Biology subjects

Koch, R.

Publications and source records attributed to Koch, R..

9 recordsLinked to original sources

bioRxiv2025

Endosymbiont hijacking of acylcarnitines regulates insect vector fecundity by suppressing the viability of stored sperm

Competition between insects and their endosymbiotic bacteria for environmentally limited nutrients can compromise the fitness of both organisms. Tsetse flies, the vectors of pathogenic African trypanosomes, harbor a species and population-specific consortium of vertically transmitted endosymbiotic bacteria that range on the functional spectrum from mutualistic to parasitic. Tsetses indigenous microbiota can include a member of the genus Spiroplasma, and infection with this bacterium causes fecundity-reducing phenotypes in the fly that include a prolonged gonotrophic cycle and a reduction in the motility of stored spermatozoa post-copulation. Herein we demonstrate that Spiroplasma and tsetse spermatozoa compete for fly-derived acylcarnitines, which in other bacteria and animals are used to maintain cell membranes and produce energy. The fat body of mated female flies increases acylcarnitine production in response to infection with Spiroplasma. Additionally, their spermathecae (sperm storage organs), and likely the sperm within, up-regulate expression of carnitine O-palmitoyltransferase-1, which is indicative of increased acylcarnitine metabolism and thus increased energy demand and energy production in this organ. These compensatory measures are insufficient to rescue the motility defect of spermatozoa stored in the spermathecae of Spiroplasma-infected females and thus results in reduced fly fecundity. Tsetses taxonomically simple and highly tractable indigenous microbiota make the fly an efficient model system for studying the biological processes that facilitate the maintenance of bacterial endosymbioses, and how these relationships impact conserved mechanisms (mammalian spermatozoa also use acylcarnitines as an energy source) that regulated animal host fecundity. In the case of insect pests and vectors, a better understanding of the metabolic mechanisms that underlie these associations can lead to the development of novel control strategies. Author SummaryAnimals and the endosymbiotic bacteria that live inside them often compete for nutrients that both organisms require in order to survive. Tsetse flies, which transmit pathogenic African trypanosomes, can house several endosymbionts that have different impacts on their hosts physiological well-being. Female tsetse flies that are infected with one of these bacteria, Spiroplasma, produce fewer offspring than do their uninfected counterparts. In this study we demonstrate that the bacterium and the flys sperm cells (spermatozoa) compete for a specific type of lipid called acylcarnitines. When mated female tsetse flies are experimentally manipulated to produce less acylcarnitine Spiroplasma density decreases. Additionally, spermatozoa stored in the sperm storage organs of acylcarnitine depleted females lose motility and are eventually resorbed, thus rendering the females reproductively sterile. These findings mechanistically demonstrate how endosymbiotic bacteria can manipulate their hosts reproductive potential. In the case of arthropods that transmit pathogenic microbes, this relationship has significant implications for disease transmission and epidemiology.

microbiology↗

bioRxiv2025

The Chlamydia trachomatis secreted effector protein CT181 binds to Mcl-1 to prolong neutrophil survival

Chlamydia trachomatis (C.t) infections can lead to severe complications due to the pathogens ability to evade the host immune response, often resulting in asymptomatic infections. The mechanisms underlying this immune subversion remain incompletely understood but likely involve specific bacterial effector proteins. Here, we identify CT181 as a novel effector that directly binds to Mcl-1, a key regulator of neutrophil survival. While a C.t. CT181 mutant exhibited only modest defects in epithelial cell replication and inclusion development, it was essential for C.t. survival in neutrophils, correlating with Mcl-1 stabilization. Using a murine infection model, we demonstrate that CT181 is required for C.t. colonization and cytokine production in vivo. Our findings establish CT181 as the first bacterial effector protein known to bind Mcl-1 to enhance neutrophil survival, revealing a critical strategy by which C.t. promotes immune dysregulation, facilitating bacterial persistence while driving C.t. pathogenesis.

microbiology↗

bioRxiv2023

Examining the potential involvement of NONO in TDP-43 proteinopathy in Drosophila

The misfolding and aggregation of TAR DNA binding protein-43 (TDP-43), leading to the formation of cytoplasmic inclusions, emerge as a key pathological feature in a spectrum of neurodegenerative diseases, including amyotrophic lateral sclerosis (ALS) and frontotemporal lobar dementia (FTLD). TDP-43 shuttles between the nucleus and cytoplasm but forms nuclear bodies (NBs) in response to stress. These NBs partially colocalize with nuclear speckles and paraspeckles that sequester RNAs and proteins, thereby regulating many cellular functions. The laboratory of Steven Brown has recently found that the non-POU domain-containing octamer-binding protein (NONO), a component of paraspeckles, forms novel nuclear speckle-like structures in mouse cortical neurons in response to stress and sleep deprivation. These findings suggest the possibility of a functional link between NONO and TDP-43, potentially contributing to TDP-43 proteinopathy. Here, we demonstrate that loss of function in the Drosophila homolog of NONO, no on or off transient A (NonA), exacerbates pathological phenotypes caused by TDP-43 gain of function, leading to locomotor defects and life span shortening. These results provide supporting evidence for the functional link between NONO and TDP-43 and lay the foundation for dissecting underlying mechanisms.

neuroscience↗

bioRxiv2023

Resolving spatiotemporal electrical signaling within the islet via CMOS microelectrode arrays

Glucose-stimulated beta-cells synchronize calcium waves across the islet to recruit more beta-cells for insulin secretion. Compared to calcium dynamics, the formation and cell-to-cell propagation of electrical signals within the islet are poorly characterized. To determine factors that influence the propagation of electrical activity across the islet underlying calcium oscillations and beta-cell synchronization, we used high-resolution CMOS multielectrode arrays (MEA) to measure voltage changes associated with the membrane potential of individual cells within intact mouse islets. We measured both fast (milliseconds, spikes) and slow (seconds, waves) voltage changes and analyzed the spatiotemporal voltage dynamics. Treatment of islets from C57BL6 mice with increasing glucose concentrations revealed that single spike activity and wave signal velocity were both glucose-dependent. A repeated glucose stimulus involved a highly active subset of cells in terms of spike activity. When islets were pretreated for 72 hours with glucolipotoxic medium, the wave velocity was significantly reduced. Network analysis confirmed that the synchrony of islet cells was affected due to slower propagating electrical waves and not due to altered spike activity. In summary, this approach provided novel insight regarding the propagation of electrical activity and opens a wide field for further studies on signal transduction in the islet cell network. Article HighlightsThis study presents a new method for characterizing islet spatiotemporal electrical dynamics and subpopulations of beta-cells. We asked whether a high-resolution CMOS-MEA is suited to detect electrical signals on a level close to single cells, and whether we can track the propagation of electrical activity through the islet on a cellular scale. A highly active subpopulation of islet cells was identified by action potential-like spike activity, whereas slower waves were a measure for synchronized electrical activity. Further, propagating waves were slowed by glucolipotoxicity. The technique is a useful tool for exploring the pancreatic islet network in health and disease.

physiology↗

bioRxiv2023

Evolution of circadian behavioral plasticity through cis-regulatory divergence of a neuropeptide gene

Widely-distributed species experience substantial environmental variation, which they often accommodate through behavioral plasticity. Although this ability is integral to fitness, we have little understanding of the mechanistic basis by which plasticity evolves. One factor that varies seasonally and by latitude is photoperiod (day length). Many organisms, including the cosmopolitan Drosophila melanogaster display circadian plasticity, adjusting to fluctuating photoperiod by varying the timing of their activity to coincide with changing dawn/dusk intervals1. Here, we compare D. melanogaster with the closely-related ecological specialist Drosophila sechellia, an equatorial island endemic that experiences minimal photoperiod variation, to investigate the molecular-genetic basis of circadian plasticity evolution2,3. We discover that D. sechellia displays exceptionally little circadian plasticity compared to D. melanogaster and other non-equatorial drosophilids. Through a screen of circadian mutants in D. melanogaster/D. sechellia hybrids, we identify a role of the neuropeptide Pigment-dispersing factor (Pdf) in this loss. While the coding sequence of Pdf is conserved, we show that Pdf has undergone cis-regulatory divergence in these drosophilids. We document species-specific temporal dynamic properties of Pdf RNA and protein expression, as well as Pdf neuron morphological plasticity, and demonstrate that modulating Pdf expression in D. melanogaster can influence the degree of behavioral plasticity. Furthermore, we find that the Pdf regulatory region exhibits signals of selection across populations of D. melanogaster from different latitudes. Finally, we provide evidence that plasticity confers a selective advantage for D. melanogaster at higher latitudes, while D. sechellia likely suffers fitness costs through reduced copulation success outside its range. Our work defines Pdf as a locus of evolution for circadian plasticity, which might have contributed to both D. melanogasters global distribution and D. sechellias habitat specialization. Moreover, together with spatial changes in Pdf expression reported in high-latitude drosophilid species4,5, our findings highlight this neuropeptide gene as a hotspot for circadian plasticity evolution.

evolutionary biology↗

bioRxiv2023

Identifying the core genome of the nucleus-forming bacteriophage family and characterization of Erwinia phage RAY

We recently discovered that some bacteriophages establish a nucleus-like replication compartment (phage nucleus), but the core genes that define nucleus-based phage replication and their phylogenetic distribution were unknown. By studying phages that encode the major phage nucleus protein chimallin, including previously sequenced yet uncharacterized phages, we discovered that chimallin-encoding phages share a set of 72 highly conserved genes encoded within seven distinct gene blocks. Of these, 21 core genes are unique to this group, and all but one of these unique genes encode proteins of unknown function. We propose that phages with this core genome comprise a novel viral family we term Chimalliviridae. Fluorescence microscopy and cryo-electron tomography studies of Erwinia phage vB_EamM_RAY confirm that many of the key steps of nucleus-based replication encoded in the core genome are conserved among diverse chimalliviruses, and reveal that non-core components can confer intriguing variations on this replication mechanism. For instance, unlike previously studied nucleus-forming phages, RAY doesnt degrade the host genome, and its PhuZ homolog appears to form a five-stranded filament with a lumen. This work expands our understanding of phage nucleus and PhuZ spindle diversity and function, providing a roadmap for identifying key mechanisms underlying nucleus-based phage replication.

microbiology↗

bioRxiv2022

Pharmacological PP2A reactivation overcomes multikinase inhibitor tolerance across brain tumor cell models

BackgroundGlioblastoma is characterized by hyperactivation of kinase signaling pathways. Regardless, most glioblastoma clinical trials targeting kinase signaling have failed. We hypothesized that overcoming the glioblastoma kinase inhibitor tolerance requires efficient shut-down of phosphorylation-dependent signaling rewiring by simultaneous inhibition of multiple critical kinases combined with reactivation of Protein Phosphatase 2A (PP2A). MethodsLive-cell imaging and colony growth assays were used to determine long-term impact of therapy effects on ten brain tumor cell models. Immunoblotting, MS-phosphoproteomics, and Seahorse metabolic assay were used for analysis of therapy-induced signaling rewiring. BH3 profiling was used to understand the mitochondrial apoptosis mechanisms. Medulloblastoma models were used to expand the importance to other brain cancer. Intracranial xenografts were used to validate the in vivo therapeutic impact of the triplet therapy. ResultsCollectively all tested ten glioblastoma and medulloblastoma cell models were effectively eradicated by the newly discovered triplet therapy combining inhibition of AKT and PDK1-4 kinases with pharmacological PP2A reactivation. Mechanistically, the brain tumor cell selective lethality of the triplet therapy could be explained by its combinatorial effects on therapy-induced signaling rewiring, OXPHOS, and apoptosis priming. The brain-penetrant triplet combination had a significant in vivo efficacy in intracranial glioblastoma and medulloblastoma models. ConclusionThe results confirm highly heterogenous responses of brain cancer cells to mono - and doublet combination therapies targeting phosphorylation-dependent signaling. However, the brain cancer cells cannot escape the triplet therapy targeting of AKT, PDK1-4, and PP2A. The results encourage evaluation of brain tumor PP2A status for design of future kinase inhibitor combination trials. Key PointsO_LIDevelopment of triplet kinase-phosphatase targeting therapy strategy for overcoming therapy tolerance across brain tumor models. C_LIO_LIIdentification of interplay between therapy-induced signaling rewiring, OXPHOS, and BH3 protein-mediated apoptosis priming as a cause for kinase inhibitor tolerance in brain cancers. C_LIO_LIValidation of the results in intracranial in vivo models with orally bioavailable and brain penetrant triplet therapy combination. C_LI Importance of the StudyBased on current genetic knowledge, glioblastoma should be particularly suitable target for kinase inhibitor therapies, However, in glioblastoma alone over 180 clinical trials with kinase inhibitors have failed. In this manuscript, we recapitulate this clinical observation by demonstrating broad tolerance of brain cancer cell models to kinase inhibitors even when combined with reactivation of PP2A. However, we discover that the therapy-induced signaling rewiring, and therapy tolerance, can be overcome by triplet targeting of AKT, PDK1-4 and PP2A. We provide strong evidence for the translatability of the findings by orally dosed brain penetrant triplet therapy combination in intracranial brain cancer models. The results encourage biomarker profiling of brain tumors for their PP2A status for clinical trials with combination of AKT and PDK1-4 inhibitors. Further, the results indicate that rapidly developing PP2A reactivation therapies will constitute an attractive future therapy option for brain tumors when combined with multi-kinase inhibition.

cancer biology↗

bioRxiv2022

Triplet kinase-phosphatase targeting to overcome kinase inhibitor tolerance in brain tumor cells

Therapeutic resistance to kinase inhibitors constitutes a major unresolved clinical challenge in cancer and especially in glioblastoma. Multi-kinase inhibitors may be used for simultaneous targeting of multiple target kinase and thereby potentially overcome kinase inhibitor resistance. However, in most cases identification of the target kinases mediating therapeutic effects of multi-kinase inhibitors has been challenging. To tackle this important problem, we developed an Actionable Targets of Multi-kinase Inhibitors (AToMI) strategy and used it for characterization of glioblastoma target kinases of staurosporine derivatives displaying synergy with protein phosphatase 2A (PP2A) reactivation. AToMI consists of interchangeable modules combining drug-kinase interaction assay, siRNA high-throughput screening, bioinformatics analysis and validation screening with more selective target kinase inhibitors. As a result, AToMI analysis revealed AKT and mitochondrial pyruvate dehydrogenase kinase PDK1 and PDK4 as kinase targets of staurosporine derivatives UCN-01, CEP-701, and K252a that synergized with PP2A activation across heterogeneous glioblastoma cells. Based on these proof-of-principle results we propose that application and further development of AToMI for clinically applicable multi-kinase inhibitors could provide significant benefits in overcoming the challenge of lack of knowledge of target specificity of multi-kinase inhibitors.

cancer biology↗

bioRxiv2021

Binding and activation of serotonergic G-protein coupled receptors by the multimodal antidepressant vortioxetine.

G-protein coupled receptors are important pharmacological targets. Despite substantial progress, important questions still remain concerning the details of activation: how can a ligand act as an agonist in one receptor, but as an antagonist in a homologous receptor, and how can agonists activate a receptor despite lacking polar functional groups able to interact with helix 5? Studying vortioxetine, an important multimodal antidepressant drug, may elucidate both questions. Herein, we present a thorough in silico analysis of vortioxetine binding to 5-HT1A, 5-HT1B, and 5-HT7 receptors and compare to available experimental data. We are able to rationalize the differential mode of action of vortioxetine at different receptors, but also, in the case of the 5-HT1A receptor, we observe the initial steps of activation suggesting that interaction with helix 5 does not necessarily require a hydrogen bond as previously suggested. The results extend our current understanding of agonist and antagonist action at GPCRs.

biophysics↗