The role of bryozoans in fossil reefs—an example from the Middle Devonian of the Western Sahara

Stenolaemate bryozoans with their stable calcitic skeletons play a significant role in reef building. In the Middle Devonian Sabkhat Lafayrina reef complex (Western Sahara), bryozoans are abundant and diverse. Although they do not form part of the principal framework of reefs, bryozoans are involved significantly in reef growth, especially in the initial stage. In this way, bryozoans are important with respect to initiating reef growth. They contribute greatly to sediment stabilization, making it possible for principal reef builders to grow on hardened and stabilized substrates, and also play sediment-baffling and sediment-filling roles. The aim of this study is to document the diversity of bryozoans in a Middle Devonian reef complex and to estimate their potential for initiation and contribution to reef structures.     

Electron microscopy holdings of the Protein Data Bank: the impact of the resolution revolution,new validation tools,and implications for the future

As a discipline, structural biology has been transformed by the three-dimensional electron microscopy (3DEM) “Resolution Revolution” made possible by convergence of robust cryo-preservation of vitrified biological materials, sample handling systems, and measurement stages operating a liquid nitrogen temperature, improvements in electron optics that preserve phase information at the atomic level, direct electron detectors (DEDs), high-speed computing with graphics processing units, and rapid advances in data acquisition and processing software. 3DEM structure information (atomic coordinates and related metadata) are archived in the open-access Protein Data Bank (PDB), which currently holds more than 11,000 3DEM structures of proteins and nucleic acids, and their complexes with one another and small-molecule ligands (~ 6% of the archive). Underlying experimental data (3DEM density maps and related metadata) are stored in the Electron Microscopy Data Bank (EMDB), which currently holds more than 21,000 3DEM density maps. After describing the history of the PDB and the Worldwide Protein Data Bank (wwPDB) partnership, which jointly manages both the PDB and EMDB archives, this review examines the origins of the resolution revolution and analyzes its impact on structural biology viewed through the lens of PDB holdings. Six areas of focus exemplifying the impact of 3DEM across the biosciences are discussed in detail (icosahedral viruses, ribosomes, integral membrane proteins, SARS-CoV-2 spike proteins, cryogenic electron tomography, and integrative structure determination combining 3DEM with complementary biophysical measurement techniques), followed by a review of 3DEM structure validation by the wwPDB that underscores the importance of community engagement.     

Two msbB genes encoding maximal acylation of lipid A are required for invasive Shigella flexneri to mediate inflammatory rupture and destruction of the intestinal epithelium

Shigella flexneri is a Gram-negative pathogen that invades and causes inflammatory destruction of the human colonic epithelium, thus leading to bloody diarrhea and dysentery. A type III secretion system that delivers effector proteins into target eukaryotic cells is largely responsible for cell and tissue invasion. However, the respective role of this invasive phenotype and of lipid A, the endotoxin of the Shigella LPS, in eliciting the inflammatory cascade that leads to rupture and destruction of the epithelial barrier, was unknown. We investigated whether genetic detoxification of lipid A would cause significant alteration in pathogenicity. We showed that S. flexneri has two functional msbB genes, one carried by the chromosome (msbB1) and the other by the virulence plasmid (msbB2), the products of which act in complement to produce full acyl-oxy-acylation of the myristate at the 3' position of the lipid A glucosamine disaccharide. A mutant in which both the msbB1 and msbB2 genes have been inactivated was impaired in its capacity to cause TNF-alpha production by human monocytes and to cause rupture and inflammatory destruction of the epithelial barrier in the rabbit ligated intestinal loop model of shigellosis, indicating that lipid A plays a significant role in aggravating inflammation that eventually destroys the intestinal barrier. In addition, neutralization of TNF-alpha during invasion by the wild-type strain strongly impaired its ability to cause rupture and inflammatory destruction of the epithelial lining, thus indicating that TNF-alpha is a major effector of epithelial destruction by Shigella.     

Salt-induced changes in the plasma membrane proteome of the halotolerant alga Dunaliella salina as revealed by blue native gel electrophoresis and nano-LC-MS/MS analysis

The halotolerant alga Dunaliella salina is a recognized model photosynthetic organism for studying plant adaptation to high salinity. The adaptation mechanisms involve major changes in the proteome composition associated with energy metabolism and carbon and iron acquisition. To clarify the molecular basis for the remarkable resistance to high salt, we performed a comprehensive proteomics analysis of the plasma membrane. Plasma membrane proteins were recognized by tagging intact cells with a membrane-impermeable biotin derivative. Proteins were resolved by two-dimensional blue native/SDS-PAGE and identified by nano-LC-MS/MS. Of 55 identified proteins, about 60% were integral membrane or membrane-associated proteins. We identified novel surface coat proteins, lipid-metabolizing enzymes, a new family of membrane proteins of unknown function, ion transporters, small GTP-binding proteins, and heat shock proteins. The abundance of 20 protein spots increased and that of two protein spots decreased under high salt. The major salt-regulated proteins were implicated in protein and membrane structure stabilization and within signal transduction pathways. The migration profiles of native protein complexes on blue native gels revealed oligomerization or co-migration of major surface-exposed proteins, which may indicate mechanisms of stabilization at high salinity.     

A bryozoan fauna from the Santa Lucia Formation (Lower–Middle Devonian) of Cantabrian Mountains,NW Spain

The Lower to Middle Devonian Santa Lucia Formation of NW Spain contains a rich and well-preserved bryozoan fauna. An assemblage containing 14 species is described from two localities in Cantabrian Mountains, Abelgas, and Paradilla. One new genus with one new species is described: Isostylus abelgasensis n. sp. n. gen. Another two new species have been found: Microcampylus minor n. sp. and Acanthoclema parvula n. sp. Additionally, the following species were identified: Cyclotrypa communis (Ulrich 1890), Fistuliphragma gracilis Ernst 2008a, Leioclema attenuatum Duncan 1939, Hemitrypa cf. tenella Barrande in Počta 1894, Fenestella aff. parallela Hall 1881, Anastomopora adnata (Hall 1883), Semicoscinium rhombicum Ulrich 1890, Quadrisemicoscinium discretum (Prantl 1932). Three species are described in open nomenclature: Trepostomata sp. indet.1 and 2, and Semicoscinium sp. The described bryozoan fauna shows connections to the Lower Devonian of Bohemia, and to the Middle Devonian of Rhenish Massif and North America.     

Increase of histidine content in Brassica rapa subsp. oleifera by over-expression of histidine-rich fusion proteins

An approach that enables the increase of the quantity of a specific amino acid in crop plants is reported. Oleosin gene from Arabidopsis thaliana or 30K movement protein gene of Tobacco mosaic virus (TMV; genus Tobamovirus) were cloned under the control of napin or hybrid promoters, and in fusion to synthetic poly-histidine (poly-His) sequences for transformation into spring turnip rape (Brassica rapa subsp. oleifera; synonym to B. campestris). The most stable expression cassettes for the poly-His production prior to the plant transformation were selected by analyzing the protein expression in in vitro translation and in transient plant expression systems using GFP as marker. Expression of the poly-His-constructs in transgenic Brassica rapa plants was analyzed using dot and western blotting and PCR. The constructs were stably expressed in the third generation of the transgenic plant lines. Histidine content was measured from the seeds of the transgenic plants, and some plant lines had more than 20% increase in histidine content compared to wild type. The methodology may be widely applicable to increase the content of any amino acid in crop plants including those encoded by rare codons.     

The power and the limitations of cross-species protein identification by mass spectrometry-driven sequence similarity searches

Mass spectrometry-driven BLAST (MS BLAST) is a database search protocol for identifying unknown proteins by sequence similarity to homologous proteins available in a database. MS BLAST utilizes redundant, degenerate, and partially inaccurate peptide sequence data obtained by de novo interpretation of tandem mass spectra and has become a powerful tool in functional proteomic research. Using computational modeling, we evaluated the potential of MS BLAST for proteome-wide identification of unknown proteins. We determined how the success rate of protein identification depends on the full-length sequence identity between the queried protein and its closest homologue in a database. We also estimated phylogenetic distances between organisms under study and related reference organisms with completely sequenced genomes that allow substantial coverage of unknown proteomes.     

Using Integrative Modeling Platform to compute,validate, and archive a model of a protein complex structure

Biology is advanced by producing structural models of biological systems, such as protein complexes. Some systems are recalcitrant to traditional structure determination methods. In such cases, it may still be possible to produce useful models by integrative structure determination that depends on simultaneous use of multiple types of data. An ensemble of models that are sufficiently consistent with the data is produced by a structural sampling method guided by a data‐dependent scoring function. The variation in the ensemble of models quantified the uncertainty of the structure, generally resulting from the uncertainty in the input information and actual structural heterogeneity in the samples used to produce the data. Here, we describe how to generate, assess, and interpret ensembles of integrative structural models using our open source Integrative Modeling Platform program ( https://integrativemodeling.org ).