Biology subjects

Smith, S. E.

Publications and source records attributed to Smith, S. E..

14 recordsLinked to original sources

bioRxiv2026

Simulated field potentials capture multiple simultaneous processes underlying human temporal attention

Essential cognitive processes like attention, memory, and decision-making require the brain to perform many operations simultaneously. Disentangling the neural mechanisms underlying these processes is challenging because they often unfold concurrently with similar dynamics. In auditory temporal expectation, these mechanisms include the accumulation of incoming sensory information in the auditory cortex interacting with cortical signals biasing activity in anticipation of future stimuli. Here, we leverage invasive human field potential recordings from participants performing a subjective temporal expectation task. Rather than relying on traditional neuroimaging methods that average or transform neural responses, we instead simulate each trial's predicted field potential, based on known field potential physiology combined with components of established cognitive theories of the temporal expectation task: sensory input, top-down bias, and sensory anticipation. Our predicted field potentials strongly correlate with neural activity in auditory regions and in regions associated with auditory temporal perception, including the parietal, temporal, and prefrontal cortices. By differentiating individual cognitive mechanisms through simulation, we identify neural signatures of attentional bias and anticipatory sensory activity across multiple cortical regions that predict trial-by-trial perceptual accuracy and confidence.

neuroscience↗

bioRxiv2024

Metagenomic analyses of single phages and phage cocktails show instances of contamination with temperate phages and bacterial DNA

Increasing antibiotic resistance has led to renewed attention being paid to bacteriophage therapy. Commercial phage cocktails are available but often their contents of the phages are not well defined. Some metagenomic studies have been done to retrospectively characterise these cocktails, but little is known about the replication cycle of the included phages, or about the possible bacterial DNA contamination. In this study, published metagenomic sequences were reanalysed using recent advances in viromics tools. Signs of temperate phage contigs were found in all cocktail metagenomes, as well as host DNA, which could poses a risk as it may lead to horizontal gene transfer of virulence factors to commensals and pathogens. This suggests the need to implement further quality measures before using phage cocktails therapeutically.

microbiology↗

bioRxiv2024

Enhanced pollen tube performance at high temperature contributes to thermotolerant fruit production in tomato.

Rising temperature extremes during critical reproductive periods threaten the yield of major grain and fruit crops. Flowering plant reproduction depends on development of sufficient numbers of pollen grains and on their ability to generate a cellular extension, the pollen tube, which elongates through the pistil to deliver sperm cells to female gametes for double fertilization. These critical phases of the life cycle are sensitive to temperature and limit productivity under high temperature (HT). Previous studies have investigated the effects of HT on pollen development, but little is known about how HT applied during the pollen tube growth phase affects fertility. Here, we used tomato as a model fruit crop to determine how HT affects the pollen tube growth phase, taking advantage of cultivars noted for fruit production in exceptionally hot growing seasons. We found that exposure to HT solely during the pollen tube growth phase limits fruit biomass and seed set more significantly in thermosensitive cultivars than in thermotolerant cultivars. Importantly, we found that pollen tubes from the thermotolerant Tamaulipas cultivar have enhanced growth in vivo and in vitro under HT. Analysis of the pollen tube transcriptomes response to HT allowed us to develop hypotheses for the molecular basis of cellular thermotolerance in the pollen tube and we define two response modes (enhanced induction of stress responses, and higher basal levels of growth pathways repressed by heat stress) associated with reproductive thermotolerance. Importantly, we define key components of the pollen tube stress response identifying enhanced ROS homeostasis and pollen tube callose synthesis and deposition as important components of reproductive thermotolerance in Tamaulipas. Our work identifies the pollen tube growth phase as a viable target to enhance reproductive thermotolerance and delineates key pathways that are altered in crop varieties capable of fruiting under HT conditions. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=95 SRC="FIGDIR/small/606234v1_ufig1.gif" ALT="Figure 1"> View larger version (35K): org.highwire.dtl.DTLVardef@25e5acorg.highwire.dtl.DTLVardef@98fb1org.highwire.dtl.DTLVardef@1b8b9cborg.highwire.dtl.DTLVardef@adcca4_HPS_FORMAT_FIGEXP M_FIG C_FIG

plant biology↗

bioRxiv2023

Defining a core configuration for human centromeres during mitosis

The biorientation of sister chromatids on the mitotic spindle, essential for accurate sister chromatid segregation, relies on critical centromere components including cohesin, the centromere-specific H3 variant CENP-A, and centromeric DNA. Centromeric DNA is highly variable between chromosomes yet must accomplish a similar function. Moreover, how the 50 nm cohesin ring, proposed to encircle sister chromatids, accommodates inter-sister centromeric distances of hundreds of nanometers on the metaphase spindle is a conundrum. Insight into the 3D organization of centromere components would help resolve how centromeres function on the mitotic spindle. We used ChIP-seq and super-resolution microscopy to examine the geometry of essential centromeric components on human chromosomes. ChIP-seq demonstrates that cohesin subunits are depleted in -satellite arrays where CENP-A nucleosomes and kinetochores assemble. Cohesin is instead enriched at pericentromeric DNA. Structured illumination microscopy of sister centromeres is consistent, revealing a non-overlapping pattern of CENP-A and cohesin. We used single particle averaging of hundreds of mitotic sister chromatids to develop an average centromere model. CENP-A clusters on sister chromatids, connected by -satellite, are separated by [~]562 nm with a perpendicular intervening [~]190 nM wide axis of cohesin. Two differently sized -satellite arrays on chromosome 7 display similar inter-sister CENP-A cluster distance, demonstrating different sized arrays can achieve a common spacing. Our data suggest a working model for a common core configuration of essential centromeric components that includes CENP-A nucleosomes at the outer edge of extensible -satellite DNA and pericentromeric cohesion. This configuration helps reconcile how centromeres function and serves as a foundation for future studies of additional components required for centromere function.

cell biology↗

bioRxiv2023

Morphological changes and two Nodal paralogs drive left-right asymmetry in the squamate veiled chameleon (C. calyptratus).

The ancestral mode of left-right (L-R) patterning involves cilia in the L-R organizer. However, the mechanisms regulating L-R patterning in non-avian reptiles remains an enigma, since most squamate embryos are undergoing organogenesis at oviposition. In contrast, veiled chameleon (Chamaeleo calyptratus) embryos are pre-gastrula at oviposition, making them an excellent organism for studying L-R patterning evolution. Here we show that veiled chameleon embryos lack motile cilia in their L-R organizer, consistent with the loss of motile cilia being a synapomorphy of all reptiles. Furthermore, in contrast to avians, geckos and turtles, which have one Nodal gene, veiled chameleon exhibits expression of two paralogs of Nodal in the left lateral plate mesoderm, albeit in non-identical patterns. Using live imaging, we observed asymmetric morphological changes that precede, and likely trigger, asymmetric expression of the Nodal cascade. Thus, veiled chameleons are a new and unique model for studying the evolution of L-R patterning.

developmental biology↗

bioRxiv2022

State-dependent protein-protein interactions mediating 4-1BB CAR Signaling

Cells rely on activity-dependent protein-protein interactions to convey biological signals, but the state-dependent interactome is notoriously cell-specific and undercharacterized1. In the case of chimeric antigen receptor (CAR) T cells containing a 4-1BB costimulatory domain, receptor engagement is thought to trigger the formation of protein complexes similar to those triggered by T cell receptor (TCR)-mediated signaling, but the number and type of protein-interaction-mediating binding domains differ between CARs and TCRs. Here, we performed co-immunoprecipitation mass spectrometry of a 2nd generation CD19-directed 4-1BB:zeta CAR (referred to as bb{zeta}CAR) and identified 67 proteins that increased their co-association after target engagement. We compared activity-induced TCR and CAR signalosomes using quantitative multiplex co-immunoprecipitation and showed that bb{zeta}CAR engagement leads to activation of two modules of protein interactions, one similar to TCR signaling that is more weakly engaged in bb{zeta}CAR vs. TCR, and one composed of TRAF signaling complexes that is not engaged by the TCR. Batch-to-batch and inter-individual variations in IL2 production correlated with differences in the magnitude of protein network activation. Future CAR T cell manufacturing protocols could measure, and eventually control, biological variation by monitoring these signalosome activation markers. One Sentence SummaryWe define a network of protein interactions engaged by chimeric antigen receptors following target binding, and show that the magnitude of network activation correlates with IL-2 secretion, a proxy measure for CAR T cell function.

immunology↗

bioRxiv2021

The SEEL Motif and Members of the MYB-related REVEILLE Transcription Factor Family are Important for the Expression of LORELEI in the Synergid Cells of the Arabidopsis Female Gametophyte

Synergid cells in the micropylar end of the female gametophyte are required for critical cell-cell signaling interactions between the pollen tube and the ovule that precede double fertilization and seed formation in flowering plants. LORELEI (LRE) encodes a GPI-anchored protein that is expressed primarily in the synergid cells, and together with FERONIA, a receptor-like kinase, it controls pollen tube reception by the receptive synergid cell. Still, how LRE expression is controlled in synergid cells remains poorly characterized. We identified candidate cis-regulatory elements enriched in LRE and other synergid cell-expressed genes. One of the candidate motifs ( TAATATCT) in the LRE promoter was an uncharacterized variant of the Evening Element motif that we named as the Short Evening Element-like (SEEL) motif. Deletion or point mutations in the SEEL motif of the LRE promoter resulted in decreased reporter expression in synergid cells, demonstrating that the SEEL motif is important for expression of LRE in synergid cells. Additionally, we found that LRE expression is decreased in the loss of function mutants of REVEILLE (RVE) transcription factors, which are clock genes known to bind the SEEL and other closely related motifs. We propose that RVE transcription factors regulate LRE expression in synergid cells by binding to the SEEL motif in the LRE promoter. Identification of a cis-regulatory element and transcription factors involved in the expression of LRE will serve as a foundation to characterize the gene regulatory networks in synergid cells and investigate the potential connection between circadian rhythm and fertilization. One sentence summaryA newly identified SEEL motif in the promoter of LORELEI and at least three members of the REVEILLE transcription factor family are important for LORELEI expression in synergid cells of the Arabidopsis female gametophyte.

plant biology↗

bioRxiv2021

A distinct inner nuclear membrane proteome in Saccharomyces cerevisiae gametes

The inner nuclear membrane (INM) proteome regulates gene expression, chromatin organization, and nuclear transport, however, it is poorly understood how changes in INM protein composition contribute to developmentally regulated processes, such as gametogenesis. Using a split-GFP complementation system, we compared the distribution of all C-terminally tagged transmembrane proteins in Saccharomyces cerevisiae in gametes to that of mitotic cells. Gametes contain a distinct INM proteome needed to complete gamete formation, including expression of genes linked to cell wall biosynthesis, lipid biosynthetic and metabolic pathways, protein degradation and unknown functions. Based on the inheritance pattern, INM components are made de novo in the gametes. Whereas mitotic cells show a strong preference for proteins with small extraluminal domains, gametes do not exhibit this size preference likely due to the changes in the nuclear permeability barrier during gametogenesis.

cell biology↗

bioRxiv2020

A computational method for immune repertoire mining that identifies novel binders from different clonotypes, demonstrated by identifying anti-Pertussis toxoid antibodies

Due to their shared genetic history, antibodies from the same clonotype often bind to the same epitope. This knowledge is used in immune repertoire mining, where known binders are used to search bulk sequencing repertoires to identify new binders. However current computational methods cannot identify epitope convergence between antibodies from different clonotypes, limiting the sequence diversity of antigen-specific antibodies which can be identified. We describe how the antibody binding site, the paratope, can be used to cluster antibodies with common antigen reactivity from different clonotypes. Our method, paratyping, uses the predicted paratope to identify these novel cross clonotype matches. We experimentally validated our predictions on a Pertussis toxoid dataset. Our results show that even the simplest abstraction of the antibody binding site, using only the length of the loops involved and predicted binding residues, is sufficient to group antigen-specific antibodies and provide additional information to conventional clonotype analysis.

bioinformatics↗

bioRxiv2020

A novel therapeutic antibody screening method using bacterial high-content imaging reveals functional antibody binding phenotypes of Escherichia coli ST131

The increase of antimicrobial resistance (AMR), and lack of new classes of licensed antimicrobials, have made alternative treatment options for AMR pathogens increasingly attractive. Recent studies have demonstrated anti-bacterial efficacy of a humanised monoclonal antibody (mAb) targeting the O25b O-antigen of Escherichia coli ST131. To evaluate the phenotypic effects of antibody binding to diverse clinical E. coli ST131 O25b bacterial isolates in high-throughput, we designed a novel mAb screening method using high-content imaging (HCI) and image-based morphological profiling to screen a mAb targeting the O25b O-antigen. Screening the antibody against a panel of 86 clinical E. coli ST131 O25:H4 isolates revealed 4 binding phenotypes: no binding (18.60%), weak binding (4.65%), strong binding (69.77%) and strong agglutinating binding (6.98%). Impaired antibody binding could be explained by the presence of insertion sequences or mutations in O-antigen or lipopolysaccharide core biosynthesis genes, affecting the amount, structure or chain length of the O-antigen. The agglutinating binding phenotype was linked with lower O-antigen density, enhanced antibody-mediated phagocytosis and increased serum susceptibly. This study highlights the need to screen candidate mAbs against large panels of clinically relevant isolates, and that HCI can be used to evaluate mAb binding affinity and potential functional efficacy against AMR bacteria.

microbiology↗

bioRxiv2020

JNK signaling controls branching, nucleokinesis, and positioning of centrosomes and primary cilia in migrating cortical interneurons

Aberrant migration of inhibitory interneurons can alter the formation of cortical circuitry and lead to severe neurological disorders including epilepsy, autism, and schizophrenia. However, mechanisms involved in directing the migration of these cells remain incompletely understood. In the current study, we used live-cell confocal microscopy to explore the mechanisms by which the c-Jun NH2-terminal kinase (JNK) pathway coordinates leading process branching and nucleokinesis, two cell biological processes that are essential for the guided migration of cortical interneurons. Pharmacological inhibition of JNK signaling disrupts the kinetics of leading process branching, rate and amplitude of nucleokinesis, and leads to the rearward mislocalization of the centrosome and primary cilium to the trailing process. Genetic loss of Jnk from interneurons corroborates our pharmacological observations and suggests that important mechanics of interneuron migration depend on the intrinsic activity of JNK. These findings suggest that JNK signaling regulates leading process branching, nucleokinesis, and the trafficking of centrosomes and cilia during interneuron migration, and further implicates JNK signaling as an important mediator of cortical development. Summary StatementLoss of JNK signaling reduces growth cone branching frequency, limits interstitial side branch duration, alters rate and amplitude of nucleokinesis, and mislocalizes centrosomes and primary cilia in migrating cortical interneurons.

neuroscience↗

bioRxiv2020

Repeated evolution of circadian clock dysregulation in cavefish populations

Circadian rhythms are nearly ubiquitous throughout nature, suggesting they are critical for survival in diverse environments. Organisms inhabiting environments with arrhythmic days, such as caves, offer a unique opportunity to study the evolution of circadian rhythms in response to changing ecological pressures. Here we demonstrate that the cave environment has led to the repeated disruption of the biological clock across multiple populations of Mexican cavefish, with the circadian transcriptome showing widespread reductions in rhythmicity and changes to the timing of the activation/repression of genes in the core pacemaker. Then, we investigate the function of two genes with decreased rhythmic expression in cavefish. Mutants of these genes phenocopy reductions in sleep seen in multiple cave populations, suggesting a link between circadian dysregulation and sleep reduction. Altogether, our results reveal that evolution in an arrhythmic environment has resulted in dysregulation to the biological clock across multiple populations by diverse molecular mechanisms.

evolutionary biology↗

bioRxiv2018

Asi1 regulates the distribution of proteins at the inner nuclear membrane in Saccharomyces cerevisiae

Inner nuclear membrane (INM) protein composition regulates nuclear function, affecting processes such as gene expression, chromosome organization, nuclear shape and stability. Mechanisms that drive changes in the INM proteome are poorly understood in part because it is difficult to definitively assay INM composition rigorously and systematically. Using a split-GFP complementation system to detect INM access, we examined the distribution of all C-terminally tagged Saccharomyces cerevisiae membrane proteins in wild-type cells and in mutants affecting protein quality control pathways, such as INM-associated degradation (INMAD), ER-associated degradation (ERAD) and vacuolar proteolysis. Deletion of the E3 ligase Asi1 had the most pronounced effect on the INM compared to mutants in vacuolar or ER-associated degradation pathways, consistent with a role for Asi1 in the INMAD pathway. Our data suggests that Asi1 not only removes mis-targeted proteins at the INM, but it also controls the levels and distribution of native INM components, such as the membrane nucleoporin Pom33. Interestingly, loss of Asi1 does not affect Pom33 protein levels but instead alters Pom33 distribution in the NE through Pom33 ubiquitination, which drives INM redistribution. Taken together, our data demonstrate that the Asi1 E3 ligase has a novel function in INM protein regulation in addition to protein turnover.

cell biology↗

bioRxiv2018

Membrane insertion function for SUN-KASH complex revealed by high resolution analysis of yeast centrosomes

Bipolar spindle formation in yeast requires insertion of centrosomes (known as spindle pole bodies (SPBs)) into fenestrated regions of the nuclear envelope (NE). Using structured-illumination microscopy and bimolecular fluorescence complementation, we map protein distribution at SPB fenestra and interrogate protein-protein interactions with high spatial resolution. We find that the Sad1-UNC-84 (SUN) protein Mps3 forms a ring-like structure around the SPB, similar to toroids seen for components of the SPB insertion network (SPIN). Mps3 and the SPIN component Mps2 (a Klarsicht-ANC-1-Syne-1 domain (KASH)-like protein) form a novel non-canonical linker of nucleoskeleton and cytoskeleton (LINC) complex that is connected in both luminal and extraluminal domains. This hairpin-like LINC complex forms during SPB insertion, suggesting it functions in NE reorganization at the pore membrane. The LINC complex also controls the distribution of a soluble SPIN component Bbp1. Taken together our work shows that Mps3 is a fifth SPIN component and suggests both direct and indirect roles for the LINC complex in NE remodeling.

cell biology↗