Biology subjects

Stein, C.

Publications and source records attributed to Stein, C..

10 recordsLinked to original sources

bioRxiv2025

The combination of elevated neuronal activity and mitochondrial damage induces Pink1-dependent mitophagy in axons

Mitochondria are critical for synaptic function. At the synapse, mitochondria produce ATP and buffer calcium, both of which are required for synapse function. Defects in mitochondrial maintenance are linked to neurodegenerative disease, yet we know little about what regulates the need for mitophagy at the synapse. We assessed the impact of neuron type, activity, and mitochondrial damage on mitophagy rate in axons of larval zebrafish. Using electron and confocal microscopy, we show that mitophagy occurs in the axon terminal of postsynaptic sensory neurons and presynaptic motor neurons at similar rates. Increasing neuronal activity or mitochondria damage does not impact the amount of mitophagy in axons. Only by combining neuronal activity and mitochondrial damage does the rate of mitophagy increase in the axon and this increase requires Pink1. Together, our data support a model in which increased mitophagic demand in axons is rare and uniquely sensitive to Pink1 disruption.

neuroscience↗

bioRxiv2025

Retrograde mitochondrial transport regulates mitochondrial biogenesis in zebrafish neurons

To maintain a functional mitochondrial population in a long-lived cell like a neuron, mitochondria must be continuously replenished through the process of mitochondrial biogenesis. Because most mitochondrial proteins are nuclear encoded, mitochondrial biogenesis requires communication between mitochondria and the nucleus. This can be a challenge in a large, compartmentalized cell like a neuron in which a large portion of the mitochondrial population is in neuronal compartments far from the nucleus. Using in vivo assessments of mitochondrial biogenesis in zebrafish neurons, we determined that mitochondrial transport between distal axonal compartments and the cell body is required for sustained mitochondrial biogenesis. Estrogen-related receptor transcriptional activation links transport with nuclear expression of mitochondrial genes. Together, our data support a role for retrograde feedback between axonal mitochondria and the nucleus for regulation of mitochondrial biogenesis in neurons.

neuroscience↗

bioRxiv2025

Distinct antibody-based signatures and functionality distinguish latent and active pediatric tuberculosis

BackgroundTuberculosis (TB), caused by Mycobacterium tuberculosis (Mtb), is among the leading causes of death from an infectious agent among children worldwide. Children represent a particularly vulnerable population due to the greater challenges in diagnosis and the higher risk of progression to severe forms of the disease. However, whether different pediatric outcomes relate to distinct immunologic responses remains incompletely understood. Emerging data suggest that Mtb-specific humoral immune responses represent a correlate of protection against Mtb both following natural infection and vaccination. MethodsTo determine if immune profiles can distinguish children across the spectrum from Mtb infection to TB disease, as well as children with TB from non-TB lower respiratory tract infection, we mapped the humoral immune response across a panel of 4 dozen Mtb antigens across children presenting with symptoms of active TB (ATB), children with evidence of latent TB infection (LTBI) and children exhibiting non-TB lower respiratory tract infection (non-TB LRTI). Using a custom Luminex assay, Mtb-specific antibody subclass/isotype, Fc receptor (FcR) binding profiles, and functions were profiled across the pediatric groups. FindingsA robust humoral immune response was observed in children with active TB compared to non-TB LRTI, marked by a strong IgA response, that exhibited high FcR binding. Conversely, children with LTBI uniquely elicited Mtb-specific antibodies with enhanced opsinophagocytic Fc{gamma}R2A binding, as well as a higher capacity to activate NK cells and neutrophils. InterpretationThere are significant differences in humoral immune profiles across the landscape of pediatric TB, potentially contributing to differential mycobacterial control, and highlighting biomarkers that could guide both diagnostic and therapeutic approaches. FundingUS National Institutes of Health.

immunology↗

bioRxiv2024

CCSer2 gates dynein activity at the cell periphery

Cytoplasmic dynein-1 (dynein) is a microtubule-associated, minus end-directed motor that traffics hundreds of different cargos. Dynein must discriminate between cargos and traffic them at the appropriate time from the correct cellular region. How dyneins trafficking activity is regulated in time or cellular space remains poorly understood. Here, we identify CCSer2 as the first known protein to gate dynein activity in the spatial dimension. CCSer2 promotes the migration of developing zebrafish primordium cells and of cultured human cells by facilitating the trafficking of cargos that are acted on by cortically localized dynein. CCSer2 inhibits the interaction between dynein and its regulator Ndel1 exclusively at the cell periphery, resulting in localized dynein activation. Our findings suggest that the spatial specificity of dynein is achieved by the localization of proteins that disinhibit Ndel1. We propose that CCSer2 defines a broader class of proteins that activate dynein in distinct microenvironments via Ndel1 inhibition.

cell biology↗

bioRxiv2024

Epigenetic programming of host lipid metabolism associates with resistance to TST/IGRA conversion after exposure to Mycobacterium tuberculosis

Mycobacterium tuberculosis (Mtb) exposure leads to a range of outcomes including clearance, latent TB infection (LTBI), and pulmonary tuberculosis (TB). Some heavily exposed individuals resist tuberculin skin test (TST) and interferon gamma release assay (IGRA) conversion (RSTR), which suggests that they employ IFN{gamma}-independent mechanisms of Mtb control. Here, we compare monocyte epigenetic profiles of RSTR and LTBI from a Ugandan household contact cohort. Chromatin accessibility did not differ between uninfected RSTR and LTBI monocytes. In contrast, methylation significantly differed at 174 CpG sites and across 63 genomic regions. Consistent with previous transcriptional findings in this cohort, differential methylation was enriched in lipid and cholesterol associated pathways including in the genes APOC3, KCNQ1, and PLA2G3. In addition, methylation was enriched in Hippo signaling, which is associated with cholesterol homeostasis and includes CIT and SHANK2. Lipid export and Hippo signaling pathways were also associated with gene expression in response to Mtb in RSTR as well as IFN stimulation in monocyte-derived macrophages (MDMs) from an independent healthy donor cohort. Moreover, serum-derived HDL from RSTR had elevated ABCA1-mediated cholesterol efflux capacity (CEC) compared to LTBI. Our findings suggest that resistance to TST/IGRA conversion is linked to regulation of lipid accumulation in monocytes, which could facilitate early Mtb clearance among RSTR subjects through IFN{gamma}-independent mechanisms. IMPORTANCETuberculosis (TB) remains an enduring global health challenge with millions of deaths and new cases each year. Despite recent advances in TB treatment, we lack an effective vaccine or a durable cure. While heavy exposure to Mycobacterium tuberculosis often results in latent TB latent infection (LTBI), subpopulations exist who are either resistant to infection or contain Mtb with IFN{gamma}-independent mechanisms not indicative of LTBI. These resisters provide an opportunity to investigate mechanisms of TB disease and discover novel therapeutic targets. Here, we compare monocyte epigenetic profiles of RSTR and LTBI from a Ugandan household contact cohort. We identify methylation signatures in host lipid and cholesterol pathways with potential relevance to early TB clearance before the sustained IFN responses indicative of LTBI. This adds to a growing body of literature linking TB disease outcomes to host lipids.

immunology↗

bioRxiv2023

Clinal variation in drought response is consistent across life stages but not between native and non-native ranges

Clinal variation, i.e., intraspecific variation that corresponds to environmental gradients, is common in widely distributed species. Studies on clinal variation across multiple ranges and life stages are lacking, but can enhance our understanding of specie[s] adaptive potential to abiotic environments and may aid in predicting future species distributions. This study examined clinal variation in drought responses of 59 Conyza canadensis populations across large aridity gradients from the native and non-native ranges in three greenhouse studies. Experimental drought was applied to recruitment, juveniles, and adult stages. Drought reduced growth at all three life stages. However, contrasting patterns of clinal variation emerged between the two ranges. Native populations from xeric habitats were less inhibited by drought than mesic populations, but such clinal variation was not apparent for non-native populations. These range-specific patterns of clinal variation were consistent across the life stages. The experiments suggest that invaders may succeed without complete local adaptation to their new abiotic environments, and that long-established invaders may still be evolving to the abiotic environment. These findings may explain lag times in some invasions and raise concern about future expansions.

evolutionary biology↗

bioRxiv2023

Nanopore sequencing for accurate bacterial outbreak tracing

Our study investigated the effectiveness of Oxford Nanopore Technologies for accurate outbreak tracing by resequencing 33 isolates of a three-year-long Klebsiella pneumoniae outbreak with Illumina short read sequencing data as the point of reference. We detected considerable base errors through cgMLST and phylogenetic analysis of genomes sequenced with Oxford Nanopore Technologies, leading to the false exclusion of some outbreak-related strains from the outbreak cluster. Nearby methylation sites cause these errors and can also be found in other species besides K. pneumoniae. Based on this data, we explored PCR-based sequencing and a masking strategy, which both successfully addressed these inaccuracies and ensured accurate outbreak tracing. We offer our masking strategy as a bioinformatic workflow (MPOA is freely available on GitHub under the GNUv3 license: github.com/replikation/MPOA) to identify and mask problematic genome positions in a reference-free manner. Our research highlights limitations in using Oxford Nanopore Technologies for sequencing prokaryotic organisms, especially for investing outbreaks. For time-critical projects that cannot wait for further technological developments by Oxford Nanopore Technologies, our study recommends either PCR-based sequencing or using our provided bioinformatic workflow. We would advise that read mapping-based quality control of genomes should be provided when publishing results.

molecular biology↗

bioRxiv2022

kimma: flexible linear mixed effects modeling with kinship for RNA-seq data

We introduce kimma (Kinship In Mixed Model Analysis), an open-source R package for flexible linear mixed effects modeling of RNA-seq including covariates, weights, random effects, covariance matrices, and fit metrics. In simulated datasets, kimma detects differentially expressed genes (DEGs) with similar specificity, sensitivity, and computational time as limma unpaired and dream paired models. Unlike other software, kimma supports covariance matrices as well as fit metrics like AIC. Utilizing genetic kinship covariance, kimma revealed that kinship impacts model fit and DEG detection in a related cohort. Thus, kimma equals or outcompetes current DEG pipelines in sensitivity, computational time, and model complexity.

bioinformatics↗

bioRxiv2021

Modelling altered signalling of G-protein coupled receptors in inflamed environment to advance drug design

We previously reported the successful design, synthesis and testing of the prototype opioid painkiller NFEPP that does not elicit adverse side effects. Uniquely, this design was based on mathematical modelling of extracellular interactions between G-protein coupled receptors (GPCRs) and ligands, recognizing that GPCRs function differently under pathological versus healthy conditions. We now present a novel stochastic model of GPCR function that includes intracellular dissociation of G-protein subunits and modulation of plasma membrane calcium channels associated with parameters of inflamed and healthy tissue (pH, radicals). The model is validated against in vitro experimental data for NFEPP and fentanyl ligands at different pH values. We found markedly reduced calcium channel inhibition induced by NFEPP at normal pH compared to lower pH, in contrast to the effect of fentanyl, and enhanced constitutive G-protein activation but lower probability of ligand binding with increasing radical concentrations. By means of molecular dynamics simulations, we also assessed qualitative changes of reaction rates due to additional disulfide bridges inside the GPCR binding pocket. The results suggest that, compared to radicals, low pH is a more important determinant of overall GPCR function in an inflamed environment. Future drug design efforts should take this into account.

systems biology↗

bioRxiv2019

Interaction between host genes and M. tuberculosis lineage can affect tuberculosis severity: evidence for co-evolution

Genetic studies of both the human host and Mycobacterium tuberculosis (MTB) demonstrate independent association with tuberculosis (TB) risk. However, neither explains a large portion of disease risk or severity. Based on studies in other infectious diseases and animal models of TB, we hypothesized that the genomes of the two interact to modulate risk of developing active TB or increasing the severity of disease, when present. We examined this hypothesis in our TB household contact study in Kampala, Uganda, in which there were 3 MTB lineages of which L4-Ugandan (L4.6) is the most recent. TB severity, measured using the Bandim TBscore, was modeled as a function of host SNP genotype, MTB lineage, and their interaction, within two independent cohorts of TB cases, N=113 and 122. No association was found between lineage and severity, but association between multiple polymorphisms in IL12B and TBscore was replicated in two independent cohorts (most significant rs3212227, combined p=0.0006), supporting previous associations of IL12B with TB susceptibility. We also observed significant interaction between a single nucleotide polymorphism (SNP) in SLC11A1 and the L4-Ugandan lineage in both cohorts (rs17235409, meta p=0.0002). Interestingly, the presence of the L4-Uganda lineage in the presence of the ancestral human allele associated with more severe disease. These findings demonstrate that IL12B is associated with severity of TB in addition to susceptibility, and that the association between TB severity and human genetics can be due to an interaction between genes in the two species, providing evidence of host-pathogen coevolution in TB.\n\nAUTHOR SUMMARYSusceptibility to tuberculosis (TB) is affected by genetic variation in both the human host and the causative bacterium, Mycobacterium tuberculosis. However, prior studies of the genetics of each species have not explained a large part of TB risk. The possibility exists that risk can be better estimated from patterns of variation in two species as a unit, such that some combinations provide increased risk, or in the presence of TB, increased disease severity. We hypothesized that alleles in the two species that have co-existed for long periods are more likely to reduce disease severity so as to promote prolonged co-occurrence. We tested this by studying TB severity in two patient cohorts from Uganda for which paired MTB-human DNA were available. We examined severity, as measured by the Bandim TBscore, and assessed whether there was an interaction between MTB lineage and SNPs in the host with this metric. Our results indicate that the most recent TB lineage (L4.6/Uganda) when found together with an ancestral allele in SLC11A1 resulted in more severe disease. This finding is consistent with the conclusion that MTB and human have coevolved to modulate TB severity.

evolutionary biology↗