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King, C.

Publications and source records attributed to King, C..

At least 19 recordsLinked to original sources

Lateral recurrence dynamically refines visual representations for task performance

Recurrent connections are pervasive in visual cortex, and visual representations evolve as recurrent processing unfolds. Yet the mechanism underlying this temporal transformation remains poorly understood. To address this question, we trained recurrent convolutional networks on face and object recognition and tested them on a parametric stimulus set in which category, identity, and pose were systematically varied. This design isolates how lateral recurrence reshapes neural-population representational geometry and single-unit tuning across recurrent timesteps. Within the trained domain, recurrence increased the separability between identities in deeper layers, while leaving the non-trained domain representations largely unchanged. Gating lateral weights abolished the across-timestep gain in trained-domain identity decoding, demonstrating that lateral recurrence drives this refinement. At the single-unit level, recurrence strengthened the correlation between each unit's category preference and its within-domain identity sensitivity, providing a unit-level mechanism for the population-level changes. These results identify lateral recurrence as a dynamic mechanism for refining learned representational structure within a fixed neural population, yielding testable predictions for time-resolved recordings in domain-selective cortex.

neuroscience↗

Genomic SEM reveals shared genetic architecture and expands locus discovery for locomotor activity in heterogeneous stock rats

Locomotor response to a novel environment is a heritable, addiction-relevant behavioral phenotype in rodents. We characterized its genetic basis in 9,341 male and female heterogeneous stock (HS) rats from eight studies. Locomotor activity was heritable across studies, while pairwise genetic correlations were positive but variable, indicating shared yet nonidentical genetic influences. Given nonuniform genetic architecture, we compared three complementary approaches to locus discovery that analyzed SNP effects 1) on a pooled locomotor phenotype (MEGA), 2) across studies under a fixed-effect model (METAL), or 3) on a shared genetic factor (Genomic SEM). To ensure comparability, all three locus-discovery approaches were applied to a common subset of seven studies. Together, these approaches identified 38 loci, with both convergence and method-specific discovery across analyses. Genomic SEM identified 31 loci, compared to 8 for MEGA and 13 for METAL. To interpret the loci, we integrated brain molecular QTLs, predicted coding and splice consequences, and open-field locomotor phenotypes from knockouts of orthologous mouse genes. Our results highlighted established neurobehavioral candidates spanning locomotor and reward-related biology (Negr1 and Lmo4), serotonergic signaling (Htr1b), and GABAergic signaling (Gabrg2), as well as relatively novel genes including Tcerg1l, Pcdh10, and Tmem132e. Our results illustrate how Genomic SEM can integrate genetically correlated yet heterogeneous behavioral measures and enable discovery of loci underlying their shared genetic architecture, highlighting its utility for complex-trait studies in rodents.

genetics↗

Recombination and repetitive genomic landscapes are decoupled in a close relative of Caenorhabditis elegans

Genomes exhibit chromosomal heterogeneity. Distributions of genes, repetitive elements, and polymorphisms are not uniform along chromosomes in multiple species. One explanation for these patterns is recombination rate variation. As recombination interacts with selection to shape the evolutionary fates of alleles, recombination rate variation could promote differences in the chromosomal distribution of genomic features. Thus, clarifying the relationship between recombination rates and genomic organization is a major goal of genetics. In the nematode Caenorhabditis elegans, recombination rate correlates with multiple genomic features that are non-uniformly distributed along chromosomes; recombination rates and repeats are higher on chromosome ends compared to chromosome centers. Its closest known relative, C. inopinata, harbors a radically altered genome with nearly uniform chromosomal distributions of repetitive elements. Is this dramatic change in genomic organization connected to the evolution of recombination rates? Here, we describe a genetic map of C. inopinata constructed via whole-genome sequencing of 180 individual F2 recombinants. This reveals four chromosomes have a conserved recombination rate domain structure whereas two other chromosomes harbor divergent, more uniform recombination rate distributions. Comparisons of these intrachromosomal recombination rates with genomic features reveal little covariation between recombination rate, diversity, gene density, and repeat content in C. inopinata (in stark contrast to most Caenorhabditis species). This suggests that the evolution of recombination may not be entirely responsible for the atypical uniform distribution of repetitive elements across C. inopinata chromosomes. Taken together, these observations reveal that recombination rates can be decoupled from the genomic organization of repetitive elements.

genomics↗

Comparative benchmarking of single-cell transcriptomes and immune repertoires across technologies

Immune receptor profiling enables tracking of individual T or B cell clones across time and tissues, providing insight into immune responses, cancer, and autoimmunity. When combined with single-cell transcriptomics, it links clonotype identity to cellular function, revealing the diversity and dynamics of immune cell populations. This study presents a head-to-head benchmarking comparison of two single-cell immune profiling technologies: droplet-based microfluidics from 10x Genomics (10x) and combinatorial barcoding from Parse Biosciences (Parse). Using matched human samples from PBMCs, the analysis evaluates performance across transcriptomic and T cell immune receptor features to assess data quality, reproducibility, and chain-specific recovery. The findings provide a framework for interpreting single-cell immune profiling platforms and emphasize the importance of accounting for technology-specific biases in bioinformatic analyses.

bioinformatics↗

DENDRO: Recovery and denoising of whole-tree dendritic voltagefrom 2D voltage movies

The ability to image voltage at high spatiotemporal resolution across an entire dendritic tree would represent a major advance in systems and circuit neuroscience. Recent advances in genetically encoded voltage indicators (GEVIs) have brought this possibility closer to reality. However, due to fundamental tradeoffs between imaging speed, resolution, SNR, and volume, this goal has remained out of reach. Here we develop a computational method that fuses 3D anatomical information with 2D voltage video data, yielding full time-varying 3D voltage estimates. Our method, termed DENDRO, comprises two steps. In step one, we use the anatomical data to build a microscope model which maps from voltages along the tree to observed fluorescence at the imaging plane. By exploiting local spatial smoothness of the voltage signal, we parameterize the voltage signal using a set of local basis functions, which reduces the dimensionality of the problem and allows us to approximately invert the microscope model. This step leverages spatial but not temporal smoothness of the underlying signal and yields noisy 3D estimates. In step two, we train a lightweight self-supervised neural network to perform spatiotemporal denoising of the inferred voltages. On simulated data, we find that DENDRO is able to recover voltages at high accuracy across an entire dendritic tree. On real voltage movies from hippocampal slices, DENDRO recovers known dendritic phenomena at single trial resolution and millisecond time-scales, and allows visualization of backpropagating action potentials in 3D.

neuroscience↗

An IL-21R hypomorph circumvents functional redundancy to define STAT1 signaling in germinal center responses

Interleukin-21 receptor engagement initiates a Janus Kinase (JAK) - signal transducer and activator of transcription (STAT) signalling cascade that activates several STAT proteins that drive the germinal center response. However, the relative effects of IL-21 on individual STAT proteins during the differentiation of T follicular helper cells and germinal center B cells has been difficult to distinguish. Here, we characterise a novel mutation in the interleukin-21 receptor (IL-21REINS) that creates a unique defect in the activation of STAT1. Our findings provide evidence that IL-21 mediated activation of STAT1 has a nonredundant role in the differentiation of T follicular (Tfh) cells following T dependent immunisation. IL-21REINS Tfh cells were even more impaired than Tfh cells genetically deficient in IL-21R, questioning our current understanding of the role of IL-21 derived from protein knockout mice. The observation that functional compensation fails in the presence of the IL-21R hypomorph provides insight into how underlying compensation can impact our interpretation of a complex biological system.

immunology↗

KlebPhaCol: A community-driven resource for Klebsiella research identified a novel gut phage order associated with the human gut

The growing threat of multidrug-resistant Klebsiella pneumoniae, coupled with its role in gut colonisation, has intensified the search for new treatments, including bacteriophage therapy. Despite increasing documentation of Klebsiella-targeting phages, clinical applications remain limited, with key phage-bacteria interactions still poorly understood. A major obstacle is fragmented access to well-characterised phage-bacteria pairings, restricting the collective advancement of therapeutic and mechanistic insights. To address this gap, we created the Klebsiella Phage Collection (KlebPhaCol), an open-source resource comprising 53 phages and 74 Klebsiella isolates, all fully characterised. These phages span five families - including a novel order, Felixvirales, associated with the human gut - and target 27 sequence types (including ST258, ST11, ST14) and 28 capsular-locus types (including KL1 and KL2), across six Klebsiella species. Freely accessible at www.klebphacol.org, KlebPhaCol invites the scientific community to both use and contribute to this resource, fostering collaborative research and a deeper understanding of Klebsiella-phage interactions beyond therapeutic use.

microbiology↗

Somatic mutant selection is altered by prior NOTCH1 mutation in aging esophageal epithelium

The ageing mid esophagus is progressively colonized by somatic mutations, including oncogenic TP53 and antioncogenic NOTCH1 mutants. Most esophageal squamous carcinomas are TP53 mutant, NOTCH1 wild type. By middle age, much of the esophagus is NOTCH1 mutant. We hypothesised that prior NOTCH1 mutation may alter subsequent mutant selection. Here we show that selection of mutant NOTCH2 was increased and mutant TP53 decreased within NOTCH1 mutant epithelium. Potential mechanisms include spatial competition between adjacent clones and/or epistasis altering the fitness NOTCH1 double mutants. We investigated these possibilities in transgenic mice. Spontaneous Notch2 mutant clones emerged in aged Notch1 null, but not wild type, esophagus, consistent with epistasis increasing Notch1/Notch2 double mutant fitness. However, there was no evidence of epistasis between Notch1 and Trp53, as Notch1 mutation alone increased fitness. In consequence, over time, Trp53 mutant Notch1 wild type clones were outcompeted by expanding Notch1 and Trp53/Notch1 mutants. NOTCH1 mutant takeover of the aging esophagus alters the trajectory of somatic evolution, decreasing selection of TP53 mutants and potentially reducing cancer risk.

genomics↗

Multiple Unfolded Protein Response pathways cooperate to link cytosolic dsDNA release to Stimulator of Interferon Gene (STING) activation

The double-stranded DNA (dsDNA) sensor STING has been increasingly implicated in responses to "sterile" endogenous threats and pathogens without nominal DNA or cyclic di-nucleotide stimuli. Previous work showed an endoplasmic reticulum (ER) stress response, known as the unfolded protein response (UPR), activates STING. Herein, we sought to determine if ER stress generated a STING ligand, and to identify the UPR pathways involved. Induction of IFN-{beta} expression following stimulation with the UPR inducer thapsigargin (TPG) or oxygen glucose deprivation required both STING and the dsDNA-sensing cyclic GMP-AMP synthase (cGAS). Furthermore, TPG increased cytosolic mitochondrial DNA, and immunofluorescence visualized dsDNA punctae in murine and human cells, providing a cGAS stimulus. N-acetylcysteine decreased IFN-{beta} induction by TPG, implicating reactive oxygen species (ROS). However, mitoTEMPO, a mitochondrial oxidative stress inhibitor did not impact TPG-induced IFN. On the other hand, inhibiting the inositol requiring enzyme 1 (IRE1) ER stress sensor and its target transcription factor XBP1 decreased the generation of cytosolic dsDNA. iNOS upregulation was XBP1-dependent, and an iNOS inhibitor decreased cytosolic dsDNA and IFN-{beta}, implicating ROS downstream of the IRE1-XBP1 pathway. Inhibition of the PKR-like ER kinase (PERK) pathway also attenuated cytoplasmic dsDNA release. The PERK-regulated apoptotic factor Bim was required for both dsDNA release and IFN-{beta} mRNA induction. Finally, XBP1 and PERK pathways contributed to cytosolic dsDNA release and IFN-induction by the RNA virus, Vesicular Stomatitis Virus (VSV). Together, our findings suggest that ER stressors, including viral pathogens without nominal STING or cGAS ligands such as RNA viruses, trigger multiple canonical UPR pathways that cooperate to activate STING and downstream IFN-{beta} via mitochondrial dsDNA release.

immunology↗

A de novo variant in PAK2 detected in an individual with Knobloch type 2 syndrome

P21-activated kinase 2 (PAK2) is a serine/threonine kinase essential for a variety of cellular processes including signal transduction, cellular survival, proliferation, and migration. A recent report proposed monoallelic PAK2 variants cause Knobloch syndrome type 2 (KNO2)--a developmental disorder primarily characterized by ocular anomalies. Here, we identified a novel de novo heterozygous missense variant in PAK2, NM_002577.4:c.1273G>A, p.(D425N), by whole genome sequencing in an individual with features consistent with KNO2. Notable clinical phenotypes include global developmental delay, congenital retinal detachment, mild cerebral ventriculomegaly, hypotonia, FTT, pyloric stenosis, feeding intolerance, patent ductus arteriosus, and mild facial dysmorphism. The p.(D425N) variant lies within the protein kinase domain and is predicted to be functionally damaging by in silico analysis. Previous clinical genetic testing did not report this variant due to unknown relevance of PAK2 variants at the time of testing, highlighting the importance of reanalysis. Our findings also substantiate the candidacy of PAK2 variants in KNO2 and expand the KNO2 clinical spectrum.

genomics↗

Radiation of pain: Psychophysical evidence of population coding mechanism

The spread of pain across body locations remains poorly understood but may provide important insights into the encoding of sensory features of noxious stimuli by populations of neurons. In this psychophysical experiment, we hypothesized that more intense noxious stimuli would lead to spread of pain, but more intense light stimuli would not produce perceptual radiation. Fifty healthy volunteers (27 females, 23 males, ages 14-44) participated in this study wherein noxious stimuli (43, 45, 47 and 49{degrees}C) were applied to glabrous (hand) and hairy skin (forearm) skin with 5s and 10s durations. Also, visual stimuli displayed on the target bodily area were utilized as a control. Participants provided pain (and light) spatial extent ratings as well as pain (and light) intensity ratings. In the extent rating procedure, participants adjusted the extent of the square displayed on the screen with the extent of pain (or light) which they experienced. Pain extent ratings showed statistically significant radiation of pain indicated by 12.42x greater spatial spread of pain (pain extent) than the area of the stimulation with 49{degrees}C (p<0.001), in contrast to visual ratings which closely approximated the size of the stimulus (1.22x). Pain radiation was more pronounced in hairy than glabrous skin (p<0.05) and was more pronounced with longer stimulus duration (p<0.001). Pain intensity explained only 14% of the pain radiation variability. The relative independence of the pain radiation from pain intensity indicates that distinct components of population coding mechanisms may be involved in the spatial representation of pain versus intensity coding.

neuroscience↗

ASPYRE-Lung: Validation of a simple, fast, robust and novel method for multi-variant genomic analysis of actionable NSCLC variants in tissue

Genomic variant testing of tumors is a critical gateway for patients to access the full potential of personalized oncology therapeutics. Current methods such as next-generation sequencing are costly and challenging to interpret, while PCR assays are limited in the number of variants they can cover. We developed ASPYRE(R) (Allele-Specific PYrophosphorolysis REaction) technology to address the urgent need for rapid, accessible and affordable diagnostics informing actionable genomic target variants of a given cancer. The targeted ASPYRE-Lung panel for non-small cell carcinoma covers 114 variants in 11 genes (ALK, BRAF, EGFR, ERBB2, KRAS, RET, ROS1, MET & NTRK1/2/3) to robustly inform clinical management. The assay detects single nucleotide variants, insertions, deletions, and gene fusions from tissue-derived DNA and RNA simultaneously. We tested the limit of detection, specificity, analytical accuracy and analytical precision of ASPYRE-Lung using FFPE lung tissue samples from patients with non-small cell lung carcinoma, variant-negative FFPE tissue from healthy donors, and FFPE-based contrived samples with controllable variant allele fractions. The sensitivity of ASPYRE-Lung was determined to be [&le;] 3% variant allele fraction for single nucleotide variants and insertions or deletions, 100 copies for fusions, and 200 copies for MET exon 14 skipping. The specificity was 100% with no false positive results. The analytical accuracy test yielded no discordant calls between ASPYRE-Lung and expected results for clinical samples (via orthogonal testing) or contrived samples, and results were replicable across operators, reagent lots, runs, and real-time PCR instruments with a high degree of precision. The technology is simple and fast, requiring only four reagent transfer steps using standard laboratory equipment (PCR and qPCR instruments) with analysis via a cloud-based analysis algorithm. The ASPYRE-Lung assay has the potential to be transformative in facilitating access to rapid, actionable molecular profiling of tissue for patients with non-small cell carcinoma.

cancer biology↗

Reduction of retinal ganglion cell death in mouse models of familial dysautonomia using AAV-mediated gene therapy and splicing modulators

Familial dysautonomia (FD) is a rare neurodevelopmental and neurodegenerative disease caused by a splicing mutation in the Elongator Acetyltransferase Complex Subunit 1 (ELP1) gene. The reduction in ELP1 mRNA and protein leads to the death of retinal ganglion cells (RGCs) and visual impairment in all FD patients. Currently, patient symptoms are managed, but there is no treatment for the disease. We sought to test the hypothesis that restoring levels of Elp1 would thwart the death of RGCs in FD. To this end, we tested the effectiveness of two therapeutic strategies for rescuing RGCs. Here we provide proof-of-concept data that gene replacement therapy and small molecule splicing modifiers effectively reduce the death of RGCs in mouse models for FD and provide pre-clinical data foundation for translation to FD patients.

neuroscience↗

ANXA11 biomolecular condensates facilitate protein-lipid phase coupling on lysosomal membranes

Phase transitions of cellular proteins and lipids play a key role in governing the organisation and coordination of intracellular biology. The frequent juxtaposition of proteinaceous biomolecular condensates to cellular membranes raises the intriguing prospect that phase transitions in proteins and lipids could be co-regulated. Here we investigate this possibility in the ribonucleoprotein (RNP) granule-ANXA11-lysosome ensemble, where ANXA11 tethers RNP granule condensates to lysosomal membranes to enable their co-trafficking. We show that changes to the protein phase state within this system, driven by the low complexity ANXA11 N-terminus, induce a coupled phase state change in the lipids of the underlying membrane. We identify the ANXA11 interacting proteins ALG2 and CALC as potent regulators of ANXA11-based phase coupling and demonstrate their influence on the nanomechanical properties of the ANXA11-lysosome ensemble and its capacity to engage RNP granules. The phenomenon of protein-lipid phase coupling we observe within this system offers an important template to understand the numerous other examples across the cell whereby biomolecular condensates closely juxtapose cell membranes. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=177 SRC="FIGDIR/small/533832v1_ufig1.gif" ALT="Figure 1"> View larger version (76K): org.highwire.dtl.DTLVardef@16f3e10org.highwire.dtl.DTLVardef@538f39org.highwire.dtl.DTLVardef@1e00d4eorg.highwire.dtl.DTLVardef@89967d_HPS_FORMAT_FIGEXP M_FIG C_FIG

cell biology↗

Epithelioids: Self-sustaining 3D epithelial cultures to study long-term processes

Studying long-term biological processes such as the colonization of aging epithelia by somatic mutant clones has been slowed by the lack of suitable culture systems. Here we describe epithelioids, a facile, cost-effective method of culturing multiple mouse and human epithelia. Esophageal epithelioids self-maintain without passaging for at least a year, recapitulating the 3D structure, cell dynamics, transcriptome, and genomic stability of the esophagus. Live imaging over 5 months showed epithelioids replicate in vivo cell dynamics. Epithelioids enable the study of cell competition and mutant selection in 3D epithelia, and how anti-cancer treatments modulate the competition between transformed and wild type cells. Epithelioids are a novel method with a wide range of applications in epithelial tissues, particularly the study of long term processes, that cannot be accessed using other culture models.

cell biology↗

A dynamic epibiont community associated with the bone eating worm Osedax

Osedax, the deep-sea annelid found at sunken whalefalls, is known to host Oceanospirillales bacterial endosymbionts intracellularly in specialized roots, that help it feed exclusively on vertebrate bones. Past studies, however, have also made mention of external bacteria on their trunks. During a 14-year study, we reveal a dynamic, yet persistent, succession of Campylobacterales integrated into the epidermis of Osedax, that change over time as the whale carcass degrades on the sea floor. The Campylobacterales associated with seven species of Osedax, which comprise 67% of the bacterial community on the trunk, are initially dominated by the genus Arcobacter (at early time points < 24 months), the Sulfurospirillum at intermediate stages (~ 50 months), and the Sulfurimonas at later stages (>140 months) of whale carcass decomposition. Metagenome analysis of the epibiont metabolic capabilities suggests a transition from heterotrophy to autotrophy along the successional gradient, and differences in their capacity to metabolize oxygen, carbon, nitrogen, and sulfur. Compared to free living relatives, the Osedax epibionts were highly enriched in transposable elements, implicating genetic exchange on the host surface, and contained numerous secretions systems with eukaryotic-like protein domains, suggesting a long evolutionary history with these enigmatic, yet widely distributed deep-sea worms

microbiology↗

Spatial tuning in nociceptive processing is driven by attention

When the source of nociception expands across a body area, the experience of pain increases due to the spatial integration of nociceptive information. This well-established effect is called spatial summation of pain (SSp) and has been the subject of multiple investigations. Here, we used cold-induced SSp to investigate the effect of attention on the spatial tuning of nociceptive processing. Forty pain-free volunteers (N=40, 20 females) participated in this experiment. They took part in an SSp paradigm based on three hand immersions into cold water (5{degrees}C): Participants either immersed the ulnar segment ("a"), radial segment ("b") or both hand segments ("a+b") and provided overall pain ratings. In some trials based on "a+b" immersions, they were also asked to provide divided (i.e., first pain in "a" then in "b"; or reversed) and directed attention ratings (i.e., pain only in "a" or "b"). Results confirmed a clear SSp effect in which reported pain during immersions of "a" or "b" was less intense than pain during immersions of "a+b" (p<0.001). Data also confirmed that spatial tuning was altered. SSp was fully abolished when participants provided two ratings in a divided fashion (p<0.001). Furthermore, pain was significantly lower when attention was directed only to one segment ("a" OR "b") during "a+b" immersion (p<0.001). We conclude that spatial tuning is dynamically driven by attention as reflected in abolished SSp. Directed attention was sufficient to focus spatial tuning and abolish SSp. Results support the role of cognitive processes such as attention in spatial tuning. PerspectiveThis article presents experimental investigation of spatial tuning in pain and offers mechanistic insights of contiguous spatial summation of pain in healthy volunteers. Depending on how pain is evaluated in terms of attentional derivative (overall pain, directed, divided attention) the pain is reduced and spatial summation abolished.

neuroscience↗

A natural variation-based screen in mouse cells reveals USF2 as a regulator of the DNA damage response and cellular senescence

Cellular senescence is a program of cell cycle arrest, apoptosis resistance, and cytokine release induced by stress exposure in metazoan cells. Landmark studies in laboratory mice have characterized a number of master senescence regulators, including p16INK4a, p21, NF-kB, p53, and C/EBP{beta}. To discover other molecular players in senescence, we developed a screening approach to harness the evolutionary divergence between mouse species. We found that primary cells from the Mediterranean mouse Mus spretus, when treated with DNA damage to induce senescence, produced less cytokine and had less-active lysosomes than cells from laboratory M. musculus. We used allele-specific expression profiling to catalog senescence-dependent cis-regulatory variation between the species at thousands of genes. We then tested for correlation between these expression changes and interspecies sequence variants in the binding sites of transcription factors. Among the emergent candidate senescence regulators, we chose a little-studied cell cycle factor, USF2, for molecular validation. In acute irradiation experiments, cells lacking USF2 had compromised DNA damage repair and response. Longer-term senescent cultures without USF2 mounted an exaggerated senescence regulatory program--shutting down cell cycle and DNA repair pathways, and turning up cytokine expression, more avidly than wild-type. We interpret these findings under a model of pro-repair, anti-senescence regulatory function by USF2. Our study affords new insights into the mechanisms by which cells commit to senescence, and serves as a validated proof of concept for natural variation-based regulator screens.

genomics↗