Biology subjects

Jacob, C.

Publications and source records attributed to Jacob, C..

8 recordsLinked to original sources

bioRxiv2026

Striatal cholinergic and dopaminergic driven astrocyte Ca2+ activity is disrupted in Parkinsonian mice

Brain neuromodulatory systems exert powerful effects on local neuronal circuit function and behavior. In addition to classical actions directly on neurons, growing evidence indicates that neuromodulators also recruit Ca2+-dependent astrocyte mechanisms to regulate synaptic plasticity and network function. The dorsal striatum integrates cortical and thalamic inputs under strong dopamine (DA) and acetylcholine (ACh) neuromodulatory control. To what extent the circuit and behavioral effects of striatal ACh and DA depend on astrocyte Ca2+ activity remains unclear. We show that locomotion elicits robust DA, ACh, and astrocyte Ca2+ activity in the dorsolateral striatum (DLS). DA and ACh release exhibits a negative correlation on a fast time scale but shows a positive correlation during continuous locomotion as slower astrocyte Ca2+ activity builds. Higher ACh and DA release is associated with higher astrocyte events, suggesting that both neurotransmitters drive astrocyte activity. In agreement, pharmacological blockade of muscarinic ACh or D1/D2 DA receptors decreases locomotion-evoked astrocyte Ca2+. Closed-loop optogenetic inhibition of striatal cholinergic interneurons (CINs) during locomotion reduces astrocyte Ca2+ activity, demonstrating a causal contribution of ACh release to astrocyte activity. Locomotion related ACh release was severely compromised in a mouse model of Parkinsons disease (PD), with the dual loss of DA and ACh attenuating astrocyte Ca2+ activity. Facilitating astrocyte cholinergic signaling via chemogenetics improved both calcium activity and motor deficits in our recent work. Thus, the pathophysiology of PD in part involves attenuated astrocyte Ca2+ signaling, placing these non-neuronal cells as a prime underexplored therapeutic target for PD.

neuroscience↗

bioRxiv2025

Expanding Targeted Instrumentation for Discovery Applications: Complement Reporter Ion Quantification with a Quadrupole-Ion Trap Instrument

Proteomics workflows have traditionally been divided into discovery-based and targeted approaches, with instrumentation optimized specifically for each. Discovery experiments typically utilize high-resolution analyzers while targeted workflows rely on the sensitivity and specificity of triple quadrupole systems. Recently, a quadrupole-ion trap instrument (Stellar MS) demonstrated superior performance for targeted applications compared to conventional triple quadrupoles. In this study, we expand the capabilities of this platform to multiplexed shotgun proteomics using complement reporter ion quantification in an ion trap (iTMTproC). Benchmarking experiments with defined standards show that iTMTproC achieves quantification accuracy and interference reduction comparable to MultiNotch MS3 on the Orbitrap Fusion Lumos, a dedicated quadrupole-ion trap-Orbitrap tribrid instrument optimized for this purpose. Notably, iTMTproC quantifies slightly more proteins than MultiNotch MS3. We further validate this approach through a developmental time-series analysis of frog embryos, obtaining proteomic data nearly indistinguishable from MultiNotch MS3, with slightly increased protein quantification depth. These findings significantly extend the functionality of targeted instrumentation, underscoring the versatility of quadrupole-ion trap systems and providing cost-effective access to highly accurate, multiplexed quantitative shotgun proteomics.

systems biology↗

bioRxiv2025

Development and Clinical Evaluation of a Multiplexed Health Surveillance Panel Using Ultra High-Throughput PRM-MS in an Inflammatory Bowel Disease Cohort.

Despite advances in clinical proteomics, translating protein biomarker discoveries into clinical use remains challenging due to the technical complexity of the validation process. Targeted MS-based proteomics approaches such as parallel reaction monitoring (PRM) offer sensitive and specific assays for biomarker translation. In this study, we developed a multiplex PRM assay using the Stellar mass spectrometry platform to quantify 57 plasma proteins, including 21 FDA-approved proteins. Loading curves (11-points) were performed at 4 sample throughputs (100, 144, 180, and 300 samples per day) using independent, optimized, and scheduled PRM methods. Following optimization, an inflammatory bowel disease (IBD) cohort of plasma samples (493 IBD, 509 matched controls) was analyzed at a throughput of 180 SPD. To monitor system performance, the study also included 1,000 additional injections for system suitability tests, low-, middle-, and high-quality controls, washes, and blanks. Using this approach, we observed high quantifiability (linearity, sensitivity, reproducibility) in the PRM assay and consistent in data acquisition across a large cohort. We also validated the candidate IBD markers, C-reactive protein and orosomucoid protein, identified in a recent discovery experiment.

biochemistry↗

bioRxiv2024

Exploiting the inherent promiscuity of the acyl transferase of the stambomycin polyketide synthase for the mutasynthesis of analogues

The polyketide specialized metabolites of bacteria are attractive targets for generating analogues, with the goal of improving their pharmaceutical properties. Here, we aimed to produce C-26 derivatives of the giant anti-cancer stambomycin macrolides using a mutasynthesis approach, as this position has been shown previously to directly impact bioactivity. For this, we leveraged the intrinsically broad specificity of the acyl transferase domain (AT12) of the modular polyketide synthase (PKS), which is responsible for the alkyl branching functionality at this position. Feeding of a panel of synthetic and commercially available dicarboxylic acid mutasynthons to an engineered strain of Streptomyces ambofaciens (Sa) deficient in synthesis of the native -carboxyacyl-CoA extender units, resulted in six new series of stambomycin derivatives as judged by LC-HRMS and NMR. Notably, the highest product yields were observed for substrates which were only poorly accepted when AT12 was transplanted into a different PKS module, suggesting a critical role for domain context in the overall functioning of PKS proteins. We also demonstrate the superiority of this mutasynthesis approach - both in terms of absolute titers and yields relative to the parental compounds - in comparison to the alternative precursor-directed strategy in which monoacid building blocks are supplied to the wild type strain. We further identify a malonyl-CoA synthetase, MatB_Sa, with specificity distinct from previously identified promiscuous enzymes, making it a useful addition to a mutasynthesis toolbox for generating atypical, CoA activated extender units. Finally, we show that two of the obtained (deoxy)-butyl-stambomycins exhibit antibacterial and antiproliferative activities similar to the parental stambomycins, while an unexpected butyl-demethyl congener is less potent. Overall, this works confirms the interest of biosynthetic pathways which combine a dedicated route to extender unit synthesis and a broad specificity AT domain for producing bioactive derivatives of fully-elaborated complex polyketides.

synthetic biology↗

bioRxiv2024

Nuclease-Resistant L-DNA Tension Probes Enable Long-TermForce Mapping of Single Cells and Cell Consortia

DNA-based tension probes with precisely programmable force response provide important insights into cellular mechanosensing. However, their degradability in cell culture limits their use for long-term imaging, for instance, when cells migrate, divide, and differentiate. This is a critical limitation for providing insights into mechanobiology for these longer-term processes. Here, we present DNA-based tension probes that are entirely designed based on the stereoisomer of biological D-DNA, i.e., L-DNA. We demonstrate that L-DNA tension probes are essentially indestructible by nucleases and provide days-long imaging without significant loss in image quality. We also show their superiority already for short imaging times commonly used for classical D-DNA tension probes. We showcase the potential of these resilient probes to image minute movements, and for generating long term force maps of single cells and for the first time, of collectively migrating cell populations.

biophysics↗

bioRxiv2024

A novel hybrid high speed mass spectrometer allows rapid translation from biomarker candidates to targeted clinical tests using 15N labeled proteins

Recent developments in affinity binder or mass spectrometry (MS)-based plasma proteomics are now producing panels of potential biomarker candidates for diagnosis or prognosis. However, clinical validation and implementation of these biomarkers remain limited by the reliance on dated triple quadrupole MS technology. Here, we evaluate a novel hybrid high-speed mass spectrometer, Stellar MS, which integrates the robustness of triple quadrupoles with the enhanced capabilities of an advanced linear ion trap analyzer. This instrument allows for extremely rapid and sensitive parallel reaction monitoring (PRM) and MS3 targeting. The Stellar MS allowed targeting thousands of peptides originally measured on Orbitrap Astral MS, achieving high reproducibility and low coefficients of variation (CV) as well as sensitivity and specificity sufficient for more than the top 1000 plasma proteins. Furthermore, we developed targeted assays for alcohol-related liver disease (ALD) biomarkers, showcasing the potential of Stellar MS in clinical applications. Absolute quantification is typically a requirement for clinical assays and we explore the use of 15 N labeled protein standards in a rapid, streamlined and generic manner. Our results indicate that Stellar MS can bridge the gap between proteomics discovery and routine clinical testing, enhancing the diagnostic and prognostic utility of protein biomarkers.

biochemistry↗

bioRxiv2023

Repair oligodendrocytes demyelinating and disintegrating damaged axons after injury

After a spinal cord injury, axons fail to regrow, which results in permanent loss of function1. This is in contrast with peripheral axons that can regrow efficiently after injury2. These differences are partly due to the different plasticity of myelinating cells, Schwann cells and oligodendrocytes, in these two systems3. The molecular mechanisms underlying this different plasticity remain however poorly understood. Here, we show that the phosphatase Dusp64 is a master inhibitor of oligodendrocyte plasticity after spinal cord injury. Dusp6 is rapidly downregulated in Schwann cells and upregulated in oligodendrocytes after axon injury. Simultaneously, the MAP kinases ERK1/2 are activated and the transcription factor c-Jun is upregulated in Schwann cells5,6, but not in oligodendrocytes. Ablation or inactivation of Dusp6 induces rapid ERK1/2 phosphorylation, c-Jun upregulation and filopodia formation in oligodendrocytes, leading to mechanically-induced, fast disintegration of distal ends of injured axons, myelin clearance and axonal regrowth. Together, our findings provide understanding of the mechanisms underlying the different plasticity of Schwann cells and oligodendrocytes after injury and a method to convert mature oligodendrocytes exhibiting inhibitory cues for axonal regrowth into repair oligodendrocytes reminiscent of repair Schwann cells. We show that repair oligodendrocytes successfully increase the compatibility of the spinal cord environment with axonal regrowth after injury, suggesting a potential use of repair oligodendrocytes as future therapeutic approach to treat spinal cord injuries.

neuroscience↗

bioRxiv2020

Moderate prenatal ethanol exposure increases total length of L1 axons in E15.5 mice

Public health campaigns broadcast the link between heavy alcohol consumption during pregnancy and physical, cognitive, and behavioral birth defects; however, they appear less effective in deterring moderate consumption prevalent in women who are pregnant or of childbearing age. The incidence of mild Fetal Alcohol Spectrum Disorders (FASD) is likely underestimated because the affected individuals lack physical signs such as retarded growth and facial dysmorphology and cognitive/behavioral deficits are not commonly detected until late childhood. Sensory information processing is distorted in FASD, but alcohol effects on the development of axons that mediate these functions are not widely investigated. We hypothesize that alcohol exposure alters axon growth and guidance contributing to the aberrant connectivity that is a hallmark of FASD. To test this, we administered alcohol to pregnant dams from embryonic day (E) 7.5 to 14.5, during the time that axons which form the major forebrain tracts are growing. We found that moderate alcohol exposure had no effect on body weight of E15.5 embryos, but significantly increased the length of L1+ axons. Our findings support our hypothesis. Future studies will investigate cellular, molecular, and functional mechanisms that underlie these effects.

developmental biology↗