Prokaryotic membrane coat - like proteins: An update.

Membrane coat proteins are essential players in the eukaryotic endomembrane traffic system. Previous work identified proteins with the membrane-coat architecture in prokaryotes, specifically in the Planctomycetes, Verrucomicrobia and Chlamydiae (PVC) superphylum, bacteria that display the most developed prokaryotic endomembrane system. Hence, the membrane coat-like (MCL) proteins are predicted to play a central role in this system but their actual function is still unknown. In this work we strengthened previous structure predictions for these prokaryotic MCL proteins. We also detected new putative MCL proteins in the Planctomycete Gemmata obscuriglobus. Structural analysis of these revealed the presence of additional domains apart from the ß-propeller and α-solenoid combination, characteristic of the membrane-coat architecture. Functions associated with these domains include some related to carbohydrate or membrane/lipid binding. Using homology-based methods, we found MCL proteins in other bacterial phyla, but the most abundant hits are still restricted to Planctomycetes and Verrucomicrobia. Detailed inspection of neighbouring genes of MCL in G. obscuriglobus supports the idea that the function of these proteins is related to membrane manipulation. No significant hits were found in Archaea, including Asgard archaea. More than 10 years after their original detection, PVC bacteria are still uniquely linked to eukaryotes through the structure of the MCL proteins sustaining their endomembrane system.

The squalene route to C30 carotenoid biosynthesis and the origins of carotenoid biosynthetic pathways.

Carotenoids are isoprenoid lipids found across the tree of life with important implications in oxidative stress adaptations, photosynthetic metabolisms, as well as in membrane dynamics. The canonical view is that C40 carotenoids are synthesized from phytoene and C30 carotenoids from diapophytoene. Squalene is mostly associated with the biosynthesis of polycyclic triterpenes, although there have been suggestions that it could also be involved in the biosynthesis of C30 carotenoids. However, demonstration of the existence of this pathway in nature is lacking. Here, we demonstrate that C30 carotenoids are synthesized from squalene in the Planctomycetes bacteria and that this squalene route to C30 carotenoids is the most widespread in prokaryotes. Using the evolutionary history of carotenoid and squalene amino oxidases, we propose an evolutionary scenario to explain the origin and diversification of the different carotenoid and squalene-related pathways. We show that carotenoid biosynthetic pathways have been constantly transferred and neofunctionalized during prokaryotic evolution. One possible origin of the squalene pathway connects it with the one of C40 carotenoid synthesis of Cyanobacteria. The widespread occurrence of the squalene route to C30 carotenoids in Bacteria increases the functional repertoire of squalene, establishing it as a general hub of carotenoids and polycyclic triterpenes synthesis.

Early origin and evolution of the FtsZ/tubulin protein family.

The origin of the FtsZ/tubulin protein family was extremely relevant for life since these proteins are present in nearly all organisms, carrying out essential functions such as cell division or forming a major part of the cytoskeleton in eukaryotes. Therefore, investigating the early evolution of the FtsZ/tubulin protein family could reveal crucial aspects of the diversification of the three domains of life. In this study, we revisited the phylogenies of the FtsZ/tubulin protein family in an extensive prokaryotic diversity, focusing on the main evolutionary events that occurred during its evolution. We found evidence of its early origin in the last universal common ancestor since FtsZ was present in the last common ancestor of Bacteria and Archaea. In bacteria, ftsZ genes are genomically associated with the bacterial division gene cluster, while in archaea, ftsZ duplicated prior to archaeal diversification, and one of the copies is associated with protein biosynthesis genes. Archaea have expanded the FtsZ/tubulin protein family with sequences closely related to eukaryotic tubulins. In addition, we report novel CetZ-like groups in Halobacterota and Asgardarchaeota. Investigating the C-termini of prokaryotic paralogs basal to eukaryotic tubulins, we show that archaeal CetZ, as well as the plasmidic TubZ from Firmicutes, most likely originated from archaeal FtsZ. Finally, prokaryotic tubulins are restricted to Odinarchaeaota and Prosthecobacter species, and they seem to belong to different molecular systems. However, their phylogenies suggest that they are closely related to α/ß-tubulins pointing to a potential ancestrality of these eukaryotic paralogs of tubulins.

Functional diversity of isoprenoid lipids in Methylobacterium extorquens PA1.

Hopanoids and carotenoids are two of the major isoprenoid-derived lipid classes in prokaryotes that have been proposed to have similar membrane ordering properties as sterols. Methylobacterium extorquens contains hopanoids and carotenoids in their outer membrane, making them an ideal system to investigate the role of isoprenoid lipids in surface membrane function and cellular fitness. By genetically knocking out hpnE and crtB we disrupted the production of squalene and phytoene in M. extorquens PA1, which are the presumed precursors for hopanoids and carotenoids respectively. Deletion of hpnE revealed that carotenoid biosynthesis utilizes squalene as a precursor resulting in pigmentation with a C30 backbone, rather than the previously predicted canonical C40 phytoene-derived pathway. Phylogenetic analysis suggested that M. extorquens may have acquired the C30 pathway through lateral gene transfer from Planctomycetes. Surprisingly, disruption of carotenoid synthesis did not generate any major growth or membrane biophysical phenotypes, but slightly increased sensitivity to oxidative stress. We further demonstrated that hopanoids but not carotenoids are essential for growth at higher temperatures, membrane permeability and tolerance of low divalent cation concentrations. These observations show that hopanoids and carotenoids serve diverse roles in the outer membrane of M. extorquens PA1.

Homology and Modular Evolution of CATCHR at the Origin of the Eukaryotic Endomembrane System.

The membrane trafficking is an essential process of eukaryotic cells, as it manages vesicular trafficking toward different parts of the cell. In this process, membrane fusions between vesicles and target membranes are mediated by several factors, including the multisubunit tethering complexes. One type of multisubunit tethering complex, the complexes associated with tethering containing helical rods (CATCHR), encompasses the exocyst, COG, GARP, and DSL1 complexes. The CATCHR share similarities at sequence, structural, and protein-complex organization level although their actual relationship is still poorly understood. In this study, we have re-evaluated CATCHR at different levels, demonstrating that gene duplications followed by neofunctionalization, were key for their origin. Our results, reveals that there are specific homology relationships and parallelism within and between the CATCHR suggesting that most of these complexes are composed by modular tetramers of four different kinds of proteins, three of them having a clear common origin. The extension of CATCHR family occurred concomitantly with the protein family expansions of their molecular partners, such as small GTPases and SNAREs, among others, and likely providing functional specificity. Our results provide novel insights into the structural organization and mechanism of action of CATCHR, with implications for the evolution of the endomembrane system of eukaryotes and promoting CATCHR as ideal candidates to study the evolution of multiprotein complexes.

Comparative Genomics of Peroxisome Biogenesis Proteins: Making Sense of the PEX Proteins.

PEX genes encode proteins involved in peroxisome biogenesis and proliferation. Using a comparative genomics approach, we clarify the evolutionary relationships between the 37 known PEX proteins in a representative set of eukaryotes, including all common model organisms, pathogenic unicellular eukaryotes and human. A large number of previously unknown PEX orthologs were identified. We analyzed all PEX proteins, their conservation and domain architecture and defined the core set of PEX proteins that is required to make a peroxisome. The molecular processes in peroxisome biogenesis in different organisms were put into context, showing that peroxisomes are not static organelles in eukaryotic evolution. Organisms that lack peroxisomes still contain a few PEX proteins, which probably play a role in alternative processes. Finally, the relationships between PEX proteins of two large families, the Pex11 and Pex23 families, were analyzed, thereby contributing to the understanding of their complicated and sometimes incorrect nomenclature. We provide an exhaustive overview of this important eukaryotic organelle.

Structural, Evolutionary, and Functional Analysis of the Protein O-Mannosyltransferase Family in Pathogenic Fungi.

Protein O-mannosyltransferases (Pmts) comprise a group of proteins that add mannoses to substrate proteins at the endoplasmic reticulum. This post-translational modification is important for the faithful transfer of nascent glycoproteins throughout the secretory pathway. Most fungi genomes encode three O-mannosyltransferases, usually named Pmt1, Pmt2, and Pmt4. In pathogenic fungi, Pmts, especially Pmt4, are key factors for virulence. Although the importance of Pmts for fungal pathogenesis is well established in a wide range of pathogens, questions remain regarding certain features of Pmts. For example, why does the single deletion of each pmt gene have an asymmetrical impact on host colonization? Here, we analyse the origin of Pmts in fungi and review the most important phenotypes associated with Pmt mutants in pathogenic fungi. Hence, we highlight the enormous relevance of these glycotransferases for fungal pathogenic development.

Origin and Evolution of Polycyclic Triterpene Synthesis.

Polycyclic triterpenes are members of the terpene family produced by the cyclization of squalene. The most representative polycyclic triterpenes are hopanoids and sterols, the former are mostly found in bacteria, whereas the latter are largely limited to eukaryotes, albeit with a growing number of bacterial exceptions. Given their important role and omnipresence in most eukaryotes, contrasting with their scant representation in bacteria, sterol biosynthesis was long thought to be a eukaryotic innovation. Thus, their presence in some bacteria was deemed to be the result of lateral gene transfer from eukaryotes. Elucidating the origin and evolution of the polycyclic triterpene synthetic pathways is important to understand the role of these compounds in eukaryogenesis and their geobiological value as biomarkers in fossil records. Here, we have revisited the phylogenies of the main enzymes involved in triterpene synthesis, performing gene neighborhood analysis and phylogenetic profiling. Squalene can be biosynthesized by two different pathways containing the HpnCDE or Sqs proteins. Our results suggest that the HpnCDE enzymes are derived from carotenoid biosynthesis ones and that they assembled in an ancestral squalene pathway in bacteria, while remaining metabolically versatile. Conversely, the Sqs enzyme is prone to be involved in lateral gene transfer, and its emergence is possibly related to the specialization of squalene biosynthesis. The biosynthesis of hopanoids seems to be ancestral in the Bacteria domain. Moreover, no triterpene cyclases are found in Archaea, invoking a potential scenario in which eukaryotic genes for sterol biosynthesis assembled from ancestral bacterial contributions in early eukaryotic lineages.

Essentiality of sterol synthesis genes in the planctomycete bacterium Gemmata obscuriglobus.

Sterols and hopanoids are chemically and structurally related lipids mostly found in eukaryotic and bacterial cell membranes. Few bacterial species have been reported to produce sterols and this anomaly had originally been ascribed to lateral gene transfer (LGT) from eukaryotes. In addition, the functions of sterols in these bacteria are unknown and the functional overlap between sterols and hopanoids is still unclear. Gemmata obscuriglobus is a bacterium from the Planctomycetes phylum that synthesizes sterols, in contrast to its hopanoid-producing relatives. Here we show that sterols are essential for growth of G. obscuriglobus, and that sterol depletion leads to aberrant membrane structures and defects in budding cell division. This report of sterol essentiality in a prokaryotic species advances our understanding of sterol distribution and function, and provides a foundation to pursue fundamental questions in evolutionary cell biology.

Transcriptome, proteome and draft genome of Euglena gracilis.

BACKGROUND: Photosynthetic euglenids are major contributors to fresh water ecosystems. Euglena gracilis in particular has noted metabolic flexibility, reflected by an ability to thrive in a range of harsh environments. E. gracilis has been a popular model organism and of considerable biotechnological interest, but the absence of a gene catalogue has hampered both basic research and translational efforts. RESULTS: We report a detailed transcriptome and partial genome for E. gracilis Z1. The nuclear genome is estimated to be around 500 Mb in size, and the transcriptome encodes over 36,000 proteins and the genome possesses less than 1% coding sequence. Annotation of coding sequences indicates a highly sophisticated endomembrane system, RNA processing mechanisms and nuclear genome contributions from several photosynthetic lineages. Multiple gene families, including likely signal transduction components, have been massively expanded. Alterations in protein abundance are controlled post-transcriptionally between light and dark conditions, surprisingly similar to trypanosomatids. CONCLUSIONS: Our data provide evidence that a range of photosynthetic eukaryotes contributed to the Euglena nuclear genome, evidence in support of the 'shopping bag' hypothesis for plastid acquisition. We also suggest that euglenids possess unique regulatory mechanisms for achieving extreme adaptability, through mechanisms of paralog expansion and gene acquisition.