Biology subjects

Schulze, J.

Publications and source records attributed to Schulze, J..

9 recordsLinked to original sources

bioRxiv2026

AdamTS-B protease is required for morphogenesis of the Drosophila respiratory system

Epithelial tube morphogenesis is critical for the function of many organs. Basement membranes underlie epithelia and their remodeling is a key step to reach the correct size and shape. Key regulators that mediate basement membrane remodeling for tube elongation and branching remain largely unknown. We analyze the expression and function of AdamTS-B, a matrix metalloprotease, in the respiratory system of Drosophila. Here we show, that AdamTS-B is expressed early in tracheal development during placode formation. We generated a mutant line of AdamTS-B, which is lethal. Analysis of trachea morphogenesis in this AdamTS-B mutant reveal a function in tube elongation and cell migration. Our results suggest that AdamTS-B control BM remodeling required for organ shape.

developmental biology↗

bioRxiv2025

Non-destructive radiocarbon dating of bone

Since the 1950s, radiocarbon measurements have anchored archaeological chronologies dating back to 50,000 years, with bone collagen being a commonly dated material. Despite advances in collagen extraction protocols, the process remains destructive, requiring sampling by sawing, drilling or crushing of dry bone, that can damage or destroy physical features and archaeological evidence, and often consumes the entire specimen. While non-destructive approaches have recently been applied in ancient genomics and palaeoproteomics, no equivalent approach has been established for radiocarbon dating of bone. Here, we outline a non-destructive collagen extraction workflow that avoids invasive sampling (cutting or drilling) and produces no visible damage to the bone. By heating the bone in hot water for several hours, collagen is solubilized, and the resulting collagenous solution can be purified and AMS dated. Here we show that the amino acid composition, C/N atomic ratios, {delta}13C and {delta}15N values of the hot-water-extracted collagen are comparable to collagen isolated from the same bones using classic decalcification methodologies. The hot water extraction method was tested on various bones, ranging from the Bronze Age to the Middle and Upper Paleolithic periods, which had been dated previously using routine destructive methods that involved acid demineralization. Our results show that non-destructive collagen extraction, coupled with an additional purification step e.g., ultrafiltration, yields identical radiocarbon ages to those obtained via the routine destructive methods.

paleontology↗

bioRxiv2024

varVAMP: automated pan-specific primer design for tiled full genome sequencing and qPCR of highly diverse viral pathogens.

Time- and cost-saving surveillance of viral pathogens is achieved by tiled sequencing in which a viral genome is amplified in overlapping PCR amplicons and qPCR. However, designing pan-specific primers for viral pathogens that have high genomic variability represents a major challenge. Here, we present a bioinformatics command-line tool, called varVAMP (variable virus amplicons). It relies on multiple sequence alignments of highly variable virus sequences and enables automatic pan-specific primer design for qPCR or tiled amplicon whole genome sequencing. The varVAMP software guarantees pan-specificity by two means: it designs primers in regions with minimal variability and introduces degenerate nucleotides into primer sequences to compensate for common sequence variations. We demonstrate varVAMPs utility by designing and evaluating novel pan-specific primer schemes suitable for sequencing the genomes of SARS-CoV-2, Hepatitis E virus, rat Hepatitis E virus, Hepatitis A virus, Borna-disease-virus-1, and Poliovirus. Moreover, we established highly sensitive and specific Poliovirus qPCR assays that could potentially simplify current Poliovirus surveillance. Importantly, wet-lab and bioinformatic techniques established for SARS-CoV-2 tiled amplicon sequencing were readily transferable to these new primer schemes and will allow sequencing laboratories to extend their established methodology to other human pathogens.

bioinformatics↗

bioRxiv2023

Neural Extracellular Matrix Remodeling Signatures in Genetic and Acquired Mouse Models of Epilepsy

Epilepsies are multifaceted neurological disorders characterized by abnormal brain activity, e.g., caused by imbalanced synaptic excitation and inhibition. The neural extracellular matrix (ECM) is dynamically modulated by physiological and pathophysiological activity and critically involved in controlling the brains excitability. We used different epilepsy models, i.e. mice lacking the presynaptic scaffolding protein Bassoon at excitatory, inhibitory or all synapse types as genetic models for rapidly generalizing early-onset epilepsy, and intra-hippocampal kainate injection, a model for acquired temporal lobe epilepsy, to study the relationship between epileptic seizures and ECM composition. Electroencephalogram recordings revealed Bassoon deletion at excitatory or inhibitory synapses having diverse effects on epilepsy-related phenotypes. While constitutive Bsn mutants and GABAergic neuron-specific knockouts (BsnDlx5/6cKO) displayed severe epilepsy with more and stronger seizures than kainate-injected animals, mutants lacking Bassoon solely in excitatory forebrain neurons (BsnEmx1cKO) showed only mild impairments. By semiquantitative immunoblotting and immunohistochemistry we show model-specific patterns of neural ECM remodeling, and we also demonstrate significant upregulation of the ECM receptor CD44 in null and BsnDlx5/6cKO mutants. ECM-associated WFA-binding chondroitin sulfates were strongly augmented in seizure models. Strikingly, Brevican, Neurocan, Aggrecan and link protein Hapln1 levels reliably predicted seizure properties across models, suggesting a link between ECM state and epileptic phenotype.

neuroscience↗

bioRxiv2022

Loss of FOXP3 function causes expansion of two pools of autoreactive T cells in patients with IPEX syndrome.

The monogenic autoimmune disease Immunedysregulation polyendocrynopathy entheropathy X-linked syndrome (IPEX) has elucidated the essential function of the transcription factor FOXP3 and of thymic-derived regulatory T (Treg) cells in controlling autoimmunity. However, the presence of autoreactive T cells in IPEX remains undetermined, thus representing a crucial gap in understanding the origin of autoimmunity in a FOXP3 deficient immune system. Combining epigenetic analysis as a lineage marker of Treg identity and TCR sequencing to assess the self-reactive clones, we showed that IPEX patients have two pools of expanded autoreactive T cells. The first originates from the expansion of autoreactive effector T cells (Teff), likely due to loss of Treg suppressive function since it is absent in carrier mothers, in whom Treg cells are functional. The second pool originates, unexpectedly, from Treg cells which lose their phenotypic markers, including CD25 and FOXP3. We call these loss of identity Treg cells and show that they are i) suppressed by healthy donor Treg in a patient post hematopoietic transplantation despite low donor chimerism, and ii) not detectable in patients with Autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy syndrome (APECED), a monogenic autoimmune disease of thymic origin. Moreover, we demonstrate that FOXP3 knock-out in Treg cells leads to increased Treg expansion and production of Th17 and Th2 cytokines, known to be increased in IPEX patients. These results suggest that the loss of identity Treg cells could directly contribute to immune dysregulation in IPEX. Collectively, we provide a better understanding of autoimmunity and novel ways to monitor the effects of Treg cell therapies in IPEX disease or other autoimmune diseases. One Sentence SummaryMutations of FOXP3 gene in humans cause expansion of autoreactive T cells originating from both effector T cells and regulatory T cells which gain effector function.

immunology↗

bioRxiv2021

Antiviral RNAi response against the insect-specific Agua Salud alphavirus

Arboviruses transmitted by mosquitoes are responsible for the death of millions of people each year. In addition to arboviruses, many insect-specific viruses (ISVs) have been discovered in mosquitoes in the last decade. ISVs, in contrast to arboviruses transmitted by mosquitoes to vertebrates, cannot replicate in vertebrate cells even when they are evolutionarily closely related to arboviruses. The alphavirus genus includes many arboviruses, although only a few ISVs have been discovered from this genus so far. Here, we investigate the interactions of a recently isolated insect-specific alphavirus, Agua-Salud alphavirus (ASALV), with its mosquito host. RNAi is one of the essential antiviral responses against arboviruses, although there is little knowledge on the interactions of RNAi with ISVs. Through knock-down of transcripts of the different key RNAi pathway (siRNA, miRNA and piRNA) proteins, we show the antiviral role of Ago2 (siRNA), Ago1 (miRNA), and Piwi4 proteins against ASALV in Aedes aegypti derived cells. ASALV replication increased in Dicer2 and Ago2 knock-out cells, confirming the antiviral role of the siRNA pathway. In infected cells, mainly ASALV-specific siRNAs are produced while piRNAs, with the characteristic nucleotide bias resulting from ping-pong amplification, are only produced in Dicer2 knock-out cells. Taken together, ASALV interactions with the mosquito RNAi response differs from arthropod-borne alphaviruses in some aspects, although they also share some commonalities. Further research is needed to understand whether the identified differences can be generalised to other insect-specific alphaviruses.

microbiology↗

bioRxiv2021

Post-entry, spike-dependent replication advantage of B.1.1.7 and B.1.617.2 over B.1 SARS-CoV-2 in an ACE2-deficient human lung cell line

Epidemiological data demonstrate that SARS-CoV-2 variants of concern (VOC) B.1.1.7 and B.1.617.2 are more transmissible and infections are associated with a higher mortality than non-VOC virus infections. Phenotypic properties underlying their enhanced spread in the human population remain unknown. B.1.1.7 virus isolates displayed inferior or equivalent spread in most cell lines and primary cells compared to an ancestral B.1 SARS-CoV-2, and were outcompeted by the latter. Lower infectivity and delayed entry kinetics of B.1.1.7 viruses were accompanied by inefficient proteolytic processing of spike. B.1.1.7 viruses failed to escape from neutralizing antibodies, but slightly dampened induction of innate immunity. The bronchial cell line NCI-H1299 supported 24- and 595-fold increased growth of B.1.1.7 and B.1.617.2 viruses, respectively, in the absence of detectable ACE2 expression and in a spike-determined fashion. Superior spread in NCI-H1299 cells suggests that VOCs employ a distinct set of cellular cofactors that may be unavailable in standard cell lines.

microbiology↗

bioRxiv2020

Split-HaloTag Imaging Assay for in vivo 3D-Microscopy and Subdiffractional Analyses of Protein-Protein Interactions

An ever-increasing number of intracellular multi-protein networks have been identified in plant cells. Split-GFP based protein-protein interaction assays combine the advantages of in vivo interaction studies in a native environment with additional visualisation of protein complex localisation. Due to its simple protocols, it has become one of the most frequently used methods. However, standard fluorescent proteins entail several drawbacks for sophisticated microscopy. With the HaloTag(R) system, these drawbacks can be overcome as this reporter forms covalent irreversible bonds with synthetic photostable fluorescent ligands. Dyes can be used in adjustable concentrations and are suitable for advanced microscopy methods. Therefore, we established the Split-HaloTag(R) imaging assay in plants which is based on the reconstitution of a functional HaloTag(R) protein upon protein-protein interaction and subsequent covalent binding of an added fluorescent ligand. Its suitability and robustness were demonstrated using well-characterised interactions as an example for protein-protein interaction at cellular structures: the molybdenum cofactor biosynthesis complex anchoring to filamentous actin. Additionally, a specific interaction was visualised with subdiffractional polarisation microscopy in a more distinctive manner as example for sophisticated imaging. Split-GFP and Split-HaloTag(R) can complement one another as Split-HaloTag(R) represents an alternative option and an addition to the large toolbox of in vivo methods. Therefore, this promising new Split-HaloTag(R) imaging assay provides a unique and sensitive approach for more detailed characterization of protein-protein interaction with specific microscopic techniques such as 3D-imaging, single molecule tracking and super-resolution microscopy.

plant biology↗

bioRxiv2018

A specific, glycomimetic Langerin ligand for human Langerhans cell targeting

Langerhans cells are a subset of dendritic cells residing in the epidermis of the human skin. As such, they are key mediators of immune regulation and have emerged as prime targets for novel transcutaneous cancer vaccines. Importantly, the induction of protective T cell immunity by these vaccines requires the efficient and specific delivery of both tumor-associated antigens and adjuvants. Langerhans cells uniquely express Langerin (CD207), an endocytic C-type lectin receptor. Here, we report the discovery of a specific, glycomimetic Langerin ligand employing a heparin-inspired design strategy that integrated NMR spectroscopy and molecular docking. The conjugation of these glycomimetics to liposomes enabled the specific and efficient targeting of Langerhans cells in the human skin. This delivery platform provides superior versatility and scalability over antibody-based approaches and thus addresses current limitations of dendritic cell-based immunotherapies.

immunology↗