Biology subjects

Zimmer, A.

Publications and source records attributed to Zimmer, A..

18 recordsLinked to original sources

bioRxiv2026

Efficient CRISPR/Cas9-mediated homology independent sequence replacement in vivo and non-dividing primary cells

Precise sequence replacement in non-dividing cells remains a major challenge for genome editing. Here we show that REPLACE (Rational end-joining protocol delivering a targeted sequence exchange), a homology-independent CRISPR/Cas9-based replacement strategy, enables exon- and gene-scale substitution in primary cells, in vivo tissues and post-mitotic human cardiomyocytes. REPLACE uses two guide RNAs to excise a defined genomic region and inserts a donor lacking homology arms through non-homologous end joining (NHEJ). In primary mouse hepatocytes, REPLACE mediated exon replacement in 35% of all cells. In adult mouse liver, editing efficiency could be increased to [~]20% when Cas9-sgRNA ribonucleoproteins were delivered via engineered virus-like particles (eVLPs) together with an adeno-associated virus (AAV) donor. REPLACE also supported large-segment replacement, enabling one-step exchange of a [~]27-kb mouse Ace2 interval with the human ACE2 coding region in zygotes, followed by germline transmission and tissue-specific expression. Finally, we applied REPLACE to a disease-relevant mutation that is not readily addressable by base editing and was poorly corrected by prime editing in post-mitotic cardiomyocytes. At the LMNA locus, REPLACE corrected the K117fs frameshift mutation in patient-derived post-mitotic cardiomyocytes with precise exon replacement and restored Lamin A/C protein expression and nuclear lamina localization. These findings establish REPLACE as a versatile platform for homology-independent sequence replacement and as a complementary approach for genetic correction in settings where homology-directed repair (HDR), base editing (BE) or prime editing (PE) are inefficient or not applicable.

genomics↗

bioRxiv2025

Industrialization drives convergent microbial and physiological shifts in the human metaorganism

Understanding how host lifestyle and industrialization shape the human gut microbiome and intestinal physiology requires multimodal analyses across diverse global host contexts. Here, we generate multivariate data from the Global Microbiome Conservancy cohort, including gut microbiome, IgA-sequencing, host genotyping, diet, lifestyle and fecal biomarker profiles, to investigate host-microbiome interactions across gradients of industrialization and geography. We show that industrialization is associated with homogenized microbial compositions, reduced microbial diversity, and lower community stability, independent of host confounders. We further show that industrialization is linked to elevated markers of gut stress, increased IgA secretion, and altered patterns of IgA-bacteria interactions. Finally, we show that microbiome-based disease predictors trained on industrialized populations lose accuracy in less industrialized cohorts, highlighting limited cross-population transferability. Together, our results suggest profound restructuring of host-microbiome interactions due to industrialized lifestyles, and emphasize the need for inclusive, globally representative data to improve translational microbiome applications across diverse human populations.

microbiology↗

bioRxiv2024

FAAH Inhibition as a Path to Mitigate Alzheimer's Disease Progression in Mouse Models of Amyloidosis

The endocannabinoid N-arachidonoylethanolamine (AEA) is a pro-homeostatic bioactive lipid known for its anti-inflammatory, anti-oxidative, immunomodulatory, and neuroprotective properties, which may contrast/mitigate Alzheimers disease (AD) pathology. This study explores the therapeutic potential of targeting fatty acid amide hydrolase (FAAH), the major enzyme degrading AEA, in mouse models of amyloidosis APP/PS1 and Tg2576. Enhancing AEA signaling by genetic deletion of FAAH delayed cognitive deficits in APP/PS1 mice and improved cognitive symptoms in 12-month-old AD-like mice. Chronic pharmacological FAAH inhibition fully reverted neurocognitive decline, attenuated neuroinflammation, and promoted neuroprotective mechanisms in Tg2576 mice. Additionally, pharmacological FAAH inactivation robustly suppressed {beta}-amyloid production and accumulation, associated with decreased expression of {beta}-site amyloid precursor protein cleaving enzyme 1 (BACE1), possibly through a cannabinoid receptor 1-dependent epigenetic mechanism. These findings improve our understanding of AEA signaling in AD pathogenesis, and provide proof-of-concept that selective targeting of FAAH activity could be a promising therapeutic strategy against AD.

biochemistry↗

bioRxiv2024

Acute high-fat high-sugar diet rapidly increases blood-brain barrier permeability in mice

BackgroundThe blood-brain barrier (BBB) maintains brain homeostasis by protecting the brain from pathological stimuli and controlling the entry of physiological substances from the periphery. Consequently, alterations in BBB permeability may pose a threat to brain health. Long-term consumption of a high-fat high-sugar/Western diet (HFD) is known to induce BBB dysfunction. However, nothing is known about the immediate effects of acute HFD consumption on the BBB. MethodsAfter consumption of either HFD or standard chow, mice were injected into the tail vein with fluorescent tracers of different sizes, including the drug doxorubicin. Individual brain regions were homogenized and analyzed for tracer extravasation using spectrophotometry. Localized tracer leakage over time in the somatosensory cortex was studied using high-resolution in vivo 2-photon microscopy. ResultsWe demonstrate region-specific BBB leakage already after 1 hour of HFD for low- and high-molecular-weight tracers. Acute HFD also significantly increased BBB permeability to the anticancer drug doxorubicin. ConclusionThese previously unknown effects of acute HFD may have direct and drastic implications for the clinical use of drugs depending on the dietary habits of the patient. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=110 SRC="FIGDIR/small/589405v1_ufig1.gif" ALT="Figure 1"> View larger version (39K): org.highwire.dtl.DTLVardef@731c74org.highwire.dtl.DTLVardef@1512efcorg.highwire.dtl.DTLVardef@e3cc19org.highwire.dtl.DTLVardef@16c9248_HPS_FORMAT_FIGEXP M_FIG C_FIG

neuroscience↗

bioRxiv2024

Reduction in activity and abundance of mitochondrial electron transport chain supercomplexes in pulmonary hypertension-induced right ventricular dysfunction

Pulmonary hypertension (PH) results in RV hypertrophy, fibrosis and dysfunction resulting in RV failure which is associated with impaired RV metabolism and mitochondrial respiration. Mitochondrial supercomplexes (mSC) are assemblies of multiple electron transport chain (ETC) complexes that consist of physically associated complex I, III and IV that may enhance respiration and lower ROS generation. The goal of this study was to determine if mSCs are reduced in RV dysfunction associated with PH. We induced PH in Sprague-Dawley rats by Sugen/Hypoxia (3 weeks) followed by normoxia (4 weeks). Control and PH rats were subjected to echocardiography, blue and clear native-PAGE to assess mSC abundance and activity, and cardiomyocyte isolation to assess mitochondrial reactive oxygen species (ROS). mSC formation was also assessed in explanted human hearts with and without RV dysfunction. RV activity of CI and CIV and abundance of CI, CIII and CIV in mitochondrial mSCs was severely reduced in PH rats compared to control. There were no differences in total CI or CIV activity or abundance in smaller ETC assemblies. There were no changes in both RV and LV of expression of representative ETC complex subunits. PAT, TAPSE and RV Wall thickness significantly correlated with CIV and CI activity in mSC, but not total CI and CIV activity in the RV. Consistent with reduced mSC activity, isolated PH RV myocytes had increased mitochondrial ROS generation compared to control. Reduced mSC activity was also demonstrated in explanted human RV tissue from patients undergoing cardiac transplant with RV dysfunction. The right atrial pressure/pulmonary capillary wedge pressure ratio (RAP/PCWP, an indicator of RV dysfunction) negatively correlated with RV mSC activity level. In conclusion, reduced assembly and activity of mitochondrial mSC is correlated with RV dysfunction in PH rats and humans with RV dysfunction.

pathology↗

bioRxiv2023

Tracking the progression of Alzheimer's disease with peripheral blood monocytes

BackgroundAlzheimers disease (AD) is the most common form of dementia with the symptoms gradually worsening over the years. However, the driving pathological processes occur well before the appearance of symptoms. AD patients display signs of systemic inflammation, suggesting that it could precede the well-established AD hallmarks. We recently showed that the innate immune response in the form of monocyte activation is detectable at the pre-clinical stage. ObjectivesOur goal here is to characterize changes of gene expression in peripheral blood monocytes from patients at different stages of AD progression and validate potential biomarkers for a better prognosis and diagnosis of AD clinical spectrum. ResultsWe performed a whole transcriptome analysis on monocytes purified from healthy subjects, Mild Cognitive Impairment (MCI) and AD patients, and established the list of genes differentially expressed in monocytes during the disease evolution. We observed that, in the top 500 genes differentially expressed, a majority of these genes were upregulated (65%) during AD progression. These genes are mainly involved in chemokine/cytokine-mediated signaling pathways. We further confirmed several biomarkers by quantitative PCR and immunoblotting and showed that they are often deregulated at pre-clinical stages of the disease (MCI stage), supporting the hyperactivation of monocytes in MCI patients. PerspectivesOur findings provide evidence that the pre-clinical stage of AD can be detected in monocytes using a specific set of biomarkers, highlighting the importance to study the early innate immune response in AD. Our results open the possibility to use these biomarkers with different diagnostic methodologies to better predict and efficiently treat AD.

neuroscience↗

bioRxiv2022

Genomic and Transcriptomic Profiling of Phoenix Colonies

Pseudomonas aeruginosa is a Gram-negative bacterium responsible for numerous human infections. Previously, novel antibiotic tolerant variants known as phoenix colonies as well as variants similar to viable but non-culturable (VBNC) colonies were identified in response to high concentrations of aminoglycosides. In this study, the mechanisms behind phoenix colony and VBNC-like colony emergence were further explored using both whole genome sequencing and RNA sequencing. Phoenix colonies were found to have a single nucleotide polymorphism (SNP) in the PA4673 gene, which is predicted to encode a GTP-binding protein. No SNPs were identified within VBNC-like colonies compared to the founder population. RNA sequencing did not detect change in expression of PA4673 but revealed multiple differentially expressed genes that may play a role in phoenix colony emergence. One of these differentially expressed genes, PA3626, encodes for a tRNA pseudouridine synthase which when knocked out led to a complete lack of phoenix colonies. Although not immediately clear whether the identified genes in this study may have interactions which have not yet been recognized, they may contribute to the understanding of how phoenix colonies are able to emerge and survive in the presence of antibiotic exposure.

microbiology↗

bioRxiv2021

An astrocytic signaling loop for frequency-dependent control of dendritic integration and spatial learning

Dendrites of hippocampal CA1 pyramidal cells amplify clustered glutamatergic input by activation of voltage-gated sodium channels and N-methyl-D-aspartate receptors (NMDARs). NMDAR activity depends on the presence of NMDAR co-agonists such as D-serine, but how co-agonists influence dendritic integration is not well understood. Using combinations of whole-cell patch clamp, iontophoretic glutamate application, two-photon excitation fluorescence microscopy and glutamate uncaging we found that exogenous D-serine reduces the threshold of dendritic spikes and increases their amplitude. Triggering an astrocytic mechanism controlling endogenous D-serine supply via endocannabinoid receptors (CBRs) also increased dendritic spiking. Unexpectedly, this pathway was activated by pyramidal cell activity primarily in the theta range, which required HCN channels and astrocytic CB1Rs. Therefore, astrocytes close a positive and frequency-dependent feedback loop between pyramidal cell activity and their integration of dendritic input. Its disruption led to an impairment of spatial memory, which demonstrates its behavioral relevance.

neuroscience↗

bioRxiv2021

Autocrine regulation of adult neurogenesis by the endocannabinoid 2-arachidonoylglycerol (2-AG)

The endocannabinoid system modulates adult hippocampal neurogenesis by promoting the proliferation and survival of neural stem and progenitor cells (NSPCs). Specifically, deleting cannabinoid receptors on NSPCs or the constitutive deletion of the endocannabinoid 2-arachidonoylglycerol (2-AG) producing enzyme diacylglycerol lipase alpha (DAGLa) disrupts adult neurogenesis. However, it is not known which cells are the producers of 2-AG relevant to neurogenesis. In this paper, we investigated the cellular source of endocannabinoids in the subgranular zone (SGZ) of the hippocampus, an important neurogenic niche. For this purpose, we used two complementary Cre-deleter mouse strains to delete DAGLa either in neurons or astroglia and NSPCs. Surprisingly, neurogenesis was not altered in mice with a deletion of Dagla in neurons (Syn-Dagla KO), although they are the main source for the endocannabinoids in the brain. In contrast, mice with a specific inducible deletion of Dagla in NPSCs and astrocytes (GLAST-CreERT2-Dagla KO) showed a strongly impaired neurogenesis with significantly reduced proliferation and survival of newborn cells. These results identify Dagla in NSPCs in the SGZ of dentate gyrus or in astrocytes, as the cellular source for 2-AG in adult hippocampal neurogenesis. In summary, 2-AG produced by progenitor cells or astrocytes in the SGZ regulates adult hippocampal neurogenesis. SummaryDAGLa in neuronal progenitor cells in the SGZ of dentate gyrus is identified as the cellular source for 2-AG in adult hippocampal neurogenesis.

neuroscience↗

bioRxiv2021

Adherent and suspension baby hamster kidney cells have a different cytoskeleton and surface receptor repertoire

Animal cell culture, with single cells growing in suspension, ideally in a chemically defined environment, is a mainstay of biopharmaceutical production. The synthetic environment lacks exogenous growth factors and usually requires a time-consuming adaptation process to select cell clones that proliferate in suspension to high cell numbers. The molecular mechanisms that facilitate the adaptation and that take place inside the cell are largely unknown. Especially for cell lines that are used for virus antigen production such as baby hamster kidney (BHK) cells, the restriction of virus growth through the evolution of undesired cell characteristics is highly unwanted. The comparison between adherently growing BHK cells and suspension cells with different susceptibility to foot-and-mouth disease virus revealed differences in the expression of cellular receptors such as integrins and heparan sulfates, and in the organization of the actin cytoskeleton. Transcriptome analyses and growth kinetics demonstrated the diversity of BHK cell clones and confirmed the importance of well-characterized parental cell clones and mindful screening to make sure that essential cellular features do not get lost during adaptation.

cell biology↗

bioRxiv2020

Protective role of neuronal and lymphoid cannabinoid CB2 receptors in neuropathic pain

Cannabinoid CB2 receptor (CB2r) agonists are potential painkillers void of psychotropic effects. Peripheral immune cells, neurons and glia express CB2r, however the involvement of CB2r from these cells in neuropathic pain remains unresolved. We explored spontaneous neuropathic pain through on-demand self-administration of the selective CB2r agonist JWH133 in wild-type and knockout mice lacking CB2r in neurons, monocytes or constitutively. Operant self-administration reflected drug-taking to alleviate spontaneous pain, nociceptive and affective manifestations. While constitutive deletion of CB2r disrupted JWH133-taking behavior, this behavior was not modified in monocyte-specific CB2r knockouts and was increased in mice defective in neuronal CB2r knockouts suggestive of increased spontaneous pain. Interestingly, CB2r-positive lymphocytes infiltrated the injured nerve and possible CB2r transfer from immune cells to neurons was found. Lymphocyte CB2r depletion also exacerbated JWH133 self-administration and inhibited antinociception. This work identifies a simultaneous activity of neuronal and lymphoid CB2r that protects against spontaneous and evoked neuropathic pain.

physiology↗

bioRxiv2020

Actinobacillus utilizes a binding-protein dependent ABC transporter to acquire Vitamin B6

Bacteria require high efficiency uptake systems to survive and proliferate in nutrient limiting environments, such as those found in the host. The ABC transporters at the bacterial plasma membrane provide a mechanism for transport of many substrates. We recently demonstrated that an AfuABC operon, previously annotated as encoding a ferrous iron uptake system, is in fact a cyclic hexose/heptose-phosphate transporter with high selectivity and specificity for these metabolites. In this study, we examine a second operon containing a periplasmic binding protein discovered in Actinobacillus for its potential role in nutrient acquisition. Using electron density obtained from the crystal structure of the periplasmic binding protein we modeled a pyridoxal-5-phosphate (P5P/PLP/Vitamin B6) ligand into the atomic resolution electron density map. The identity of the Vitamin B6 bound to this periplasmic binding protein was verified by isothermal titration calorimetry, microscale thermophoresis, and mass spectrometry, leading us to name the protein P5PA and the operon P5PAB. To illustrate the functional utility of this uptake system, we introduced the P5PAB operon from A. pleuropneumoniae into an E. coli K-12 strain that was devoid of a key enzyme required for Vitamin B6 synthesis. The growth of this strain at low levels of Vitamin B6 supports the role of this newly identify operon in Vitamin B6 uptake.

biochemistry↗

bioRxiv2020

Gut Microbiome Pattern Reflects Healthy Aging and Predicts Extended Survival in Humans

The gut microbiome has important effects on human health, yet its importance in human aging remains unclear. Using two independent cohorts comprising 4582 individuals across the adult lifespan we demonstrate that, starting in mid-to-late adulthood, gut microbiomes become increasingly unique with age. This uniqueness pattern is strongly associated with gut microbial amino acid derivatives circulating within the bloodstream, many of which have been previously identified as longevity biomarkers. At the latest stages of human life, two distinct patterns emerge wherein individuals in good health show continued microbial drift toward a unique compositional state, while the same drift is absent in individuals who perform worse on a number of validated health measures. The identified healthy aging pattern is characterized by an overall depletion of core genera found across most humans - primarily a depletion in the nearly ubiquitous genus Bacteroides. Consistently, retaining a high Bacteroides dominance into extreme age, or, equivalently, having a low gut microbiome uniqueness score, predicts decreased survival in a four-year follow-up. Our comprehensive analysis identifies the gut microbiome as a novel component of healthy aging, with important implications for the worlds growing older population.

systems biology↗

bioRxiv2020

Immunohistochemical Characterization of Phosphorylated Ubiquitin in the Mouse Hippocampus

Mitochondrial autophagy (mitophagy) is an essential and evolutionarily conserved process that maintains mitochondrial integrity via the removal of damaged or superfluous mitochondria in eukaryotic cells. Phosphatase and tensin homolog (PTEN)-induced putative kinase 1 (PINK1) and Parkin promote mitophagy and function in a common signaling pathway. PINK1-mediated ubiquitin phosphorylation at Serine 65 (Ser65-pUb) is a key event in the efficient execution of PINK1/Parkin-dependent mitophagy. However, few studies have used immunohistochemistry to analyze Ser65-pUb in the mouse. Here, we examined the immunohistochemical characteristics of Ser65-pUb in the mouse hippocampus. Some hippocampal cells were Ser65-pUb positive, whereas the remaining cells expressed no or low levels of Ser65-pUb. PINK1 deficiency resulted in a decrease in the density of Ser65-pUb-positive cells, consistent with a previous hypothesis based on in vitro research. Interestingly, Ser65-pUb-positive cells were detected in hippocampi lacking PINK1 expression. The CA3 pyramidal cell layer and the dentate gyrus (DG) granule cell layer exhibited significant reductions in the density of Ser65-pUb-positive cells in PINK1-deficient mice. Moreover, Ser65-pUb immunoreactivity colocalized predominantly with neuronal markers. These findings suggest that Ser65-pUb may serve as a biomarker of in situ PINK1 signaling in the mouse hippocampus; however, the results should be interpreted with caution, as PINK1 deficiency downregulated Ser65-pUb only partially.

neuroscience↗

bioRxiv2018

Scale-invariant geometric data analysis (SIGDA) provides robust, detailed visualizations of human ancestry specific to individuals and populations

Scale invariance is a common property of physical laws and a key concept in perspective drawing, which aims to provide a meaningful two-dimensional representation of a more complex, three-dimensional scene. Here we describe Scale Invariant Geometric Data Analysis (SIGDA), a new, general exploratory data analysis (EDA) method based on normalization of data to scale invariance. We discuss similarities and differences between SIGDA and two widely-used EDA methods, Correspondence Analysis (CA) and Principal Components Analysis (PCA). We then illustrate SIGDAs ability to analyze and visualize population structure relationships within the data that inspired its development: genetic marker data, in which context PCA is considered a standard method. We show that SIGDA provides significant advantages over PCA of the same data, including: (a) robust detection and separation of a larger number of population axes, leading to (b) better separation of annotated populations; (c) separation of an independent allele frequency axis interpretable as a proxy for allele age, (d) visualization of marker flow between populations (population history), and (d) robust detection and visualization of relationships between closely-related individuals and among family groups. Although this illustration focuses on a specific task, SIGDA is a general-purpose EDA method and derives its advantages from its novel approach to fundamental issues in data analysis, rather than clever sampling or other task-specific methodology.\n\nOne Sentence SummaryWe illustrate the advantages of Scale Invariant Geometric Data Analysis (SIGDA), a new exploratory data analysis method similar to PCA, by applying SIGDA to derive detailed, robust visualizations of the complex history of human population structure from a large sample of single nucleotide variants.

bioinformatics↗

bioRxiv2018

Universal cancer tasks, evolutionary tradeoffs, and the functions of driver mutations

Recent advances have led to an appreciation of the vast molecular diversity of cancer. Detailed data has enabled powerful methods to sort tumors into groups with benefits for prognosis and treatment. We are still missing, however, a general theoretical framework to understand the diversity of tumor gene-expression and mutations. To address this, we present a framework based on multi-task evolution theory, using the fact that tumors evolve in the body, and that tumors are faced with multiple tasks that contribute to their fitness. In accordance with the theory, we find that tradeoff between tasks constrains tumor gene-expression to a continuum bounded by a polyhedron. The vertices of the polyhedron are gene-expression profiles each specializing in one task, allowing the tasks to be identified. We find five universal cancer tasks across tissue-types: cell-division, biomass & energy, lipogenesis, immune-interaction and invasion & tissue remodeling. Tumors whose gene-expression lies close to a vertex are task specialists. We find evidence that such specialists are more sensitive to drugs that interfere with this task. We find that driver mutations, but not passenger mutations, tune gene-expression towards specialization in specific tasks. This approach can integrate additional types of molecular data into a theoretically-based framework for understanding tumor diversity.

cancer biology↗

bioRxiv2017

A unified web platform for network-based analyses of genomic data

Functional genomics networks are widely used to identify unexpected pathway relationships in large genomic datasets. However, it is challenging to quantitatively compare the signal-to-noise ratio of different networks, the biology they describe, and to identify the optimal network to interpret a particular genetic dataset. Via GeNets users can train a machine-learning model (Quack) to make such comparisons; and they can execute, store, and share analyses of genetic and RNA sequencing datasets.

genomics↗

bioRxiv2017

Activation of Somatostatin Inhibitory Neurons by Lypd6-nAChRα2 System Restores Juvenile-like Plasticity in Adult Visual Cortex

The limitation of plasticity in the adult brain impedes functional recovery later in life from brain injury or disease. This pressing clinical issue may be resolved by enhancing plasticity in the adult brain. One strategy for triggering robust plasticity in adulthood is to reproduce one of the hallmark physiological events of experience-dependent plasticity observed during the juvenile critical period - rapidly reduce the activity of parvalbumin (PV)-expressing interneurons and disinhibit local excitatory neurons. This may be achieved through enhancement of local inhibitory inputs, particularly those of somatostatin (SST)-expressing interneurons. However, to date the means for manipulating SST interneurons for enhancing cortical plasticity in the adult brain are not known. We show that SST interneuron-selective overexpression of Lypd6, an endogenous nicotinic signaling modulator, enhances ocular dominance plasticity in the adult primary visual cortex (V1). Lypd6 overexpression mediates a rapid experience-dependent increase in the visually evoked activity of SST interneurons as well as a simultaneous reduction in PV interneuron activity and disinhibition of excitatory neurons. Recapitulating this transient activation of SST interneurons using chemogenetics similarly enhanced V1 plasticity. Notably, we show that SST-selective Lypd6 overexpression restores visual acuity in amblyopic mice that underwent early long-term monocular deprivation. Our data in both male and female mice reveal selective modulation of SST interneurons and a putative downstream circuit mechanism as an effective method for enhancing experience-dependent cortical plasticity as well as functional recovery in adulthood. Significance StatementThe decline of cortical plasticity after closure of juvenile critical period consolidates neural circuits and behavior, but this limits functional recovery from brain diseases and dysfunctions in later life. Here we show that activation of cortical SST interneurons by Lypd6, an endogenous modulator of nicotinic acetylcholine receptors (nAChRs), enhances experience-dependent plasticity and recovery from amblyopia in adulthood. This manipulation triggers rapid reduction of PV interneuron activity and disinhibition of excitatory neurons, which are known hallmarks of cortical plasticity during juvenile critical periods. Our study demonstrates modulation of SST interneurons by Lypd6 to achieve robust levels of cortical plasticity in the adult brain and may provide promising targets for restoring brain function in the event of brain trauma or disease.

neuroscience↗