Biology subjects

Johnson, D. S.

Publications and source records attributed to Johnson, D. S..

14 recordsLinked to original sources

bioRxiv2026

Scale-dependent feedback between sociality and space use in a long-lived marine predator

Spatial and social behaviours in animals are intertwined, yet the causal direction of this feedback--and how it varies across scales--remains largely unresolved. Using long-term data from false killer whales (Pseudorca crassidens) in the main Hawaiian Islands, we developed and applied a scale-explicit analytical framework to test how social reliance and resource ephemerality govern top-down versus bottom-up processes linking movement and sociality. Movements, association networks, genetic relatedness, and isotopic niches reveal that strong social bonds drive bottom-up emergence of short-term intra-group movements, while ephemeral and likely island-associated prey landscapes impose top-down constraints on inter-group dynamics across scales. These complementary processes generate persistent, fine-scale fidelity within some groups and relatively well-differentiated feeding niches among them. Our findings highlight a general mechanism by which life-history strategies and environmental stochasticity jointly determine the scale and direction of feedback between space use and sociality--shaping population structure and connectivity in mobile social predators.

ecology↗

bioRxiv2025

Discovery and Validation of Alternatives to VSV-G for Pseudotyping of Lentiviral Vectors for In Vivo Delivery of Anti-Tumor Transgenes

Though cell therapy for cancer is now widely used commercially and has been efficacious for tens of thousands of patients, conventional manufacturing methods are expensive and difficult to scale. An alternative approach is to deliver the relevant anti-tumor transgenes to T cells in vivo in the patient. Such "in vivo cell therapy" methods promise to be more scalable, with reduced cost of goods, since the same drug product can be administered to any patient. Typically, conventional cell therapy introduces anti-tumor transgenes into T cells using a lentivector pseudotyped with VSV-G. However, VSV-G is not cell type specific because its molecular target is present on a diversity of human cells, which may result in less than optimal pharmacology in vivo. Natures existing viral diversity presents the opportunity to identify alternative pseudotypes that are more optimal for in vivo cell therapy. In this study, we first performed a large-scale bioinformatic sequence search for G-proteins similar to VSV-G. We identified 166 G-proteins in the sequence search and then tested 9 in vitro for efficiency, specificity, and sensitivity of transgene delivery to various human immune cell phenotypes. We used the results from this screen to select three G-protein candidates for a pilot GFP transgene delivery study in humanized mice, using anti-CD3 antibody fragments for T cell tropism. One candidate G-protein performed significantly better than VSV-G, so we moved that candidate into tumor control studies in a humanized mouse model. This candidate G-protein was able to deliver an anti-tumor transgene to T cells, which subsequently cleared 100% of tumor burden in 100% of mice. We conclude that systematic screens for optimal lentivector designs can be used to identify optimized candidates for in vivo cell therapy.

cancer biology↗

bioRxiv2024

NaV1.7 targeted fluorescence imaging agents for nerve identification during intraoperative procedures

Surgeries and trauma result in traumatic and iatrogenic nerve damage that can result in a debilitating condition that approximately affects 189 million individuals worldwide. The risk of nerve injury during oncologic surgery is increased due to tumors displacing normal nerve location, blood turbidity, and past surgical procedures, which complicate even an experienced surgeons ability to precisely locate vital nerves. Unfortunately, there is a glaring absence of contrast agents to assist surgeons in safeguarding vital nerves. To address this unmet clinical need, we leveraged the abundant expression of the voltage-gated sodium channel 1.7 (NaV1.7) as an intraoperative marker to access peripheral nerves in vivo, and visualized nerves for surgical guidance using a fluorescently-tagged version of a potent NaV1.7-targeted peptide, Tsp1a, derived from a Peruvian tarantula. We characterized the expression of NaV1.7 in sensory and motor peripheral nerves across mouse, primate, and human specimens and demonstrated universal expression. We synthesized and characterized a total of 10 fluorescently labeled Tsp1a-peptide conjugates to delineate nerves. We tested the ability of these peptide-conjugates to specifically accumulate in mouse nerves with a high signal-to-noise ratio in vivo. Using the best-performing candidate, Tsp1a-IR800, we performed thyroidectomies in non-human primates and demonstrated successful demarcation of the recurrent laryngeal and vagus nerves, which are commonly subjected to irreversible damage. The ability of Tsp1a to enhance nerve contrast during surgery provides opportunities to minimize nerve damage and revolutionize standards of care across various surgical specialties.

biochemistry↗

bioRxiv2024

A range expanding ecosystem engineer influences historical and expanded habitats via the same causal pathways

Species are shifting their ranges in response to climate change. There remain many unknowns about relative impacts of range-expanding ecosystem engineers between historical and expanded habitats, however. The mud fiddler crab Minuca pugnax (=Uca pugnax) is shifting its range northward likely due to increased warming in the Gulf of Maine. A burrowing crab, M. pugnax affects ecosystem functioning in salt marshes south of Cape Cod, Massachusetts with unknown effects in expanded marsh habitats over 150km to the north. We therefore studied the M. pugnax range expansion to determine the extent that range expanding ecosystem engineers are influencing ecosystem functioning expanded ranges relative to historical habitats. We installed in 2017 and 2018 a series of crab-inclusion cages at both the UMass Boston Nantucket Field Station (historical range) and the Plum Island Estuary Long Term Ecological Research site (PIE-LTER, expanded range). For each site, year, and block, we measured in the beginning and end of the three-month experiment metrics of sediment strength, primary production, and decomposition. We developed and tested causal models using structural equation modeling (SEM) to determine direct and indirect effects of fiddler crabs on ecosystem functions. Despite site, year, and block variability, local environment influenced burrow density, which directly affected sediment strength and indirectly affected primary production in both ranges. Overall, understanding range-expanding ecosystem engineers in historical ranges was predictive for how they influence expanded habitats, despite inter-site heterogeneity. Therefore, it is critical to study relative impacts of range-expanding ecosystem engineers to understand total impacts of global range shifts.

ecology↗

bioRxiv2021

Stitchr: stitching coding TCR nucleotide sequences from V/J/CDR3 information

The study and manipulation of T cell receptors (TCRs) is central to multiple fields across basic and translational immunology research. Produced by V(D)J recombination, TCRs are often only recorded in the literature and data repositories as a combination of their V and J gene symbols, plus their hypervariable CDR3 amino acid sequence. However, numerous applications require full-length coding nucleotide sequences. Here we present Stitchr, a software tool developed to specifically address this limitation. Given minimal V/J/CDR3 information, Stitchr produces complete coding sequences representing a fully spliced TCR cDNA. Due to its modular design, Stitchr can be used for TCR engineering using either published germline or novel/modified variable and constant region sequences. Sequences produced by Stitchr were validated by synthesizing and transducing TCR sequences into Jurkat cells, recapitulating the expected antigen specificity of the parental TCR. Using a companion script, Thimble, we demonstrate that Stitchr can process a million TCRs in under ten minutes using a standard desktop personal computer. By systemizing the production and modification of TCR sequences, we propose that Stitchr will increase the speed, repeatability, and reproducibility of TCR research. Stitchr is available on GitHub.

immunology↗

bioRxiv2021

Lack of blocking activity in anti-CTLA-4 antibodies reduces toxicity, but not anti-tumor efficacy

Anti-CTLA-4 antibodies such as ipilimumab were among the first immune-oncology agents to show significantly improved outcomes for patients. However, existing anti-CTLA-4 therapies fail to induce a response in a majority of patients and can induce severe, immune-related adverse events. It has been assumed that checkpoint inhibition, i.e., blocking the interaction between CTLA-4 and its ligands, is the primary mechanism of action for ipilimumab. In this study we present evidence that checkpoint inhibition is not a primary mechanism of action for efficacy of anti-CTLA-4 antibodies. Instead, the primary mechanism for efficacy is FcR-mediated Treg depletion in the tumor microenvironment. First, we identified a monoclonal antibody (mAb) that binds to CTLA-4 at an epitope that differs from ipilimumabs by only a few amino acids, yet has limited checkpoint inhibitor activity. Surprisingly, the weak checkpoint inhibitor has superior anti-tumor activity compared to ipilimumab in a murine model. The weak checkpoint inhibitor also induces less Treg proliferation and has increased ability to induce in vitro FcR signaling and in vivo depletion of intratumoral Tregs. Further experiments showed that the enhanced FcR activity of the weak checkpoint inhibitor likely contributes to its enhanced anti-tumor activity. Importantly, we also showed that weak checkpoint inhibition was associated with lower toxicity in murine models. Our work suggests that new anti-CTLA-4 drugs should be optimized for Treg depletion rather than checkpoint inhibition.

immunology↗

bioRxiv2020

Single cell transcriptomics reveals the effect of PD-L1/TGF-β blockade on the tumor microenvironment

BackgroundThe anti-tumor activity of anti-PD-1/PD-L1 therapies correlates with T cell infiltration in tumors. Thus, a major goal in oncology is to find strategies that enhance T cell infiltration and efficacy of anti-PD-1/PD-L1 therapy. TGF-{beta} has been shown to contribute to T cell exclusion and anti-TGF-{beta} improves anti-PD-L1 efficacy in vivo. However, TGF-{beta} inhibition has frequently been shown to induce toxicity in the clinic, and the clinical efficacy of combination PD-L1 and TGF-{beta} blockade has not yet been proven. To identify strategies to overcome resistance to PD-L1 blockade, the transcriptional programs associated with PD-L1 and/or TGF-{beta} blockade in the tumor microenvironment should be further elucidated. ResultsFor the first time, we used single-cell RNA sequencing to characterize the transcriptomic effects of PD-L1 and/or TGF-{beta} blockade on nearly 30,000 single cells in the tumor and surrounding microenvironment. Combination treatment led to upregulation of immune response genes, including multiple chemokine genes such as CCL5, in CD45+ cells, and down-regulation of extracellular matrix genes in CD45-cells. Analysis of publicly available tumor transcriptome profiles showed that the chemokine CCL5 was strongly associated with immune cell infiltration in various human cancers. Further investigation with in vivo models showed that intratumorally administered CCL5 enhanced cytotoxic lymphocytes and the anti-tumor activity of anti-PD-L1. ConclusionsTaken together, our data could be leveraged translationally to improve anti-PD-L1 plus anti-TGF-{beta} combination therapy, for example through companion biomarkers, and/or to identify novel targets that could be modulated to overcome resistance.

genomics↗

bioRxiv2020

Cargo modulates the conformation of the nuclear pore in living cells

While the static structure of the nuclear pore complex (NPC) continues to be refined with cryo-EM and x-ray crystallography, the in vivo conformational dynamics of the NPC remain under-explored. We developed sensors that report on the orientation of NPC components by rigidly conjugating mEGFP to different NPC proteins. Our studies show conformational changes to select domains of Nups within the inner ring (Nup54, Nup58, Nup62) when transport through the NPC is perturbed and no conformational changes to Nups elsewhere in the NPC. Our results suggest that select components of the NPC are flexible and undergo conformational changes upon engaging with cargo.

cell biology↗

bioRxiv2019

Estimating northern fur seal pup production: A case study of batch mark abundance estimation

We describe a hierarchical N-mixture model for estimating northern fur seal pup production from batch mark-resight data. Our goal was to improve upon a traditional design-based estimation method used for over 50 years. To this end, we propose a hierarchical N-mixture model to account for differences in animal availability for resighting and observer detection probabilities. A Bayesian approach is used for inference with three separate methods proposed for necessary computations. First a straightforward posterior sample is drawn using MCMC. This was considered the gold standard for this analysis. However, we also consider an approximate model-based on Gaussian approximation of the Poisson and binomial distributions used in the exact hierarchical model. By using the Gaussian approximations, analytic integration can be used to marginalize over latent components. Inference can then be made by maximizing the posterior to find the mode. Following this we investigate both delta-method and parametric bootstrap approaches for calculating abundance and the associated standard errors with batch sampling data collected on northern fur seals in 2016. Each of the three newly proposed methods produced virtually identical estimates and standard errors. In addition, the parametric bootstrap approximation to a posterior sample was also virtually the same as the MCMC derived sample. There were a handful of small differences in the point estimates of abundance but these were not large enough to be especially notable. There were some noticeable differences in uncertainty estimates. The production estimate coefficients of variation ranged from 2.5-9.5% for the model-based estimates, but 0.0-12.5% for the design-based estimates. The design-based estimates of uncertainty were more volatile using the design-based method with some standard error estimates unreasonably small. Through a case study we have provided support for using Gaussian approximation in N-mixture type models when abundance is relatively large. These situations are typically challenging for fitting models using any sort of marginalization because there is no analytic solution for the exact models. However, when abundances are large using a Gaussian approximated model allows for analytic integration over latent abundance states which can greatly facilitate methodology such as maximum likelihood or posterior inference.

ecology↗

bioRxiv2018

A Continuous-Time Semi-Markov Model for Animal Movement in a Dynamic Environment

We consider an extension to discrete-space continuous-time models animal movement that have previously be presented in the literature. The extension from a continuous-time Markov formulation to a continuous-time semi-Markov formulation allows for the inclusion of temporally dynamic habitat conditions as well as temporally changing movement responses by animals to that environment. We show that with only a little additional consideration, the Poisson likelihood approximation for the Markov version can still be used within the multiple imputation framework commonly employed for analysis of telemetry data. In addition, we consider a Bayesian model selection methodology with the imputation framework. The model selection method uses a Laplace approximation to the posterior model probability to provide a computationally feasible approach. The full methodology is then used to analyze movements of 15 northern fur seal (Callorhinus ursinus) pups with respect to surface winds, geostrophic currents, and sea surface temperature. The highest posterior model probabilities belonged to those models containing only winds and current, SST did not seem to be a significant factor for modeling their movement.

ecology↗

bioRxiv2018

Timing of ESCRT-III protein recruitment and membrane scission during HIV-1 assembly

The Endosomal Sorting Complexes Required for Transport III (ESCRT-III) proteins are critical for cellular membrane scission processes with topologies inverted relative to clathrin-mediated endocytosis. Some viruses appropriate ESCRT-IIIs for their release. By imaging single assembling viral-like particles of HIV-1, we observed that ESCRT-IIIs and the ATPase VPS4 arrive after most of the virion membrane is bent, linger for tens of seconds, and depart [~]20 seconds before scission. These observations suggest ESCRT-IIIs are recruited by a combination of membrane curvature and the late domains of the HIV-1 Gag protein. ESCRT-IIIs may pull the neck into a narrower form but must leave to allow scission. If scission does not occur within minutes of ESCRT departure, ESCRT-III and VPS4 are recruited again. This mechanistic insight is likely relevant for other ESCRT dependent scission processes including cell division, endosome tubulation, multivesicular body and nuclear envelope formation, and secretion of exosomes and ectosomes.

cell biology↗

bioRxiv2018

The contribution of non-canonical splicing mutations to severe dominant developmental disorders

Mutations which perturb normal pre-mRNA splicing are significant contributors to human disease. We used exome sequencing data from 7,833 probands with developmental disorders (DD) and their unaffected parents, as well as >60,000 aggregated exomes from the Exome Aggregation Consortium, to investigate selection around the splice site, and quantify the contribution of splicing mutations to DDs. Patterns of purifying selection, a deficit of variants in highly constrained genes in healthy subjects and excess de novo mutations in patients highlighted particular positions within and around the consensus splice site of greater functional relevance. Using mutational burden analyses in this large cohort of proband-parent trios, we could estimate in an unbiased manner the relative contributions of mutations at canonical dinucleotides (73%) and flanking non-canonical positions (27%), and calculated the positive predictive value of pathogenicity for different classes of mutations. We identified 18 patients with likely diagnostic de novo mutations in dominant DD-associated genes at non-canonical positions in splice sites. We estimate 35-40% of pathogenic variants in non-canonical splice site positions are missing from public databases.

genetics↗

bioRxiv2017

Autosomal recessive coding variants explain only a small proportion of undiagnosed developmental disorders in the British Isles

Large exome-sequencing datasets offer an unprecedented opportunity to understand the genetic architecture of rare diseases, informing clinical genetics counseling and optimal study designs for disease gene identification. We analyzed 7,448 exome-sequenced families from the Deciphering Developmental Disorders study, and, for the first time, estimated the causal contribution of recessive coding variation exome-wide. We found that the proportion of cases attributable to recessive coding variants is surprisingly low in patients of European ancestry, at only 3.6%, versus 50% of cases explained by de novo coding mutations. Surprisingly, we found that, even in European probands with affected siblings, recessive coding variants are only likely to explain ~12% of cases. In contrast, they account for 31% of probands with Pakistani ancestry due to elevated autozygosity. We tested every gene for an excess of damaging homozygous or compound heterozygous genotypes and found three genes that passed stringent Bonferroni correction: EIF3F, KDM5B, and THOC6. EIF3F is a novel disease gene, and KDM5B has previously been reported as a dominant disease gene. KDM5B appears to follow a complex mode of inheritance, in which heterozygous loss-of-function variants (LoFs) show incomplete penetrance and biallelic LoFs are fully penetrant. Our results suggest that a large proportion of undiagnosed developmental disorders remain to be explained by other factors, such as noncoding variants and polygenic risk.

genetics↗

bioRxiv2016

Confronting preferential sampling in wildlife surveys: diagnosis and model-based triage

SummaryO_LIWildlife surveys are often used to estimate the density, abundance, or distribution of animal populations. Recently, model-based approaches to analyzing survey data have become popular because one can more readily accommodate departures from pre-planned survey routes and construct more detailed maps than one can with design-based procedures.\nC_LIO_LISpecies distribution models fitted to wildlife survey data often make the implicit assumption that locations chosen for sampling and animal abundance at those locations are conditionally independent given modeled covariates. However, this assumption is likely violated in many cases when survey effort is non-randomized, leading to preferential sampling.\nC_LIO_LIWe develop a hierarchical statistical modeling framework for detecting and alleviating the biasing effects of preferential sampling in species distribution models fitted to count data. The approach works by jointly modeling wildlife state variables and the locations selected for sampling, and specifying a dependent correlation structure between the two models.\nC_LIO_LIUsing simulation, we show that moderate levels of preferential sampling can lead to large (e.g. 40%) bias in estimates of animal density, and that our modeling approach can considerably reduce this bias.\nC_LIO_LIWe apply our approach to aerial survey counts of bearded seals (Erignathus barbatus) in the eastern Bering Sea. Models that included a preferential sampling effect led to lower estimates of abundance than models without, but the effect size of the preferential sampling parameter decreased in models that included explanatory environmental covariates.\nC_LIO_LIWhen wildlife surveys are conducted without a well-defined sampling frame, ecologists should recognize the potentially biasing effects of preferential sampling. Joint models, such as those described in this paper, can be used to test and correct for such biases. Predictive covariates are also useful for bias reduction, but ultimately the best way to avoid preferential sampling bias is to incorporate design-based principles such as randomization and/or systematic sampling into survey design.\nC_LI

ecology↗