34,553 research outputs found

    SChloro: directing Viridiplantae proteins to six chloroplastic sub-compartments

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    Motivation: Chloroplasts are organelles found in plants and involved in several important cell processes. Similarly to other compartments in the cell, chloroplasts have an internal structure comprising several sub-compartments, where different proteins are targeted to perform their functions. Given the relation between protein function and localization, the availability of effective computational tools to predict protein sub-organelle localizations is crucial for large-scale functional studies. Results: In this paper we present SChloro, a novel machine-learning approach to predict protein sub-chloroplastic localization, based on targeting signal detection and membrane protein information. The proposed approach performs multi-label predictions discriminating six chloroplastic sub-compartments that include inner membrane, outer membrane, stroma, thylakoid lumen, plastoglobule and thylakoid membrane. In comparative benchmarks, the proposed method outperforms current state-of-the-art methods in both single-and multi-compartment predictions, with an overall multi-label accuracy of 74%. The results demonstrate the relevance of the approach that is eligible as a good candidate for integration into more general large-scale annotation pipelines of protein subcellular localization

    Plastid proteome prediction for diatoms and other algae with secondary plastids of the red lineage

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    The plastids of ecologically and economically important algae from phyla such as stramenopiles, dinoflagellates and cryptophytes were acquired via a secondary endosymbiosis and are surrounded by three or four membranes. Nuclear-encoded plastid-localized proteins contain N-terminal bipartite targeting peptides with the conserved amino acid sequence motif ‘ASAFAP’. Here we identify the plastid proteomes of two diatoms, Thalassiosira pseudonana and Phaeodactylum tricornutum, using a customized prediction tool (ASAFind) that identifies nuclear-encoded plastid proteins in algae with secondary plastids of the red lineage based on the output of SignalP and the identification of conserved ‘ASAFAP’ motifs and transit peptides. We tested ASAFind against a large reference dataset of diatom proteins with experimentally confirmed subcellular localization and found that the tool accurately identified plastid-localized proteins with both high sensitivity and high specificity. To identify nucleus-encoded plastid proteins of T. pseudonana and P. tricornutum we generated optimized sets of gene models for both whole genomes, to increase the percentage of full-length proteins compared with previous assembly model sets. ASAFind applied to these optimized sets revealed that about 8% of the proteins encoded in their nuclear genomes were predicted to be plastid localized and therefore represent the putative plastid proteomes of these algae

    Annotating Protein Functional Residues by Coupling High-Throughput Fitness Profile and Homologous-Structure Analysis.

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    Identification and annotation of functional residues are fundamental questions in protein sequence analysis. Sequence and structure conservation provides valuable information to tackle these questions. It is, however, limited by the incomplete sampling of sequence space in natural evolution. Moreover, proteins often have multiple functions, with overlapping sequences that present challenges to accurate annotation of the exact functions of individual residues by conservation-based methods. Using the influenza A virus PB1 protein as an example, we developed a method to systematically identify and annotate functional residues. We used saturation mutagenesis and high-throughput sequencing to measure the replication capacity of single nucleotide mutations across the entire PB1 protein. After predicting protein stability upon mutations, we identified functional PB1 residues that are essential for viral replication. To further annotate the functional residues important to the canonical or noncanonical functions of viral RNA-dependent RNA polymerase (vRdRp), we performed a homologous-structure analysis with 16 different vRdRp structures. We achieved high sensitivity in annotating the known canonical polymerase functional residues. Moreover, we identified a cluster of noncanonical functional residues located in the loop region of the PB1 β-ribbon. We further demonstrated that these residues were important for PB1 protein nuclear import through the interaction with Ran-binding protein 5. In summary, we developed a systematic and sensitive method to identify and annotate functional residues that are not restrained by sequence conservation. Importantly, this method is generally applicable to other proteins about which homologous-structure information is available.ImportanceTo fully comprehend the diverse functions of a protein, it is essential to understand the functionality of individual residues. Current methods are highly dependent on evolutionary sequence conservation, which is usually limited by sampling size. Sequence conservation-based methods are further confounded by structural constraints and multifunctionality of proteins. Here we present a method that can systematically identify and annotate functional residues of a given protein. We used a high-throughput functional profiling platform to identify essential residues. Coupling it with homologous-structure comparison, we were able to annotate multiple functions of proteins. We demonstrated the method with the PB1 protein of influenza A virus and identified novel functional residues in addition to its canonical function as an RNA-dependent RNA polymerase. Not limited to virology, this method is generally applicable to other proteins that can be functionally selected and about which homologous-structure information is available

    An Outlook on the Localisation and Structure-Function Relationships of R Proteins in Solanum

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    The co-evolution of plants and plant-pathogens shaped a multi-layered defence system in plants, in which Resistance proteins (R proteins) play a significant role. A fundamental understanding of the functioning of these R proteins and their position in the broader defence system of the plant is essential. Sub-project 3 of the BIOEXPLOIT programme studies how R proteins are activated upon effector recognition and how recognition is conveyed in resistance signalling pathways, using the solanaceous R proteins Rx1 (from S. tuberosum spp. andigena; conferring extreme resistance against Potato Virus X), I-2 (from S. lycopersicon; mediating resistance to Fusarium oxysporum) and Mi-1.2 (from S. lycopersicon; conferring resistance to Meloidogyne incognita) as model systems. The results obtained in this project will serve as a model for other R proteins and will be translated to potential applications or alternative strategies for disease resistance. These include the modification of the recognition specificity of R proteins with the aim to obtain broad spectrum resistance to major pathogens in potato

    Regulation of alphaherpesvirus infections by the ICP0 family of proteins

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    Immediate-early protein ICP0 of herpes simplex virus type 1 (HSV-1) is important for the regulation of lytic and latent viral infection. Like the related proteins expressed by other alphaherpesviruses, ICP0 has a zinc-stabilized RING finger domain that confers E3 ubiquitin ligase activity. This domain is essential for the core functions of ICP0 and its activity leads to the degradation of a number of cellular proteins, some of which are involved in cellular defences that restrict viral infection. The article reviews recent advances in ICP0-related research, with an emphasis on the mechanisms by which ICP0 and related proteins counteract antiviral restriction and the roles in this process of cellular nuclear substructures known as ND10 or PML nuclear bodies. We also summarize recent advances in the understanding of the biochemical aspects of ICP0 activity. These studies highlight the importance of the SUMO conjugation pathway in both intrinsic resistance to HSV-1 infection and in substrate targeting by ICP0. The topics discussed in this review are relevant not only to HSV-1 infection, but also to cellular intrinsic resistance against herpesviruses more generally and the mechanisms by which viruses can evade this restriction

    Structure-function mapping of a heptameric module in the nuclear pore complex.

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    The nuclear pore complex (NPC) is a multiprotein assembly that serves as the sole mediator of nucleocytoplasmic exchange in eukaryotic cells. In this paper, we use an integrative approach to determine the structure of an essential component of the yeast NPC, the ~600-kD heptameric Nup84 complex, to a precision of ~1.5 nm. The configuration of the subunit structures was determined by satisfaction of spatial restraints derived from a diverse set of negative-stain electron microscopy and protein domain-mapping data. Phenotypic data were mapped onto the complex, allowing us to identify regions that stabilize the NPC's interaction with the nuclear envelope membrane and connect the complex to the rest of the NPC. Our data allow us to suggest how the Nup84 complex is assembled into the NPC and propose a scenario for the evolution of the Nup84 complex through a series of gene duplication and loss events. This work demonstrates that integrative approaches based on low-resolution data of sufficient quality can generate functionally informative structures at intermediate resolution
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