linked glycosylation. In particular, a subset of endoplasmic reticulum-associated signal peptidase complex (SPCS) proteins was necessary for proper cleavage of the flavivirus structural proteins (prM and E) and secretion of viral particles. Loss of SPCS1 expression resulted in markedly reduced yield of all Flaviviridae family members tested (West Nile, dengue, Zika, yellow fever, Japanese encephalitis, and hepatitis C viruses), but had little impact on alphavirus, bunyavirus, or rhabdovirus infection or the surface expression or secretion of diverse host proteins. We found that SPCS1 dependence could be bypassed by replacing the native prM protein leader sequences with a class I major histocompatibility complex (MHC) antigen leader sequence. Thus, SPCS1, either directly or indirectly via its interactions with host proteins, preferentially promotes the processing of specific protein cargo, and Flaviviridae have a unique dependence on this signal peptide processing pathway. SPCS1 and other signal processing pathway members could represent pharmacological targets for inhibiting infection by the expanding number of flaviviruses of medical concern.

Post-translational modifications of proteins and the domains that recognize these modifications have central roles in creating a highly dynamic relay system that reads and responds to alterations in the cellular microenvironment. Here we review the common principles of post-translational modifications and their importance in signal integration underlying epidermal growth factor receptor signaling and endocytosis, DNA-damage responses and immunity.

The twin-arginine protein translocation (Tat) system has been characterized in bacteria, archaea and the chloroplast thylakoidal membrane. This system is distinct from other protein transport systems with respect to two key features. Firstly, it accepts cargo proteins with an N-terminal signal peptide that carries the canonical twin-arginine motif, which is essential for transport. Second, the Tat system only accepts and translocates fully folded cargo proteins across the respective membrane. Here, we review the core essential features of folded protein transport via the bacterial Tat system, using the three-component TatABC system of Escherichia coli and the two-component TatAC systems of Bacillus subtilis as the main examples. In particular, we address features of twin-arginine signal peptides, the essential Tat components and how they assemble into different complexes, mechanistic features and energetics of Tat-dependent protein translocation, cytoplasmic chaperoning of Tat cargo proteins, and the remarkable proofreading capabilities of the Tat system. In doing so, we present the current state of our understanding of Tat-dependent protein translocation across biological membranes, which may serve as a lead for future investigations.

The signal recognition particle (SRP) preferentially binds peptides destined for secretion before peptide-targeting signals are translated through recognition of elements in their mRNA, including non-coding sequences. Signal recognition particle specificity As nascent proteins are generated by translating ribosomes, they are simultaneously targeted for translocation into the endoplasmic reticulum by a protein–RNA complex known as the signal recognition particle (SRP). Judith Frydman and co-workers were intrigued by how the SRP, which is in relatively low abundance, selects its substrates among a large number of nascent peptide chains emerging from ribosomes. Studying yeast cells, they find that the SRP preferentially binds substrates destined for secretion before they are fully translated — specifically, through the non-coding elements in their mRNA and before 'targeting signals' of these substrates are translated. Elsewhere in this issue of Nature , Günter Kramer and colleagues investigate the SRP interactome in the bacterium Escherichia coli and report that SRP almost exclusively targets hydrophobic transmembrane domains of inner membrane proteins, rejecting proteins of the outer membrane and the periplasmic space. Ribosome-associated factors must properly decode the limited information available in nascent polypeptides to direct them to their correct cellular fate 1 . It is unclear how the low complexity information exposed by the nascent chain suffices for accurate recognition by the many factors competing for the limited surface near the ribosomal exit site 2 , 3 . Questions remain even for the well-studied cotranslational targeting cycle to the endoplasmic reticulum, involving recognition of linear hydrophobic signal sequences or transmembrane domains by the signal recognition particle (SRP) 4 , 5 . Notably, the SRP has low abundance relative to the large number of ribosome–nascent-chain complexes (RNCs), yet it accurately selects those destined for the endoplasmic reticulum 6 . Despite their overlapping specificities, the SRP and the cotranslationally acting Hsp70 display precise mutually exclusive selectivity in vivo for their cognate RNCs 7 , 8 . To understand cotranslational nascent chain recognition in vivo , here we investigate the cotranslational membrane-targeting cycle using ribosome profiling 9 in yeast cells coupled with biochemical fractionation of ribosome populations. We show that the SRP preferentially binds secretory RNCs before their targeting signals are translated. Non-coding mRNA elements can promote this signal-independent pre-recruitment of SRP. Our study defines the complex kinetic interaction between elongation in the cytosol and determinants in the polypeptide and mRNA that modulate SRP–substrate selection and membrane targeting.

Mitochondrial β-barrel proteins are synthesized on cytosolic ribosomes and must be specifically targeted to the organelle before their integration into the mitochondrial outer membrane. The signal that assures such precise targeting and its recognition by the organelle remained obscure. In the present study we show that a specialized β-hairpin motif is this long searched for signal. We demonstrate that a synthetic β-hairpin peptide competes with the import of mitochondrial β-barrel proteins and that proteins harbouring a β-hairpin peptide fused to passenger domains are targeted to mitochondria. Furthermore, a β-hairpin motif from mitochondrial proteins targets chloroplast β-barrel proteins to mitochondria. The mitochondrial targeting depends on the hydrophobicity of the β-hairpin motif. Finally, this motif interacts with the mitochondrial import receptor Tom20. Collectively, we reveal that β-barrel proteins are targeted to mitochondria by a dedicated β-hairpin element, and this motif is recognized at the organelle surface by the outer membrane translocase. Mitochondrial β-barrel proteins are synthesized in the cytosol before being targeted to the organelle. Here, Jores et al. show that a specialized hydrophobic β-hairpin motif is the previously undefined targeting sequence and is recognized by the mitochondrial outer membrane translocase.

Malaria blood stage parasites export a large number of proteins into their host erythrocyte to change it from a container of predominantly hemoglobin optimized for the transport of oxygen into a niche for parasite propagation. To understand this process, it is crucial to know which parasite proteins are exported into the host cell. This has been aided by the PEXEL/HT sequence, a five-residue motif found in many exported proteins, leading to the prediction of the exportome. However, several PEXEL/HT negative exported proteins (PNEPs) indicate that this exportome is incomplete and it remains unknown if and how many further PNEPs exist. Here we report the identification of new PNEPs in the most virulent malaria parasite Plasmodium falciparum. This includes proteins with a domain structure deviating from previously known PNEPs and indicates that PNEPs are not a rare exception. Unexpectedly, this included members of the MSP-7 related protein (MSRP) family, suggesting unanticipated functions of MSRPs. Analyzing regions mediating export of selected new PNEPs, we show that the first 20 amino acids of PNEPs without a classical N-terminal signal peptide are sufficient to promote export of a reporter, confirming the concept that this is a shared property of all PNEPs of this type. Moreover, we took advantage of newly found soluble PNEPs to show that this type of exported protein requires unfolding to move from the parasitophorous vacuole (PV) into the host cell. This indicates that soluble PNEPs, like PEXEL/HT proteins, are exported by translocation across the PV membrane (PVM), highlighting protein translocation in the parasite periphery as a general means in protein export of malaria parasites.

Bacteria control gene expression in concert with their population density by a process called quorum sensing, which is modulated by bacterial chemical signals and environmental factors. In the human pathogen Streptococcus pyogenes , production of secreted virulence factor SpeB is controlled by a quorum-sensing pathway and environmental pH. The quorum-sensing pathway consists of a secreted leaderless peptide signal (SIP), and its cognate receptor RopB. Here, we report that the SIP quorum-sensing pathway has a pH-sensing mechanism operative through a pH-sensitive histidine switch located at the base of the SIP-binding pocket of RopB. Environmental acidification induces protonation of His144 and reorganization of hydrogen bonding networks in RopB, which facilitates SIP recognition. The convergence of two disparate signals in the SIP signaling pathway results in induction of SpeB production and increased bacterial virulence. Our findings provide a model for investigating analogous crosstalk in other microorganisms. The mechanism by which environmental pH controls the virulence of the pathogen Streptococcus pyogenes is unclear. Here, Do et al. show that changes in pH affect the activity of the virulence regulator RopB via its interaction with a quorum-sensing peptide signal.