bluuebunny/crossref_metadata_2025_split huggingface dataset:
The interactome of the UapA transporter reveals putative new players in anterograde membrane cargo trafficking
Authors: Georgiou Xenia , Dimou Sofia , Diallinas George , Samiotaki Martina 
Published: July 2023

AbstractNeosynthesized plasma membrane (PM) proteins co-translationally translocate to the ER, concentrate at regions called ER-exit sites (ERes) and pack into COPII secretory vesicles which are sorted to the early-Golgi through membrane fusion. Following Golgi maturation, membrane cargoes reach the late-Golgi, from where they exit in clathrin-coated vesicles destined to the PM, directly or through endosomes. Post-Golgi membrane cargo trafficking also involves the cytoskeleton and the exocyst. The Golgi-dependent secretory pathway is thought to be responsible for the trafficking of all major membrane proteins. However, our recent findings inAspergillus nidulansshowed that several plasma membrane cargoes, such as transporters and receptors, follow a sorting route that seems to bypass Golgi functioning. To gain insight on membrane trafficking and specifically Golgi-bypass, here we used proximity dependent biotinylation (PDB) coupled with data-independent acquisition mass spectrometry (DIA-MS) for identifying transient interactors of the UapA transporter. Our assays, which included proteomes of wild-type and mutant strains affecting ER-exit or endocytosis, identified both expected and novel interactions that might be physiologically relevant to UapA trafficking. Among those, we validated, using reverse genetics and fluorescence microscopy, that COPI coatomer is essential for ER-exit and anterograde trafficking of UapA and other membrane cargoes. We also showed that ArfAArf1GTPase activating protein (GAP) Glo3 contributes to UapA trafficking at increased temperature. This is the first report addressing the identification of transient interactions during membrane cargo biogenesis using PDB and proteomics coupled with fungal genetics. Our work provides a basis for dissecting dynamic membrane cargo trafficking via PDB assays.HighlightsA PDB-proteomics system for identifying transient interactions inA.nidulansUapA interactomes in wild-type and specific mutantsAnalysis and validation of UapA interacting proteinsCOPI and Arf1 are essential for cargo anterograde traffickingGraphical Abstract
BMP-dependent patterning of ectoderm tissue material properties modulates lateral mesendoderm cell migration during early zebrafish gastrulation
Authors: Tavano Stefania , Brückner David B. , Tasciyan Saren , Tong Xin , Kardos Roland , Schauer Alexandra , Hauschild Robert , Heisenberg Carl-Philipp 
Published: July 2023

SummaryCell migration is a fundamental process during embryonic development. Most studies on cell migrationin vivohave focussed on the migration of cells using the extracellular matrix (ECM) as their substrate for migration. In contrast, much less is known about how cells migrate on other cells, as found in early embryos when the ECM has not yet formed. Here, we show that lateral mesendoderm (LME) cells in the early zebrafish gastrula use the overlying ectoderm as their substrate for migration. We show that the lateral ectoderm is permissive for the animal pole-directed migration of LME cells, while the ectoderm at the animal pole of the gastrula halts animal pole-directed LME migration. These differences in the permissive properties of the ectoderm for LME migration are due to the lateral ectoderm being more cohesive and viscous than the animal ectoderm. Consistently, tuning cell contractility in lateral and animal ectoderm via modulation of the actomyosin cytoskeleton is sufficient to change their permissiveness for LME migration. Finally, we found that BMP signalling is critical for reducing animal ectoderm cohesion and, thus, its capacity to halt LME migration towards the animal pole. Collectively, these findings identify a critical role of ectoderm tissue cohesion and viscosity in guiding LME migration during zebrafish gastrulation.
Nucleotide-protocell interactions: A reciprocal relationship in prebiotically pertinent environments
Authors: Deshpande Kshitij , Mulewar Sahil Sunil , Rajamani Sudha 
Published: July 2023

AbstractSpontaneous interactions between nucleotides and lipid membranes are likely to have played a prominent role in the emergence of life on Earth. However, the effect of nucleotides on the physicochemical properties of model protocellular membranes is relatively less understood. To this end, we aimed to discern the effect of canonical nucleotides on the properties of single-chain amphiphile membranes under prebiotically relevant conditions of multiple wet-dry cycles. Furthermore, the change in critical aggregation concentration of the membranes, and their stability in the presence of nucleotides, was also investigated in Astrobiologically analogue environments. We report that different nucleotides, lipid headgroups, and the ionic makeup of the system affect lipid-nucleotide interactions, which in turn can modulate the effect of nucleotides on the membranes. Specifically, oleic acid membranes seemingly showed an increase in the critical aggregation concentration, and an increase in the stability against prebiotically relevant selection pressures, in the presence of certain nucleotides. Our work has implications for understanding how nucleotides might have shaped the protocellular landscape of the prebiotic Earth.
Genome Sequence and Annotation the B3 Mycobacteriophage Phayeta
Authors: Bishop Emily , Earlely Warren , Greco Alexandra , Hofseth Emma , Kinerson Emma , Lafayette Brandon , Llanot-Arocho Nestor , Mazen Brittney , Cevasco Megan , Williams Daniel C. 
Published: July 2023

AbstractPhayeta is a siphovirus extracted from soil near Myrtle Beach, South Carolina which infectsMycobacterium smegmatis. Annotation of the 68,700 base-pair circularly permuted genome identified 104 predicted protein-encoding genes, 36 of which have functional assignments.
Cdc42 mobility and membrane flows regulate fission yeast cell shape and survival
Authors: Rutkowski David M. , Vincenzetti Vincent , Vavylonis Dimitrios , Martin Sophie G. 
Published: July 2023

AbstractLocal Cdc42 GTPase activation promotes polarized exocytosis, resulting in membrane flows that deplete low-mobility membrane-associated proteins from the growth region. To investigate the self-organizing properties of the Cdc42 secretion-polarization system under membrane flow, we developed a reaction-diffusion particle model. The model includes positive feedback activation of Cdc42, hydrolysis by GTPase-activating proteins (GAPs), and flow-induced displacement by exo/endocytosis. Simulations show how polarization relies on flow-induced depletion of low mobility GAPs. To probe the role of Cdc42 mobility in the fission yeastSchizosaccharomyces pombe, we changed its membrane binding properties by replacing its prenylation site with 1, 2 or 3 repeats of the Rit1 C terminal membrane binding domain (ritC), yielding alleles with progressively lower unbinding and diffusion rates. Concordant modelling predictions and experimental observations show that lower Cdc42 mobility results in lower Cdc42 activation level and wider patches. Indeed, while Cdc42-1ritC cells are viable and polarized, Cdc42-2ritC polarize poorly and Cdc42-3ritC is inviable. The model further predicts that GAP depletion increases Cdc42 activity at the expense of loss of polarization. Experiments confirm this prediction, as deletion of Cdc42 GAPs restores viability to Cdc42-3ritC cells. Our combined experimental and modelling studies demonstrate how membrane flows are an integral part of Cdc42-driven pattern formation.Significance StatementThe delivery of new membrane from internal pools at zones of polarized secretion induces in-plane plasma membrane flows that displace slowly mobile membrane-associated proteins from the zone of secretion. However, zones of polarized secretion are themselves specified by the activity of membrane-associated polarity factors, such as the small GTPase Cdc42. Through combined modelling and experimental approaches, this work demonstrates that the fast mobility of the Cdc42 GTPase is critical to allow the establishment and maintenance of a polarity patch, which is reinforced by flow-mediated displacement of a negative regulator.
Structure of a tripartite protein complex that targets toxins to the type VII secretion system
Authors: Klein Timothy A. , Shah Prakhar Y. , Gkragkopoulou Polyniki , Grebenc Dirk W. , Kim Youngchang , Whitney John C. 
Published: July 2023

AbstractType VII secretion systems are membrane-embedded nanomachines used by Gram-positive bacteria to export effector proteins from the cytoplasm to the extracellular environment. Many of these effectors are polymorphic toxins comprised of an N-terminal Leu-x-Gly (LXG) domain of unknown function and a C-terminal toxin domain that inhibits the growth of bacterial competitors. In recent work, it was shown that LXG effectors require two cognate Lap proteins for T7SS-dependent export. Here, we present the 2.6Å structure of the LXG domain of the TelA toxin from the opportunistic pathogenStreptococcus intermediusin complex with both of its cognate Lap targeting factors. The structure reveals an elongated α-helical bundle within which each Lap protein makes extensive hydrophobic contacts with either end of the LXG domain. Remarkably, despite low overall sequence identity, we identify striking structural similarity between our LXG complex and PE-PPE heterodimers exported by the distantly related ESX type VII secretion systems of Mycobacteria implying a conserved mechanism of effector export among diverse Gram-positive bacteria. Overall, our findings demonstrate that LXG domains, in conjunction with their cognate Lap targeting factors, represent a tripartite secretion signal for a widespread family of T7SS toxins.
Macrophage Xanthine Oxidoreductase Links LPS Induced Lung Inflammatory Injury to NLRP3 Inflammasome Expression and Mitochondrial Respiration
Authors: Fini Mehdi A. , Monks Jenifer A. , Li Min , Gerasimovskaya Evgenia , Paucek Petr , Wang Kepeng , Frid Maria G. , Pugliese Steven C. , Bratton Donna , Yu Yen-Rei , Irwin David , Karin Michael , Wright Richard M. , Stenmark Kurt R. 
Published: July 2023

SUMMARYAcute lung injury (ALI) and the acute respiratory distress syndrome (ARDS) remain poorly treated inflammatory lung disorders. Both reactive oxygen species (ROS) and macrophages are involved in the pathogenesis of ALI/ARDS. Xanthine oxidoreductase (XOR) is an ROS generator that plays a central role in the inflammation that contributes to ALI. To elucidate the role of macrophage-specific XOR in endotoxin induced ALI, we developed a conditional myeloid specific XOR knockout in mice. Myeloid specific ablation of XOR in LPS insufflated mice markedly attenuated lung injury demonstrating the essential role of XOR in this response. Macrophages from myeloid specific XOR knockout exhibited loss of inflammatory activation and increased expression of anti-inflammatory genes/proteins. Transcriptional profiling of whole lung tissue of LPS insufflated XORfl/fl//LysM-Cremice demonstrated an important role for XOR in expression and activation of the NLRP3 inflammasome and acquisition of a glycolytic phenotype by inflammatory macrophages. These results identify XOR as an unexpected link between macrophage redox status, mitochondrial respiration and inflammatory activation.
Generation of an induced pluripotent stem cell line from a healthy adult indigenous Nigerian participant
Authors: Muhammad Zaid , Brown Phoebe W. , Babazau Larema , Alkhamis Abdulrahman I. , Goni Baba W. , Nggada Haruna A. , Mbaya Kefas M. , Wray Selina , Marte Isa H. , Karch Celeste M. , Serpell Louise C. , Maina Mahmoud B. 
Published: July 2023

AbstractGenetic backgrounds contribute to cellular phenotypes, drug responsiveness, and health outcomes. However, the majority of human induced pluripotent stem cell (iPSC) lines are derived from individuals of European descent. Thus, there is a major, unmet need in the generation, characterisation, and distribution of iPSCs from diverse ancestries. To begin to address this need, we have generated iPSCs from dermal fibroblasts isolated from a healthy 60-year-old indigenous Nigerian male belonging to the Babur ethnic group. The iPSCs were generated using Sendai virus, and copy number variation (CNV) analysis revealed no new major abnormalities compared to the parental fibroblasts. The iPSCs have been characterised for pluripotency markers and morphology and successfully differentiated into neural progenitor cells and astrocytes. This iPSC line could serve as a healthy control in comparative studies and can be used in disease modelling, toxicity assessments, genetic analyses, and drug discovery processes within an African genetic background. To bolster the inclusion of African models in biomedical research, this iPSC line will be made available to the broader scientific community. Ongoing efforts focus on generating more lines from diverse indigenous populations towards creating a dedicated open-access African iPSC biobank.
Dual regulation of Misshapen by Tao and Rap2l promotes collective cell migration
Authors: Roberto Gabriela Molinari , Boutet Alison , Keil Sarah , Emery Gregory 
Published: July 2023

ABSTRACTCollective cell migration occurs in various biological processes such as development, wound healing and metastasis. During Drosophila oogenesis, border cells (BC) form a cluster that migrates collectively inside the egg chamber. The Ste20-like kinase Misshapen (Msn) is a key regulator of BC migration coordinating the restriction of protrusion formation and contractile forces within the cluster. Here, we demonstrate that the kinase Tao acts as an upstream activator of Msn in BCs. Depletion of Tao significantly impedes BC migration and produces a phenotype similar to Msn loss-of-function. Furthermore, we show that the localization of Msn relies on its CNH domain, which interacts with the small GTPase Rap2l. Our findings indicate that Rap2l promotes the trafficking of Msn to the endolysosomal pathway. When Rap2l is depleted, the levels of Msn increase in the cytoplasm and at cell-cell junctions between BCs. Overall, our data suggest that Rap2l ensures that the levels of Msn are higher at the periphery of the cluster through the targeting of Msn to the degradative pathway. Together, we identified two distinct regulatory mechanisms that ensure the appropriate distribution and activation of Msn in BCs.
Listeria monocytogenesinfection in intestinal epithelial Caco-2 cells with exposure to progesterone and estradiol-17beta in a gestational infection model
Authors: Hugon Anna Marie , Golos Thaddeus G. 
Published: July 2023

AbstractListeria monocytogenes (Lm) is a food-borne pathogen associated with serious pregnancy complications, including miscarriage, stillbirth, preterm birth, neonatal sepsis, and meningitis. Although Lm infection within the gastrointestinal (GI) tract is well studied, little is known about the influence sex hormones may have on listeriosis. Estradiol (E2) and progesterone (P4) not only have receptors within the GI tract but are significantly increased during pregnancy. The presence of these hormones may play a role in susceptibility to listeriosis during pregnancy.Caco-2 cell monolayers were grown on trans-well inserts in the presence of E2, P4, both E2 and P4, or no hormones (control). Cells were inoculated with Lm for 1 hour, before rinsing with gentamycin and transfer to fresh media. Trans-epithelial resistance was recorded hourly, and bacterial burden of the apical media, intracellular lysates, and basal media were assessed at 6 hours post inoculation.There were no significant differences in bacterial replication when directly exposed to sex steroids, and Caco-2 cell epithelial barrier function was not impacted during culture with Lm. Addition of P4 significantly reduced intracellular bacterial burden compared to E2 only and no hormone controls. Interestingly, E2 only treatment was associated with significantly increased Lm within the basal compartment, compared to reduction in the intracellular and apical layers.These data indicate that increased circulating sex hormones alone do not significantly impact intestinal epithelial barrier integrity during listeriosis, but that addition of P4 and E2, alone or in combination, was associated with reduced epithelial cell bacterial burden and apical release of Lm.Summary sentenceProgesterone and estradiol inhibit infection of Caco-2 intestinal epithelial cells byListeria monocytogenes.
SYS-1/beta-catenin inheritance and regulation by Wnt-signaling during asymmetric cell division
Authors: Valdes Michel Maria F. , Phillips Bryan T. 
Published: July 2023

AbstractAsymmetric cell division (ACD) allows daughter cells of a polarized mother to acquire different developmental fates. InC. elegans, the Wnt/β-catenin Asymmetry (WβA) pathway oversees many embryonic and larval ACDs; here, a Wnt gradient induces an asymmetric distribution of Wnt signaling components within the dividing mother cell. One terminal nuclear effector of the WβA pathway is the transcriptional activator SYS-1/β-catenin. SYS-1 is sequentially negatively regulated during ACD; first by centrosomal regulation and subsequent proteasomal degradation and second by asymmetric activity of the β-catenin “destruction complex” in one of the two daughter cells, which decreases SYS-1 levels in the absence of WβA signaling. However, the extent to which mother cell SYS-1 influences cell fate decisions of the daughters is unknown. Here, we quantify inherited SYS-1 in the differentiating daughter cells and the role of SYS-1 inheritance in Wnt-directed ACD. Photobleaching experiments demonstrate the GFP::SYS-1 present in daughter cell nuclei is comprised of inherited andde novotranslated SYS-1 pools. We used a photoconvertible DENDRA2::SYS-1, to directly observe the dynamics of inherited SYS-1. Photoconversion during mitosis reveals that SYS-1 clearance at the centrosome preferentially degrades older SYS-1, and this accumulation is regulated via dynein trafficking. Photoconversion of the EMS cell during Wnt-driven ACD shows daughter cell inheritance of mother cell SYS-1. Additionally, loss of centrosomal SYS-1 increased inherited SYS-1 and, surprisingly, loss of centrosomal SYS-1 also resulted in increased levels ofde novoSYS-1 in both EMS daughter cells. Lastly, we show that daughter cell negative regulation of SYS-1 via the destruction complex member APR-1/APC is key to limit both thede novoand the inherited SYS-1 pools in both the E and the MS cells. We conclude that regulation of both inherited and newly translated SYS-1 via centrosomal processing in the mother cell and daughter cell regulation via Wnt signaling are critical to maintain sister SYS-1 asymmetry during ACD.
Sclerotome is compartmentalized by parallel Shh and Bmp signaling downstream of CaMKII
Authors: Rothschild Sarah C. , Row Richard H. , Martin Benjamin L. , Clements Wilson K. 
Published: July 2023

AbstractThe sclerotome in vertebrates comprises an embryonic population of cellular progenitors that give rise to diverse adult tissues including the axial skeleton, ribs, intervertebral discs, connective tissue, and vascular smooth muscle. In the thorax, this cell population arises in the ventromedial region of each of the segmented tissue blocks known as somites. How and when sclerotome adult tissue fates are specified and how the gene signatures that predate those fates are regulated has not been well studied. We have identified a previously unknown role for Ca2+/calmodulin-dependent protein kinase II (CaMKII) in regulating sclerotome patterning in zebrafish. Mechanistically, CaMKII regulates the activity of parallel signaling inputs that pattern sclerotome gene expression. In one downstream arm, CaMKII regulates distribution of the established sclerotome-inductive morphogen sonic hedgehog (Shh), and thus Shh-dependent sclerotome genes. In the second downstream arm, we show a previously unappreciated inductive requirement for Bmp signaling, where CaMKII activates expression ofbmp4and consequently Bmp activity. Bmp activates expression of a second subset of stereotypical sclerotome genes, while simultaneously repressing Shh-dependent markers. Our work demonstrates that CaMKII promotes parallel Bmp and Shh signaling as a mechanism to first promote global sclerotome specification, and that these pathways subsequently regionally activate and refine discrete compartmental genetic programs. Our work establishes how the earliest unique gene signatures that likely drive distinct cell behaviors and adult fates arise within the sclerotome.
Reductive amination cascades in cell-free and resting whole cell formats for valorization of lignin deconstruction products
Authors: Nain Priyanka , Dickey Roman M. , Somasundaram Vishal , Sulzbach Morgan , Kunjapur Aditya M. 
Published: July 2023

AbstractThe selective introduction of amine groups within deconstruction products of lignin could provide an avenue for valorizing waste biomass while achieving a green synthesis of industrially relevant building blocks from sustainable sources. Here, we built and characterized enzyme cascades that create aldehydes and subsequently primary amines from diverse lignin-derived carboxylic acids using a carboxylic acid reductase (CAR) and an ω-transaminase (TA). Unlike previous studies that have paired CAR and TA enzymes, here we examine multiple homologs of each of these enzymes and a broader set of candidate substrates. In addition, we compare the performance of these systems in cell-free and resting whole-cell biocatalysis formats using the conversion of vanillate to vanillyl amine as model chemistry. We also demonstrate that resting whole cells can be recycled for multiple batch reactions. We used the knowledge gained from this study to produce several amines from carboxylic acid precursors using one-pot biocatalytic reactions, several of which we report for the first time. These results expand our knowledge of these industrially relevant enzyme families to new substrates and contexts for environmentally friendly and potentially low-cost synthesis of diverse aryl aldehydes and amines.
Validation of GCN5L1/BLOC1S1/BLOS1 Antibodies Using Knockout Cells and Tissue
Authors: Bugga Paramesha , Stoner Michael W. , Manning Janet R. , Mushala Bellina A.S. , Thapa Dharendra , Scott Iain 
Published: July 2023

AbstractGCN5L1, also known as BLOC1S1 and BLOS1, is a small intracellular protein involved in a number of key biological processes. Over the last decade, GCN5L1 has been implicated in the regulation of protein lysine acetylation, energy metabolism, endo-lysosomal function, and cellular immune pathways. An increasing number of published papers have used commercially-available reagents to interrogate GCN5L1 function. However, in many cases these reagents have not been rigorously validated, leading to potentially misleading results. In this report we tested several commercially-available antibodies for GCN5L1, and found that two-thirds of those available did not unambiguously detect the protein by western blot in cultured mouse cells orex vivoliver tissue. These data suggest that previously published studies which used these unverified antibodies to measure GCN5L1 protein abundance, in the absence of other independent methods of corroboration, should be interpreted with appropriate caution.