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Date palm (Phoenix dactylifera L.) is an important crop in arid regions that is well-adapted to desert ecosystems. To understand the remarkable ability to grow and yield in water-limited environments, experiments with water-withholding for up to four weeks were conducted. In response to drought, root, rather than leaf, osmotic strength increased, with organic solutes such as sugars and amino acids contributing more to the osmolyte increase than minerals. Consistently, carbon and amino acid metabolism was acclimated toward biosynthesis at both the transcriptional and translational levels. In leaves, a remodeling of membrane systems was observed, suggesting changes in thylakoid lipid composition, which together with the restructuring of the photosynthetic apparatus, indicated an acclimation preventing oxidative damage. Thus, xerophilic date palm avoids oxidative damage under drought by combined prevention and rapid detoxification of oxygen radicals. Although minerals were expected to serve as cheap key osmotics, date palm also relies on organic osmolytes for osmotic adjustment of the roots during early drought acclimation. The diversion of these resources away from growth is consistent with date palm's strategy of generally slow growth in harsh environments and clearly indicates a trade-off between growth and stress-related physiological responses., (© The Author(s) 2024. Published by Oxford University Press on behalf of the Society for Experimental Biology.)

Chromosome segregation is crucial for the faithful inheritance of DNA to the daughter cells after DNA replication. For this, the kinetochore, a megadalton protein complex, assembles on centromeric chromatin containing the histone H3 variant CENP-A, and provides a physical connection to the microtubules. Here, we report an unanticipated role for enzymes required for β-1,6- and β-1,3-glucan biosynthesis in regulating kinetochore function in Saccharomyces cerevisiae. These carbohydrates are the major constituents of the yeast cell wall. We found that the deletion of KRE6, which encodes a glycosylhydrolase/ transglycosidase required for β-1,6-glucan synthesis, suppressed the centromeric defect of mutations in components of the kinetochore, foremost the NDC80 components Spc24, Spc25, the MIND component Nsl1, and Okp1, a constitutive centromere-associated network protein. Similarly, the absence of Fks1, a β-1,3-glucan synthase, and Kre11/Trs65, a TRAPPII component, suppressed a mutation in SPC25. Genetic analysis indicates that the reduction of intracellular β-1,6- and β-1,3-glucans, rather than the cell wall glucan content, regulates kinetochore function. Furthermore, we found a physical interaction between Kre6 and CENP-A/Cse4 in yeast, suggesting a potential function for Kre6 in glycosylating CENP-A/Cse4 or another kinetochore protein. This work shows a moonlighting function for selected cell wall synthesis proteins in regulating kinetochore assembly, which may provide a mechanism to connect the nutritional status of the cell to cell-cycle progression and chromosome segregation., Competing Interests: Conflicts of interest The author(s) declare no conflict of interest., (© The Author(s) 2023. Published by Oxford University Press on behalf of The Genetics Society of America. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Clubroot disease caused by Plasmodiophora brassicae ( P. brassicae ) severely threatens the cultivation of Cruciferous plants, especially Chinese cabbage. Recently, resistance genes in plants have been reported to encode for a Ca 2+ -permeable channel in the plasma membrane, which can mediate the cytosolic Ca 2+ increase in plant cells upon pathogen attack. However, the downstream Ca 2+ sensor and decoder are still unknown. In this study, we identified the virulent and avirulent P. brassicae isolates (Pbs) of two near isogenic lines, CR 3-2 and CS 3-2, with CR 3-2 harboring clubroot resistant gene BraCRa . The transcriptomic analysis was then conducted with CR 3-2 after inoculating with virulent isolate PbE and avirulent isolate Pb4. From the differentially expressed genes of transcriptomic data, we identified a Ca 2+ -sensor encoding gene, BraCBL1 . 2 , that was highly induced in CR 3-2 during infection by Pb4 but not by PbE. Moreover, GUS histochemical staining and subcellular localization analysis revealed that BraCBL1 . 2 was specifically expressed in the root hair cells of Arabidopsis and encoded a putative Ca 2+ sensor localized in the plasma membrane. We also developed an assay to investigate the BraCRa-mediated hypersensitive response (HR) in tobacco leaves. The results suggest that BraCBL1.2 is involved in the BraCRa-mediated plant ETI immune response against P. brassicae . In addition, we verified that overexpression of BraCBL1.2 enhanced clubroot resistance in Arabidopsis . Collectively, our data identified the involvement of a Ca 2+ sensor in BraCRa-mediated clubroot resistance in Chinese cabbage, providing a theoretical basis for further research on the resistance of Chinese cabbage to P. brassicae ., Competing Interests: The authors declare that they have no conflicts of interest., (© The Author(s) 2024. Published by Oxford University Press on behalf of Nanjing Agricultural University.)

Queuosine (Q) is a complex tRNA modification found in bacteria and eukaryotes at position 34 of four tRNAs with a GUN anticodon, and it regulates the translational efficiency and fidelity of the respective codons that differ at the Wobble position. In bacteria, the biosynthesis of Q involves two precursors, preQ0 and preQ1, whereas eukaryotes directly obtain Q from bacterial sources. The study of queuosine has been challenging due to the limited availability of high-throughput methods for its detection and analysis. Here, we have employed direct RNA sequencing using nanopore technology to detect the modification of tRNAs with Q and Q precursors. These modifications were detected with high accuracy on synthetic tRNAs as well as on tRNAs extracted from Schizosaccharomyces pombe and Escherichia coli by comparing unmodified to modified tRNAs using the tool JACUSA2. Furthermore, we present an improved protocol for the alignment of raw sequence reads that gives high specificity and recall for tRNAs ex cellulo that, by nature, carry multiple modifications. Altogether, our results show that 7-deazaguanine-derivatives such as queuosine are readily detectable using direct RNA sequencing. This advancement opens up new possibilities for investigating these modifications in native tRNAs, furthering our understanding of their biological function., (© The Author(s) 2023. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Background and Aims: The genus Buxus has high levels of endemism in the Caribbean flora, with ~50 taxa. In Cuba, 82 % grow on ultramafic substrates and 59 % are nickel (Ni) accumulators or Ni hyperaccumulators. Hence it is an ideal model group to study if this diversification could be related to adaptation to ultramafic substrates and to Ni hyperaccumulation., Methods: We generated a well-resolved molecular phylogeny, including nearly all of the Neotropical and Caribbean Buxus taxa. To obtain robust divergence times we tested for the effects of different calibration scenarios, and we reconstructed ancestral areas and ancestral character states. Phylogenetic trees were examined for trait-independent shifts in diversification rates and we used multi-state models to test for state-dependent speciation and extinction rates. Storms could have contributed to Cuba acting as a species pump and to Buxus reaching other Caribbean islands and northern South America'., Key Results: We found a Caribbean Buxus clade with Mexican ancestors, encompassing three major subclades, which started to radiate during the middle Miocene (13.25 Mya). Other Caribbean islands and northern South America were reached from ~3 Mya onwards., Conclusions: An evolutionary scenario is evident in which Buxus plants able to grow on ultramafic substrates by exaptation became ultramafic substrate endemics and evolved stepwise from Ni tolerance through Ni accumulation to Ni hyperaccumulation, which has triggered species diversification of Buxus in Cuba. Storms could have contributed to Cuba acting as a species pump and to Buxus reaching other Caribbean islands and northern South America'., (© The Author(s) 2023. Published by Oxford University Press on behalf of the Annals of Botany Company. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

DNA is compacted into individual particles or chromosomes that form the basic units of inheritance. However, different animals and plants have widely different numbers of chromosomes. This means that we cannot readily tell which chromosomes are related to which. Here, we describe a simple technique that looks at the similarity of genes on each chromosome and thus gives us a true picture of their homology or similarity through evolutionary time. We use this new system to look at the chromosomes of butterflies and moths or Lepidoptera. We term the associated synteny units, Lepidopteran Synteny Units (LSUs). Using a sample of butterfly and moth genomes from across evolutionary time, we show that LSUs form a simple and reliable method of tracing chromosomal homology back through time. Surprisingly, this technique reveals that butterfly and moth chromosomes show conserved blocks dating back to their sister group the Trichoptera. As Lepidoptera have holocentric chromosomes, it will be interesting to see if similar levels of synteny are shown in groups of animals with monocentric chromosomes. The ability to define homology via LSU analysis makes it considerably easier to approach many questions in chromosomal evolution., Competing Interests: Conflicts of interest statement The author(s) declare no conflict of interest., (© The Author(s) 2023. Published by Oxford University Press on behalf of The Genetics Society of America.)

For many years the surprising multiplicity, signal input diversity, and output specificity of c-di-GMP signaling proteins has intrigued researchers studying bacterial second messengers. How can several signaling pathways act in parallel to produce specific outputs despite relying on the same diffusible second messenger maintained at a certain global cellular concentration? Such high specificity and flexibility arise from combining modes of local and global c-di-GMP signaling in complex signaling networks. Local c-di-GMP signaling can be experimentally shown by three criteria being met: (i) highly specific knockout phenotypes for particular c-di-GMP-related enzymes, (ii) actual cellular c-di-GMP levels that remain unchanged by such mutations and/or below the K d 's of the relevant c-di-GMP-binding effectors, and (iii) direct interactions between the signaling proteins involved. Here, we discuss the rationale behind these criteria and present well-studied examples of local c-di-GMP signaling in Escherichia coli and Pseudomonas . Relatively simple systems just colocalize a local source and/or a local sink for c-di-GMP, i.e. a diguanylate cyclase (DGC) and/or a specific phosphodiesterase (PDE), respectively, with a c-di-GMP-binding effector/target system. More complex systems also make use of regulatory protein interactions, e.g. when a "trigger PDE" responds to locally provided c-di-GMP, and thereby serves as a c-di-GMP-sensing effector that directly controls a target's activity, or when a c-di-GMP-binding effector recruits and directly activates its own "private" DGC. Finally, we provide an outlook into how cells can combine local and global signaling modes of c-di-GMP and possibly integrate those into other signaling nucleotides networks., Competing Interests: None declared., (© The Author(s) 2023. Published by Oxford University Press on behalf of FEMS.)

More than 170 posttranscriptional RNA modifications are so far known on both coding and noncoding RNA species. Within this group, pseudouridine (Ψ) and queuosine (Q) represent conserved RNA modifications with fundamental functional roles in regulating translation. Current detection methods of these modifications, which both are reverse transcription (RT)-silent, are mostly based on the chemical treatment of RNA prior to analysis. To overcome the drawbacks associated with indirect detection strategies, we have engineered an RT-active DNA polymerase variant called RT-KTq I614Y that produces error RT signatures specific for Ψ or Q without prior chemical treatment of the RNA samples. Combining this polymerase with next-generation sequencing techniques allows the direct identification of Ψ and Q sites of untreated RNA samples using a single enzymatic tool., (© The Author(s) 2023. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Synthetic small RNAs (sRNAs) are gaining increasing attention in the field of synthetic biology and bioengineering for efficient post-transcriptional regulation of gene expression. However, the optimal design of synthetic sRNAs is challenging because alterations may impair functions or off-target effects can arise. Here, we introduce DIGGER-Bac, a toolbox for Design and Identification of seed regions for Golden Gate assembly and Expression of synthetic sRNAs in Bacteria. The SEEDling tool predicts optimal sRNA seed regions in combination with user-defined sRNA scaffolds for efficient regulation of specified mRNA targets. Results are passed on to the G-GArden tool, which assists with primer design for high-fidelity Golden Gate assembly of the desired synthetic sRNA constructs., (© The Author(s) 2023. Published by Oxford University Press.)

Post-translational modifications on histones are well known to regulate chromatin structure and function, but much less information is available on modifications of the centromeric histone H3 variant and their effect at the kinetochore. Here, we report two modifications on the centromeric histone H3 variant CENP-A/Cse4 in the yeast Saccharomyces cerevisiae, methylation at arginine 143 (R143me) and lysine 131 (K131me), that affect centromere stability and kinetochore function. Both R143me and K131me lie in the core region of the centromeric nucleosome, near the entry/exit sites of the DNA from the nucleosome. Unexpectedly, mutation of Cse4-R143 (cse4-R143A) exacerbated the kinetochore defect of mutations in components of the NDC80 complex of the outer kinetochore (spc25-1) and the MIND complex (dsn1-7). The analysis of suppressor mutations of the spc25-1 cse4-R143A growth defect highlighted residues in Spc24, Ndc80, and Spc25 that localize to the tetramerization domain of the NDC80 complex and the Spc24-Spc25 stalk, suggesting that the mutations enhance interactions among NDC80 complex components and thus stabilize the complex. Furthermore, the Set2 histone methyltransferase inhibited kinetochore function in spc25-1 cse4-R143A cells, possibly by methylating Cse4-K131. Taken together, our data suggest that Cse4-R143 methylation and Cse4-K131 methylation affect the stability of the centromeric nucleosome, which is detrimental in the context of defective NDC80 tetramerization and can be compensated for by strengthening interactions among NDC80 complex components., Competing Interests: Conflict of interest The authors declare that they have no conflict of interest., (© The Author(s) 2023. Published by Oxford University Press on behalf of the Genetics Society of America.)

In the thermal system, skin cooling is represented in the primary somatosensory cortex (S1) and the posterior insular cortex (pIC). Whether S1 and pIC are nodes in anatomically separate or overlapping thermal sensorimotor pathways is unclear, as the brain-wide connectivity of the thermal system has not been mapped. We address this using functionally targeted, dual injections of anterograde viruses or retrograde tracers into the forelimb representation of S1 (fS1) and pIC (fpIC). Our data show that inputs to fS1 and fpIC originate from separate neuronal populations, supporting the existence of parallel input pathways. Outputs from fS1 and fpIC are more widespread than their inputs, sharing a number of cortical and subcortical targets. While, axonal projections were separable, they were more overlapping than the clusters of input cells. In both fS1 and fpIC circuits, there was a high degree of reciprocal connectivity with thalamic and cortical regions, but unidirectional output to the midbrain and hindbrain. Notably, fpIC showed connectivity with regions associated with thermal processing. Together, these data indicate that cutaneous thermal information is routed to the cortex via parallel circuits and is forwarded to overlapping downstream regions for the binding of somatosensory percepts and integration with ongoing behavior., (© The Author(s) 2022. Published by Oxford University Press.)

The molecular heterogeneity of cancer cells contributes to the often partial response to targeted therapies and relapse of disease due to the escape of resistant cell populations. While single-cell sequencing has started to improve our understanding of this heterogeneity, it offers a mostly descriptive view on cellular types and states. To obtain more functional insights, we propose scGeneRAI, an explainable deep learning approach that uses layer-wise relevance propagation (LRP) to infer gene regulatory networks from static single-cell RNA sequencing data for individual cells. We benchmark our method with synthetic data and apply it to single-cell RNA sequencing data of a cohort of human lung cancers. From the predicted single-cell networks our approach reveals characteristic network patterns for tumor cells and normal epithelial cells and identifies subnetworks that are observed only in (subgroups of) tumor cells of certain patients. While current state-of-the-art methods are limited by their ability to only predict average networks for cell populations, our approach facilitates the reconstruction of networks down to the level of single cells which can be utilized to characterize the heterogeneity of gene regulation within and across tumors., (© The Author(s) 2023. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Evolution relies on the availability of genetic diversity for fitness-based selection. However, most deoxyribonucleic acid (DNA) viruses employ DNA polymerases (Pol) capable of exonucleolytic proofreading to limit mutation rates during DNA replication. The relative genetic stability produced by high-fidelity genome replication can make studying DNA virus adaptation and evolution an intensive endeavor, especially in slowly replicating viruses. Here, we present a proofreading-impaired Pol mutant (Y547S) of Marek's disease virus that exhibits a hypermutator phenotype while maintaining unimpaired growth in vitro and wild-type (WT)-like pathogenicity in vivo . At the same time, mutation frequencies observed in Y547S virus populations are 2-5-fold higher compared to the parental WT virus. We find that Y547S adapts faster to growth in originally non-permissive cells, evades pressure conferred by antiviral inhibitors more efficiently, and is more easily attenuated by serial passage in cultured cells compared to WT. Our results suggest that hypermutator viruses can serve as a tool to accelerate evolutionary processes and help identify key genetic changes required for adaptation to novel host cells and resistance to antiviral therapy. Similarly, the rapid attenuation achieved through adaptation of hypermutators to growth in cell culture enables identification of genetic changes underlying attenuation and virulence, knowledge that could practically exploited, e.g. in the rational design of vaccines., (© The Author(s) 2022. Published by Oxford University Press.)

Host-associated bacterial communities play an important role in host fitness and resistance to diseases. Yet, few studies have investigated tripartite interaction between a host, parasite and host-associated bacterial communities in natural settings. Here, we use 16S rRNA gene amplicon sequencing to compare gut- and body- bacterial communities of wild water fleas belonging to the Daphnia longispina complex, between uninfected hosts and those infected with the common and virulent eukaryotic gut parasite Caullerya mesnili (Family: Ichthyosporea). We report community-level changes in host-associated bacteria with the presence of the parasite infection; namely decreased alpha diversity and increased beta diversity at the site of infection, i.e. host gut (but not host body). We also report decreased abundance of bacterial taxa proposed elsewhere to be beneficial for the host, and an appearance of taxa specifically associated with infected hosts. Our study highlights the host-microbiota-infection link in a natural system and raises questions about the role of host-associated microbiota in natural disease epidemics as well as the functional roles of bacteria specifically associated with infected hosts., (© The Author(s) 2022. Published by Oxford University Press on behalf of FEMS.)

Background: Spatial sequencing methods increasingly gain popularity within RNA biology studies. State-of-the-art techniques quantify messenger RNA expression levels from tissue sections and at the same time register information about the original locations of the molecules in the tissue. The resulting data sets are processed and analyzed by accompanying software that, however, is incompatible across inputs from different technologies., Findings: Here, we present spacemake, a modular, robust, and scalable spatial transcriptomics pipeline built in Snakemake and Python. Spacemake is designed to handle all major spatial transcriptomics data sets and can be readily configured for other technologies. It can process and analyze several samples in parallel, even if they stem from different experimental methods. Spacemake's unified framework enables reproducible data processing from raw sequencing data to automatically generated downstream analysis reports. Spacemake is built with a modular design and offers additional functionality such as sample merging, saturation analysis, and analysis of long reads as separate modules. Moreover, spacemake employs novoSpaRc to integrate spatial and single-cell transcriptomics data, resulting in increased gene counts for the spatial data set. Spacemake is open source and extendable, and it can be seamlessly integrated with existing computational workflows., (© The Author(s) 2022. Published by Oxford University Press GigaScience.)

Plants evolved efficient multifaceted acclimation strategies to cope with low temperatures. Chloroplasts respond to temperature stimuli and participate in temperature sensing and acclimation. However, very little is known about the involvement of chloroplast genes and their expression in plant chilling tolerance. Here we systematically investigated cold acclimation in tobacco seedlings over 2 days of exposure to low temperatures by examining responses in chloroplast genome copy number, transcript accumulation and translation, photosynthesis, cell physiology, and metabolism. Our time-resolved genome-wide investigation of chloroplast gene expression revealed substantial cold-induced translational regulation at both the initiation and elongation levels, in the virtual absence of changes at the transcript level. These cold-triggered dynamics in chloroplast translation are widely distinct from previously described high light-induced effects. Analysis of the gene set responding significantly to the cold stimulus suggested nonessential plastid-encoded subunits of photosynthetic protein complexes as novel players in plant cold acclimation. Functional characterization of one of these cold-responsive chloroplast genes by reverse genetics demonstrated that the encoded protein, the small cytochrome b6f complex subunit PetL, crucially contributes to photosynthetic cold acclimation. Together, our results uncover an important, previously underappreciated role of chloroplast translational regulation in plant cold acclimation., (© The Author(s) 2022. Published by Oxford University Press on behalf of American Society of Plant Biologists.)

Soil microbes promote plant growth through several mechanisms such as secretion of chemical compounds including plant growth hormones. Among the phytohormones, auxins, ethylene, cytokinins, abscisic acid and gibberellins are the best understood compounds. Gibberellins were first isolated in 1935 from the fungus Gibberella fujikuroi and are synthesized by several soil microbes. The effect of gibberellins on plant growth and development has been studied, as has the biosynthesis pathways, enzymes, genes and their regulation. This review revisits the history of gibberellin research highlighting microbial gibberellins and their effects on plant health with an emphasis on the early discoveries and current advances that can find vital applications in agricultural practices., (© 2021 The Society for Applied Microbiology.)

Unordered parent-offspring (PO) relationships are an outstanding issue in pedigree reconstruction studies. Resolution of the order of these relationships would expand the results, conclusions, and usefulness of such studies; however, no such PO order resolution (POR) tests currently exist. This study describes two such tests, demonstrated using SNP array data in the outcrossing species apple (Malus × domestica) on a PO relationship of known order ("Keepsake" as a parent of "Honeycrisp") and two PO relationships previously ordered only via provenance information. The first test, POR-1, tests whether some of the extended haplotypes deduced from homozygous SNP calls from one individual in an unordered PO duo are composed of recombinant haplotypes from accurately phased SNP genotypes from the second individual. If so, the first individual would be the offspring of the second individual, otherwise the opposite relationship would be present. The second test, POR-2, does not require phased SNP genotypes and uses similar logic as the POR-1 test, albeit in a different approach. The POR-1 and POR-2 tests determined the correct relationship between "Keepsake" and "Honeycrisp". The POR-2 test confirmed "Reinette Franche" as a parent of "Nonpareil" and "Brabant Bellefleur" as a parent of "Court Pendu Plat". The latter finding conflicted with the recorded provenance information, demonstrating the need for these tests. The successful demonstration of these tests suggests they can add insights to future pedigree reconstruction studies, though caveats, like extreme inbreeding or selfing, would need to be considered where relevant., (© The Author(s) 2022. Published by Oxford University Press. All rights reserved.)

Pedigree information is of fundamental importance in breeding programs and related genetics efforts. However, many individuals have unknown pedigrees. While methods to identify and confirm direct parent-offspring relationships are routine, those for other types of close relationships have yet to be effectively and widely implemented with plants, due to complications such as asexual propagation and extensive inbreeding. The objective of this study was to develop and demonstrate methods that support complex pedigree reconstruction via the total length of identical by state haplotypes (referred to in this study as "summed potential lengths of shared haplotypes", SPLoSH). A custom Python script, HapShared, was developed to generate SPLoSH data in apple and sweet cherry. HapShared was used to establish empirical distributions of SPLoSH data for known relationships in these crops. These distributions were then used to estimate previously unknown relationships. Case studies in each crop demonstrated various pedigree reconstruction scenarios using SPLoSH data. For cherry, a full-sib relationship was deduced for 'Emperor Francis, and 'Schmidt', a half-sib relationship for 'Van' and 'Windsor', and the paternal grandparents of 'Stella' were confirmed. For apple, 29 cultivars were found to share an unknown parent, the pedigree of the unknown parent of 'Cox's Pomona' was reconstructed, and 'Fameuse' was deduced to be a likely grandparent of 'McIntosh'. Key genetic resources that enabled this empirical study were large genome-wide SNP array datasets, integrated genetic maps, and previously identified pedigree relationships. Crops with similar resources are also expected to benefit from using HapShared for empowering pedigree reconstruction., (© 2021. The Author(s).)

Genetic incompatibility between the cytoplasm and the nucleus is thought to be a major factor in species formation, but mechanistic understanding of this process is poor. In evening primroses (Oenothera spp.), a model plant for organelle genetics and population biology, hybrid offspring regularly display chloroplast-nuclear incompatibility. This usually manifests in bleached plants, more rarely in hybrid sterility or embryonic lethality. Hence, most of these incompatibilities affect photosynthetic capability, a trait that is under selection in changing environments. Here we show that light-dependent misregulation of the plastid psbB operon, which encodes core subunits of photosystem II and the cytochrome b6f complex, can lead to hybrid incompatibility, and this ultimately drives speciation. This misregulation causes an impaired light acclimation response in incompatible plants. Moreover, as a result of their different chloroplast genotypes, the parental lines differ in photosynthesis performance upon exposure to different light conditions. Significantly, the incompatible chloroplast genome is naturally found in xeric habitats with high light intensities, whereas the compatible one is limited to mesic habitats. Consequently, our data raise the possibility that the hybridization barrier evolved as a result of adaptation to specific climatic conditions., (© The Author(s) 2021. Published by Oxford University Press on behalf of American Society of Plant Biologists.)

DDX3 is an RNA chaperone of the DEAD-box family that regulates translation. Ded1, the yeast ortholog of DDX3, is a global regulator of translation, whereas DDX3 is thought to preferentially affect a subset of mRNAs. However, the set of mRNAs that are regulated by DDX3 are unknown, along with the relationship between DDX3 binding and activity. Here, we use ribosome profiling, RNA-seq, and PAR-CLIP to define the set of mRNAs that are regulated by DDX3 in human cells. We find that while DDX3 binds highly expressed mRNAs, depletion of DDX3 particularly affects the translation of a small subset of the transcriptome. We further find that DDX3 binds a site on helix 16 of the human ribosomal rRNA, placing it immediately adjacent to the mRNA entry channel. Translation changes caused by depleting DDX3 levels or expressing an inactive point mutation are different, consistent with different association of these genetic variant types with disease. Taken together, this work defines the subset of the transcriptome that is responsive to DDX3 inhibition, with relevance for basic biology and disease states where DDX3 is altered., (© The Author(s) 2021. Published by Oxford University Press on behalf of Nucleic Acids Research.)

There is a growing awareness that traits do not evolve individually but rather are organized as modular networks of covarying traits. Although the importance of multi-trait correlation has been linked to the ability to evolve in response to new environmental conditions, the evolvability of the network itself has to date rarely been assessed experimentally. By following the evolutionary dynamics of a model bacterium adapting to plant roots, we demonstrate that the whole structure of the trait correlation network is highly dynamic. We experimentally evolved Pseudomonas protegens, a common rhizosphere dweller, on the roots of Arabidopsis thaliana. We collected bacteria at regular intervals and determined a range of traits linked to growth, stress resistance, and biotic interactions. We observed a rapid disintegration of the original trait correlation network. Ancestral populations showed a modular network, with the traits linked to resource use and stress resistance forming two largely independent modules. This network rapidly was restructured during adaptation, with a loss of the stress resistance module and the appearance of new modules out of previously disconnected traits. These results show that evolutionary dynamics can involve a deep restructuring of phenotypic trait organization, pointing to the emergence of novel life history strategies not represented in the ancestral phenotype., (© 2021 The Authors. Evolution published by Wiley Periodicals LLC on behalf of The Society for the Study of Evolution.)

We report on a study that aimed at establishing a large soil-fungal culture collection spanning a wide taxonomic diversity and systematically screening the collection for bacterial associations. Fungal cultures were isolated from soil samples obtained from a natural grassland in eastern Germany and bacterial associations were assessed by PCR-amplification and sequencing of bacterial 16S rRNA. In addition, intraspecies genetic diversities of a subset of the isolated species were estimated by double-digest restriction associated DNA sequencing. A total of 688 fungal cultures, representing at least 106 fungal species from 36 different families, were obtained and even though clonal isolates were identified in almost all fungal species subjected to ddRAD-seq, relatively high genetic diversities could be observed in some of the isolated species. A total of 69% of the fungal isolates in our collection were found to be associated with bacteria and the most commonly identified bacterial genera were Pelomonas, Enterobacter and Burkholderia. Our results indicate that bacterial associations commonly occur in soil fungi, even if antibiotics are being applied during the isolation process, and provide a basis for the use of our culture collection in ecological experiments that want to acknowledge the importance of intraspecies genetic diversity., (© The Author(s) 2021. Published by Oxford University Press on behalf of FEMS.)

Although giraffes maintain the usual mammalian cervical number of seven vertebrae, their first thoracic vertebra (T1) exhibits aberrant anatomy and has been hypothesized to functionally elongate the neck. We test this "functional elongation hypothesis" by combining phylogenetically informed analyses of neck length, three-dimensional (3D) vertebral shape, and of the functional significance of shape differences across a broad sample of ruminants and camelids. Digital bone models of the cervicothoracic transition were subjected to 3D geometric morphometric analysis revealing how the shape of the seventh cervical (C7) has converged in several long-necked species. However, we find a unique "cervicalization" of the giraffe's T1. In contrast, we demonstrate a "thoracalization" of C7 for the European bison. Other giraffids (okapi and extinct Sivatherium) did not exhibit "cervicalized" T1 morphology. Quantitative range of motion (ROM) analysis at the cervicothoracic transition in ruminants and camelids confirms the "functional elongation hypothesis" for the giraffe in terms of increased mobility, especially with regard to dorsoventral flexion/extension. Additionally, other factors related to the unique morphology of the giraffe's cervicothoracic transition such as neck posture and intervertebral stability are discussed and should be considered in future studies of giraffe neck evolution., (© 2021 The Authors. Evolution published by Wiley Periodicals LLC on behalf of The Society for the Study of Evolution.)

Convergent evolution is a major topic in evolutionary biology. Low bone cortical compactness (CC, a measure of porosity of cortical bone) in the extant genera of "tree sloths," has been linked to their convergent slow arboreal ecology. This proposed relationship of low CC with a slow arboreal lifestyle suggests potential convergent evolution of this trait in other slow arboreal mammals. Femoral and humeral CC were analyzed in "tree sloths," lorisids, koala, and extinct palaeopropithecids and Megaladapis, in comparison to closely related but ecologically distinct taxa, in a phylogenetic framework. Low CC in "tree sloths" is unparalleled by any analyzed clade and the high CC in extinct sloths suggests the recent convergence of low CC in "tree sloths." A tendency for low CC was found in Palaeopropithecus and Megaladapis. However, lorisids and the koala yielded unexpected CC patterns, preventing the recognition of a straightforward convergence of low CC in slow arboreal mammals. This study uncovers a complex relationship between CC and convergent evolution of slow arboreality, highlighting the multifactorial specificity of bone microstructure., (© 2020 The Authors. Evolution published by Wiley Periodicals LLC on behalf of The Society for the Study of Evolution.)

Despite significant advances in invertebrate phylogenomics over the past decade, the higher-level phylogeny of Pycnogonida (sea spiders) remains elusive. Due to the inaccessibility of some small-bodied lineages, few phylogenetic studies have sampled all sea spider families. Previous efforts based on a handful of genes have yielded unstable tree topologies. Here, we inferred the relationships of 89 sea spider species using targeted capture of the mitochondrial genome, 56 conserved exons, 101 ultraconserved elements, and 3 nuclear ribosomal genes. We inferred molecular divergence times by integrating morphological data for fossil species to calibrate 15 nodes in the arthropod tree of life. This integration of data classes resolved the basal topology of sea spiders with high support. The enigmatic family Austrodecidae was resolved as the sister group to the remaining Pycnogonida and the small-bodied family Rhynchothoracidae as the sister group of the robust-bodied family Pycnogonidae. Molecular divergence time estimation recovered a basal divergence of crown group sea spiders in the Ordovician. Comparison of diversification dynamics with other marine invertebrate taxa that originated in the Paleozoic suggests that sea spiders and some crustacean groups exhibit resilience to mass extinction episodes, relative to mollusk and echinoderm lineages., (© The Author(s) 2020. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.)

In the Elongator-dependent modification pathway, chemical modifications are introduced at the wobble uridines at position 34 in transfer RNAs (tRNAs), which serve to optimize codon translation rates. Here, we show that this three-step modification pathway exists in Dictyostelium discoideum, model of the evolutionary superfamily Amoebozoa. Not only are previously established modifications observable by mass spectrometry in strains with the most conserved genes of each step deleted, but also additional modifications are detected, indicating a certain plasticity of the pathway in the amoeba. Unlike described for yeast, D. discoideum allows for an unconditional deletion of the single tQCUG gene, as long as the Elongator-dependent modification pathway is intact. In gene deletion strains of the modification pathway, protein amounts are significantly reduced as shown by flow cytometry and Western blotting, using strains expressing different glutamine leader constructs fused to GFP. Most dramatic are these effects, when the tQCUG gene is deleted, or Elp3, the catalytic component of the Elongator complex is missing. In addition, Elp3 is the most strongly conserved protein of the modification pathway, as our phylogenetic analysis reveals. The implications of this observation are discussed with respect to the evolutionary age of the components acting in the Elongator-dependent modification pathway., (© The Author(s) 2020. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Deciphering signal transduction processes is crucial for understanding how plants sense and respond to environmental changes. Various chemical compounds function as central messengers within deeply intertwined signaling networks. How such compounds act in concert remains to be elucidated. We have developed dual-reporting transcriptionally linked genetically encoded fluorescent indicators (2-in-1-GEFIs) for multiparametric in vivo analyses of the phytohormone abscisic acid (ABA), Ca 2+ , protons (H + ), chloride (anions), the glutathione redox potential, and H 2 O 2 Simultaneous analyses of two signaling compounds in Arabidopsis ( Arabidopsis thaliana ) roots revealed that ABA treatment and uptake did not trigger rapid cytosolic Ca 2+ or H + dynamics. Glutamate, ATP, Arabidopsis PLANT ELICITOR PEPTIDE, and glutathione disulfide (GSSG) treatments induced rapid spatiotemporally overlapping cytosolic Ca 2+ , H + , and anion dynamics, but except for GSSG, only weakly affected the cytosolic redox state. Overall, 2-in-1-GEFIs enable complementary, high-resolution in vivo analyses of signaling compound dynamics and facilitate an advanced understanding of the spatiotemporal coordination of signal transduction processes in Arabidopsis., (© 2020 American Society of Plant Biologists. All rights reserved.)

Previously, combined loss of different anticodon loop modifications was shown to impair the function of distinct tRNAs in Saccharomyces cerevisiae. Surprisingly, each scenario resulted in shared cellular phenotypes, the basis of which is unclear. Since loss of tRNA modification may evoke transcriptional responses, we characterized global transcription patterns of modification mutants with defects in either tRNAGlnUUG or tRNALysUUU function. We observe that the mutants share inappropriate induction of multiple starvation responses in exponential growth phase, including derepression of glucose and nitrogen catabolite-repressed genes. In addition, autophagy is prematurely and inadequately activated in the mutants. We further demonstrate that improper induction of individual starvation genes as well as the propensity of the tRNA modification mutants to form protein aggregates are diminished upon overexpression of tRNAGlnUUG or tRNALysUUU, the tRNA species that lack the modifications of interest. Hence, our data suggest that global alterations in mRNA translation and proteostasis account for the transcriptional stress signatures that are commonly triggered by loss of anticodon modifications in different tRNAs., (© The Author(s) 2020. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Motivation: The submission of annotated sequence data to public sequence databases constitutes a central pillar in biological research. The surge of novel DNA sequences awaiting database submission due to the application of next-generation sequencing has increased the need for software tools that facilitate bulk submissions. This need has yet to be met with the concurrent development of tools to automate the preparatory work preceding such submissions., Results: The author introduce annonex2embl, a Python package that automates the preparation of complete sequence flatfiles for large-scale sequence submissions to the European Nucleotide Archive. The tool enables the conversion of DNA sequence alignments that are co-supplied with sequence annotations and metadata to submission-ready flatfiles. Among other features, the software automatically accounts for length differences among the input sequences while maintaining correct annotations, automatically interlaces metadata to each record and displays a design suitable for easy integration into bioinformatic workflows. As proof of its utility, annonex2embl is employed in preparing a dataset of more than 1500 fungal DNA sequences for database submission., Availability and Implementation: annonex2embl is freely available via the Python package index at http://pypi.python.org/pypi/annonex2embl., Supplementary Information: Supplementary data are available at Bioinformatics online., (© The Author(s) 2020. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Studies on Glucose-6-phosphate (G6P)/phosphate translocator isoforms GPT1 and GPT2 reported the viability of Arabidopsis ( Arabidopsis thaliana ) gpt2 mutants, whereas heterozygous gpt1 mutants exhibited a variety of defects during fertilization/seed set, indicating that GPT1 is essential for this process. Among other functions, GPT1 was shown to be important for pollen and embryo-sac development. Because our previous work on the irreversible part of the oxidative pentose phosphate pathway (OPPP) revealed comparable effects, we investigated whether GPT1 may dually localize to plastids and peroxisomes. In reporter fusions, GPT2 localized to plastids, but GPT1 also localized to the endoplasmic reticulum (ER) and around peroxisomes. GPT1 contacted two oxidoreductases and also peroxins that mediate import of peroxisomal membrane proteins from the ER, hinting at dual localization. Reconstitution in yeast ( Saccharomyces cerevisiae ) proteoliposomes revealed that GPT1 preferentially exchanges G6P for ribulose-5-phosphate (Ru5P). Complementation analyses of heterozygous +/ gpt1 plants demonstrated that GPT2 is unable to compensate for GPT1 in plastids, whereas GPT1 without the transit peptide (enforcing ER/peroxisomal localization) increased gpt1 transmission significantly. Because OPPP activity in peroxisomes is essential for fertilization, and immunoblot analyses hinted at the presence of unprocessed GPT1-specific bands, our findings suggest that GPT1 is indispensable in both plastids and peroxisomes. Together with its G6P-Ru5P exchange preference, GPT1 appears to play a role distinct from that of GPT2 due to dual targeting., (© 2020 American Society of Plant Biologists. All rights reserved.)

Multiple sequence alignments (MSAs) play a pivotal role in studies of molecular sequence data, but nobody has developed a minimum reporting standard (MRS) to quantify the completeness of MSAs in terms of completely specified nucleotides or amino acids. We present an MRS that relies on four simple completeness metrics. The metrics are implemented in AliStat, a program developed to support the MRS. A survey of published MSAs illustrates the benefits and unprecedented transparency offered by the MRS., (© The Author(s) 2019. Published by Oxford University Press on behalf of NAR Genomics and Bioinformatics.)

Transcriptome analysis by RNA sequencing (RNA-seq) has become an indispensable research tool in modern plant biology. Virtually all RNA-seq studies provide a snapshot of the steady state transcriptome, which contains valuable information about RNA populations at a given time but lacks information about the dynamics of RNA synthesis and degradation. Only a few specialized sequencing techniques, such as global run-on sequencing, have been used to provide information about RNA synthesis rates in plants. Here, we demonstrate that RNA labeling with the modified, nontoxic uridine analog 5-ethynyl uridine (5-EU) in Arabidopsis ( Arabidopsis thaliana ) seedlings provides insight into plant transcriptome dynamics. Pulse labeling with 5-EU revealed nascent and unstable RNAs, RNA processing intermediates generated by splicing, and chloroplast RNAs. Pulse-chase experiments with 5-EU allowed us to determine RNA stabilities without the need for chemical transcription inhibitors such as actinomycin and cordycepin. Inhibitor-free, genome-wide analysis of polyadenylated RNA stability via 5-EU pulse-chase experiments revealed RNAs with shorter half-lives than those reported after chemical inhibition of transcription. In summary, our results indicate that the Arabidopsis nascent transcriptome contains unstable RNAs and RNA processing intermediates and suggest that polyadenylated RNAs have low stability in plants. Our technique lays the foundation for easy, affordable, nascent transcriptome analysis and inhibitor-free analysis of RNA stability in plants., (© 2020 American Society of Plant Biologists. All rights reserved.)

Denitrification is an ecosystem process linked to ongoing climate change, because it releases nitrous oxide (N2O) into the atmosphere. To date, the literature covers mostly how aboveground (i.e. plant community structure) and belowground (i.e. plant-associated soil microbes) biota separately influence denitrification in isolation of each other. We here present a mesocosm experiment where we combine a manipulation of belowground biota (i.e. addition of Rhizophagus irregularis propagules to the indigenous mycorrhizal community) with a realized gradient in plant diversity. We used a seed mix containing plant species representative of mesophytic European grasslands and by stochastic differences in species establishment across the sixteen replicates per treatment level a spontaneously established gradient in plant diversity. We address mycorrhizal-induced and plant-diversity mediated changes on denitrification potential parameters and how these differ from the existing literature that studies them independently of each other. We show that unlike denitrification potential, N2O potential emissions do not change with mycorrhiza and depend instead on realized plant diversity. By linking mycorrhizal ecology to an N-cycling process, we present a comprehensive assessment of terrestrial denitrification dynamics when diverse plants co-occur., (© FEMS 2019.)

Among many glycoproteins within the plant secretory system, KORRIGAN1 (KOR1), a membrane-anchored endo-β-1,4-glucanase involved in cellulose biosynthesis, provides a link between N- glycosylation, cell wall biosynthesis, and abiotic stress tolerance. After insertion into the endoplasmic reticulum, KOR1 cycles between the trans -Golgi network (TGN) and the plasma membrane (PM). From the TGN, the protein is targeted to growing cell plates during cell division. These processes are governed by multiple sequence motifs and also host genotypes. Here, we investigated the interaction and hierarchy of known and newly identified sorting signals in KOR1 and how they affect KOR1 transport at various stages in the secretory pathway. Conventional steady-state localization showed that structurally compromised KOR1 variants were directed to tonoplasts. In addition, a tandem fluorescent timer technology allowed for differential visualization of young versus aged KOR1 proteins, enabling the analysis of single-pass transport through the secretory pathway. Observations suggest the presence of multiple checkpoints/branches during KOR1 trafficking, where the destination is determined based on KOR1's sequence motifs and folding status. Moreover, growth analyses of dominant PM-confined KOR1-L 48 L 49 →A 48 A 49 variants revealed the importance of active removal of KOR1 from the PM during salt stress, which otherwise interfered with stress acclimation., (© 2020 American Society of Plant Biologists. All rights reserved.)

Recent work revealed that PGD2, an Arabidopsis 6-phosphogluconate dehydrogenase (6-PGD) catalysing the third step of the oxidative pentose-phosphate pathway (OPPP) in peroxisomes, is essential during fertilization. Earlier studies on the second step, catalysed by PGL3, a dually targeted Arabidopsis 6-phosphogluconolactonase (6-PGL), reported the importance of OPPP reactions in plastids but their irrelevance in peroxisomes. Assuming redundancy of 6-PGL activity in peroxisomes, we examined the sequences of other higher plant enzymes. In tomato, there exist two 6-PGL isoforms with the strong PTS1 motif SKL. However, their analysis revealed problems regarding peroxisomal targeting: reporter-PGL detection in peroxisomes required construct modification, which was also applied to the Arabidopsis isoforms. The relative contribution of PGL3 versus PGL5 during fertilization was assessed by mutant crosses. Reduced transmission ratios were found for pgl3-1 (T-DNA-eliminated PTS1) and also for knock-out allele pgl5-2. The prominent role of PGL3 showed as compromised growth of pgl3-1 seedlings on sucrose and higher activity of mutant PGL3-1 versus PGL5 using purified recombinant proteins. Evidence for PTS1-independent uptake was found for PGL3-1 and other Arabidopsis PGL isoforms, indicating that peroxisome import may be supported by a piggybacking mechanism. Thus, multiple redundancy at the level of the second OPPP step in peroxisomes explains the occurrence of pgl3-1 mutant plants., (© The Author(s) 2019. Published by Oxford University Press on behalf of the Society for Experimental Biology.)

Locomotor habits in mammals are strongly tied to limb bones' lengths, diameters, and proportions. By comparison, fewer studies have examined how limb bone cross-sectional traits relate to locomotor habit. Here, we tested whether climbing, digging, and swimming locomotor habits reflect biomechanically meaningful differences in three cross-sectional traits rendered dimensionless- cross-sectional area (CSA), second moments of area (SMA), and section modulus (MOD)-using femora, tibiae, and fibulae of 28 species of mustelid. CSA and SMA represent resistance to axial compression and bending, respectively, whereas MOD represents structural strength. Given the need to counteract buoyancy in aquatic environments and soil's high density, we predicted that natatorial and fossorial mustelids have higher values of cross-sectional traits. For all three traits, we found that natatorial mustelids have the highest values, followed by fossorial mustelids, with both of these groups significantly differing from scansorial mustelids. However, phylogenetic relatedness strongly influences diversity in cross-sectional morphology, as locomotor habit strongly correlates with phylogeny. Testing whether hind limb bone cross-sectional traits have evolved adaptively, we fit Ornstein-Uhlenbeck (OU) and Brownian motion (BM) models of trait diversification to cross-sectional traits. The cross-sectional traits of the femur, tibia, and fibula appear to have, respectively, diversified under a multi-rate BM model, a single rate BM model, and a multi-optima OU model. In light of recent studies on mustelid body size and elongation, our findings suggest that the mustelid body plan-and perhaps that of other mammals-is likely the sum of a suite of traits evolving under different models of trait diversification., (© The Author(s) 2020. Published by Oxford University Press on behalf of the Society for Integrative and Comparative Biology.)

Seeds of Arabidopsis contain ~40% oil, which is primarily in the form of triacylglycerol and it is converted to sugar to support post-germination growth. We identified an Arabidopsis T-DNA knockout mutant that is sugar-dependent during early seedling establishment and determined that the β-oxidation process involved in catabolising the free fatty acids released from the seed triacylglycerol is impaired. The mutant was confirmed to be transcriptional null for Protein Acyl Transferase 15, AtPAT15 (At5g04270), one of the 24 protein acyl transferases in Arabidopsis. Although it is the shortest, AtPAT15 contains the signature 'Asp-His-His-Cys cysteine-rich domain' that is essential for the enzyme activity of this family of proteins. The function of AtPAT15 was validated by the fact that it rescued the growth defect of the yeast protein acyl transferase mutant akr1 and it was also auto-acylated in vitro. Transient expression in Arabidopsis and tobacco localised AtPAT15 in the Golgi apparatus. Taken together, our data demonstrate that AtPAT15 is involved in β-oxidation of triacylglycerol, revealing the importance of protein S-acylation in the breakdown of seed-storage lipids during early seedling growth of Arabidopsis., (© The Author(s) 2019. Published by Oxford University Press on behalf of the Society for Experimental Biology. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Summary: Measured kinetic constants are key input data for metabolic models, but they are often uncertain, inconsistent and incomplete. Parameter balancing translates such data into complete and consistent parameter sets while accounting for predefined ranges and physical constraints. Based on Bayesian regression, it determines a most plausible parameter set as well as uncertainty ranges for all model parameters. Our tools for parameter balancing support standard model and data formats and enable an easy customization of prior distributions and constraints for biochemical constants. Modellers can balance kinetic constants, thermodynamic data and metabolomic data to obtain thermodynamically consistent metabolic states that comply with user-defined flux directions., Availability and Implementation: An online tool for parameter balancing, a stand-alone Python command line tool, a Python package and a Matlab toolbox (which uses the CPLEX solver) are freely available at www.parameterbalancing.net., (© The Author(s) 2019. Published by Oxford University Press.)

Pollinators use multiple cues whilst foraging including direct cues from flowers and indirect cues from other pollinators. The use of indirect social cues is common in social insects, such as honeybees and bumblebees, where a social environment facilitates the ability to use such cues. Bumblebees use cues to forage on flowers according to previous foraging experiences. Flowers are an essential food source for pollinators but also pose a high risk of parasite infection through the shared use of flowers leading to parasite spillover. Nevertheless, bumblebees have evolved behavioral defense mechanisms to limit parasite infection by avoiding contaminated flowers. Mechanisms underlying the avoidance of contaminated flowers by bumblebees are poorly understood. Bumblebees were recorded having the choice to forage on non-contaminated flowers and flowers contaminated by a trypan osome gut parasite, Crithidia bombi . The use of different treatments with presence or absence of conspecifics on both contaminated and non-contaminated flowers allowed to investigate the role of social visual cues on their pathogen avoidance behavior. Bumblebees are expected to use social visual cues to avoid contaminated flowers. Our study reveals that the presence of a conspecific on flowers either contaminated or not does not help bumblebee foragers avoiding contaminated flowers. Nevertheless, bumblebees whereas gaining experience tend to avoid their conspecific when placed on contaminated flower and copy it when on the non-contaminated flower. Our experiment suggests a detrimental impact of floral scent on disease avoidance behavior.

Saline-alkali soil is a major environmental constraint impairing plant growth and crop productivity. In this study, we identified a Ca 2+ sensor/kinase/plasma membrane (PM) H + -ATPase module as a central component conferring alkali tolerance in Arabidopsis ( Arabidopsis thaliana ). We report that the SCaBP3 (SOS3-LIKE CALCIUM BINDING PROTEIN3)/CBL7 (CALCINEURIN B-LIKE7) loss-of-function plants exhibit enhanced stress tolerance associated with increased PM H + -ATPase activity and provide fundamental mechanistic insights into the regulation of PM H + -ATPase activity. Consistent with the genetic evidence, interaction analyses, in vivo reconstitution experiments, and determination of H + -ATPase activity indicate that interaction of the Ca 2+ sensor SCaBP3 with the C-terminal Region I domain of the PM H + -ATPase AHA2 ( Arabidopsis thaliana PLASMA MEMBRANE PROTON ATPASE2) facilitates the intramolecular interaction of the AHA2 C terminus with the Central loop region of the PM H + -ATPase to promote autoinhibition of H + -ATPase activity. Concurrently, direct interaction of SCaPB3 with the kinase PKS5 (PROTEIN KINASE SOS2-LIKE5) stabilizes the kinase-ATPase interaction and thereby fosters the inhibitory phosphorylation of AHA2 by PKS5. Consistently, yeast reconstitution experiments and genetic analysis indicate that SCaBP3 provides a bifurcated pathway for coordinating intramolecular and intermolecular inhibition of PM H + -ATPase. We propose that alkaline stress-triggered Ca 2+ signals induce SCaBP3 dissociation from AHA2 to enhance PM H + -ATPase activity. This work illustrates a versatile signaling module that enables the stress-responsive adjustment of plasma membrane proton fluxes., (© 2019 American Society of Plant Biologists. All rights reserved.)

Posttranscriptional RNA modifications occur in all domains of life. Modifications of anticodon bases are of particular importance for ribosomal decoding and proteome homeostasis. The Elongator complex modifies uridines in the wobble position and is highly conserved in eukaryotes. Despite recent insights into Elongator's architecture, the structure and function of its regulatory factor Kti12 have remained elusive. Here, we present the crystal structure of Kti12's nucleotide hydrolase domain trapped in a transition state of ATP hydrolysis. The structure reveals striking similarities to an O-phosphoseryl-tRNA kinase involved in the selenocysteine pathway. Both proteins employ similar mechanisms of tRNA binding and show tRNASec-dependent ATPase activity. In addition, we demonstrate that Kti12 binds directly to Elongator and that ATP hydrolysis is crucial for Elongator to maintain proper tRNA anticodon modification levels in vivo. In summary, our data reveal a hitherto uncharacterized link between two translational control pathways that regulate selenocysteine incorporation and affect ribosomal tRNA selection via specific tRNA modifications., (© The Author(s) 2019. Published by Oxford University Press on behalf of Nucleic Acids Research.)

In eukaryotes, the wobble position of tRNA with a GUN anticodon is modified to the 7-deaza-guanosine derivative queuosine (Q34), but the original source of Q is bacterial, since Q is synthesized by eubacteria and salvaged by eukaryotes for incorporation into tRNA. Q34 modification stimulates Dnmt2/Pmt1-dependent C38 methylation (m5C38) in the tRNAAsp anticodon loop in Schizosaccharomyces pombe. Here, we show by ribosome profiling in S. pombe that Q modification enhances the translational speed of the C-ending codons for aspartate (GAC) and histidine (CAC) and reduces that of U-ending codons for asparagine (AAU) and tyrosine (UAU), thus equilibrating the genome-wide translation of synonymous Q codons. Furthermore, Q prevents translation errors by suppressing second-position misreading of the glycine codon GGC, but not of wobble misreading. The absence of Q causes reduced translation of mRNAs involved in mitochondrial functions, and accordingly, lack of Q modification causes a mitochondrial defect in S. pombe. We also show that Q-dependent stimulation of Dnmt2 is conserved in mice. Our findings reveal a direct mechanism for the regulation of translational speed and fidelity in eukaryotes by a nutrient originating from bacteria., (© The Author(s) 2019. Published by Oxford University Press on behalf of Nucleic Acids Research.)

The nematode Caenorhabditis elegans has been central to the understanding of metazoan biology. However, C. elegans is but one species among millions and the significance of this important model organism will only be fully revealed if it is placed in a rich evolutionary context. Global sampling efforts have led to the discovery of over 50 putative species from the genus Caenorhabditis , many of which await formal species description. Here, we present species descriptions for 10 new Caenorhabditis species. We also present draft genome sequences for nine of these new species, along with a transcriptome assembly for one. We exploit these whole-genome data to reconstruct the Caenorhabditis phylogeny and use this phylogenetic tree to dissect the evolution of morphology in the genus. We reveal extensive variation in genome size and investigate the molecular processes that underlie this variation. We show unexpected complexity in the evolutionary history of key developmental pathway genes. These new species and the associated genomic resources will be essential in our attempts to understand the evolutionary origins of the C. elegans model.

Protists are the most important predators of soil microbes like bacteria and fungi and are highly diverse in terrestrial ecosystems. However, the structure of protistan communities throughout the soil profile is still poorly explored. Here, we used Illumina sequencing to track differences in the relative abundance and diversity of Cercozoa, a major group of protists, at two depths; 10-30 cm (topsoil) and 60-75 cm (subsoil) in an agricultural field in Germany. At the two depths, we also distinguished among three soil compartments: rhizosphere, drilosphere (earthworm burrows) and bulk soil. With increasing depth, we found an overall decline in richness, but we were able to detect subsoil specific phylotypes and contrasting relative abundance patterns between topsoil and subsoil for different clades. We also found that the compartment effect disappeared in the subsoil when compared to the topsoil. More studies are now needed to describe and isolate these possibly subsoil specific phylotypes and better understand their ecology and function., (© FEMS 2019.)

Long-term suppression of recombination ultimately leads to gene loss, as demonstrated by the depauperate Y and W chromosomes of long-established pairs of XY and ZW chromosomes. The young social supergene of the Solenopsis invicta red fire ant provides a powerful system to examine the effects of suppressed recombination over a shorter timescale. The two variants of this supergene are carried by a pair of heteromorphic chromosomes, referred to as the social B and social b (SB and Sb) chromosomes. The Sb variant of this supergene changes colony social organization and has an inheritance pattern similar to a Y or W chromosome because it is unable to recombine. We used high-resolution optical mapping, k-mer distribution analysis, and quantification of repetitive elements on haploid ants carrying alternate variants of this young supergene region. We find that instead of shrinking, the Sb variant of the supergene has increased in length by more than 30%. Surprisingly, only a portion of this length increase is due to consistent increases in the frequency of particular classes of repetitive elements. Instead, haplotypes of this supergene variant differ dramatically in the amounts of other repetitive elements, indicating that the accumulation of repetitive elements is a heterogeneous and dynamic process. This is the first comprehensive demonstration of degenerative expansion in an animal and shows that it occurs through nonlinear processes during the early evolution of a region of suppressed recombination., (© The Author 2018. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.)

Potassium and nitrogen are essential nutrients for plant growth and development. Plants can sense potassium nitrate (K + /NO 3 - ) levels in soils, and accordingly they adjust root-to-shoot K + /NO 3 - transport to balance the distribution of these ions between roots and shoots. In this study, we show that the transcription factorMYB59 maintains this balance by regulating the transcription of the Arabidopsis ( Arabidopsis thaliana ) Nitrate Transporter1.5 ( NRT1.5 )/ Nitrate Transporter/Peptide Transporter Family7.3 ( NPF7.3 ) in response to low K + (LK) stress. The myb59 mutant showed a yellow-shoot sensitive phenotype when grown on LK medium. Both the transcript and protein levels of NPF7.3 were remarkably reduced in the myb59 mutant. LK stress repressed transcript levels of both MYB59 and NPF7.3 The npf7.3 and myb59 mutants, as well as the npf7.3 myb59 double mutant, showed similar LK-sensitive phenotypes. Ion content analyses indicated that root-to-shoot K + /NO 3 - transport was significantly reduced in these mutants under LK conditions. Moreover, chromatin immunoprecipitation and electrophoresis mobility shift assay assays confirmed that MYB59 bound directly to the NPF7.3 promoter. Expression of NPF7.3 in root vasculature driven by the PHOSPHATE 1 promoter rescued the sensitive phenotype of both npf7.3 and myb59 mutants. Together, these data demonstrate that MYB59 responds to LK stress and directs root-to-shoot K + /NO 3 - transport by regulating the expression of NPF7.3 in Arabidopsis roots., (© 2019 American Society of Plant Biologists. All rights reserved.)

Many experiments that measure the response of microbial communities to heavy metals increase metal concentrations abruptly in the soil. However, it is unclear whether abrupt additions mimic the gradual and often long-term accumulation of these metals in the environment where microbial populations may adapt. In a greenhouse experiment that lasted 26 months, we tested whether bacterial communities and soil respiration differed between soils that received an abrupt or a gradual addition of copper or no copper at all. Bacterial richness and other diversity indices were consistently lower in the abrupt treatment compared to the ambient treatment that received no copper. The abrupt addition of copper yielded different initial bacterial communities than the gradual addition; however, these communities appeared to converge once copper concentrations were approximately equal. Soil respiration in the abrupt treatment was initially suppressed but recovered after four months. Afterwards, respiration in both the gradual and abrupt treatments wavered between being below or equal to the ambient treatment. Overall, our study indicates that gradual and abrupt additions of copper can yield similar bacterial communities and respiration, but these responses may drastically vary until copper concentrations are equal.