Biology subjects

Hirsch, J.

Publications and source records attributed to Hirsch, J..

12 recordsLinked to original sources

bioRxiv2026

Tie2 signaling in the tumor microenvironment orchestrates breast cancer cell dissemination through TMEM doorways

During breast cancer metastasis, tumor cells migrate toward intratumoral blood vessels and intravasate through stable structures known as TMEM (Tumor Microenvironment of Metastasis) doorways. TMEM doorways, composed of a Mena-expressing tumor cell, a Tie2hi/VEGFhi macrophage, and an endothelial cell, are clinically validated prognostic markers of distant metastasis in breast cancer and represent the exclusive sites of tumor cell intravasation. We previously demonstrated that Tie2 signaling is essential for TMEM doorway function and tumor cell intravasation. In this study, we investigated how Tie2 signaling promotes tumor cell intravasation and metastasis. Because all three TMEM doorway-associated cell types can express Tie2, we sought to determine which of these cells contribute to the Tie2 signaling-dependent vascular opening at TMEM doorways and tumor cell dissemination. We found that endothelial cells associated with TMEM doorways secrete Ang2, which stimulates VEGF-A expression in Tie2hi macrophages. Elevated VEGF-A levels at TMEM doorways increase vascular permeability, facilitating tumor cell entry into the bloodstream. Using tissue staining and line-scan analysis of Tie2 and lineage markers in human and mouse breast cancer models, we observed Tie2 expression in macrophages, tumor cells, and endothelial cells. To assess functional contributions, we selectively disrupted Tie2 in macrophages, endothelial cells, and cancer cells using CRISPR-Cas9 and RNAi approaches and tested in which of these cell-knockouts of Tie2 expression affected transendothelial migration in vitro. Macrophage-specific Tie2 deletion had the greatest impact on tumor cell intravasation. To confirm this finding in vivo, we generated a mouse model with inducible, macrophage-specific Tie2 knockout. Acute, targeted loss of Tie2 specifically in macrophages significantly reduced TMEM doorway associated vascular opening and tumor cell intravasation. Together, these findings establish macrophage Tie2 signaling as a critical driver of TMEM doorway-mediated vascular permeability and metastatic dissemination in breast cancer.

cancer biology↗

bioRxiv2025

Live-cell quantitative monitoring reveals distinct, high-affinity Gβγ regulations of GIRK2 and GIRK1/2 channels

Gi/o protein-coupled receptors (GPCRs) inhibit cardiac and neuronal excitability via G protein-activated K+ channels (GIRK), assembled by combinations of GIRK1 - GIRK4 subunits. GIRKs are activated by direct binding of the G{beta}{gamma} dimer of inhibitory Gi/o proteins. However, key aspects of this textbook signaling pathway remain debated. Recent studies suggested no Gi/o-GIRK pre-coupling and low (>250 {micro}M) G{beta}{gamma}-GIRK interaction affinity, contradicting earlier sub-{micro}M estimates and implying low signaling efficiency. We show that G{gamma} prenylation, which mediates G{beta}{gamma} membrane attachment required for GIRK activation, also contributes to the G{beta}{gamma}-GIRK interaction, explaining the poor affinity obtained with non-prenylated G{beta}{gamma}. Using quantitative protein titration and electrophysiology in live Xenopus oocytes, G{beta}{gamma} affinity for homotetrameric GIRK2 ranged from 4-30 {micro}M. Heterotetrameric GIRK1/2 showed a higher G{beta}{gamma} apparent affinity due to unique G{beta}{gamma}-docking site (anchor) in GIRK1, which enriches G{beta}{gamma} at the channel. Biochemical approaches and molecular dynamic simulations revealed that the G{beta}{gamma} anchor is formed by interacting N-terminal and distal C-terminal domains of the GIRK1 subunits, distinct from the G{beta}{gamma}-binding "activation" site(s) underlying channel opening. Thus, the affinity of G{beta}{gamma}-GIRK interaction is within the expected physiological range, while dynamic pre-coupling of G{beta}{gamma} to GIRK1-containing channels through high-affinity interactions further enhances the GPCR-Gi/o-GIRK signaling efficiency.

physiology↗

bioRxiv2025

Historic Genomes Uncover Demographic Shifts and Kinship Structures in Post-Roman Central Europe

Many European towns and villages trace their origins to Early Medieval foundations. In former Roman territories, their emergence has traditionally been linked to mass migrations from outside the Roman Empire. However, recent studies have emphasised local continuity with some individual-level mobility. We generated and analysed 248 historic genomes from Late Roman (3rd and 4th century CE) and Early Medieval (5th-8th century CE) burial sites in southern Germany, comparing them to over 2,500 contemporary and Iron Age genomes in addition to 1,344 modern-day genomes from Germany, Italy and Great-Britain. Despite small inferred Early Medieval period community sizes, genetic diversity exceeded that of modern German cities. In the Altheim graveyard, established in the 5th century by a group of Northern European descent, we inferred a demographic shift in the 6th century with the integration of newcomers with ancestry typical of a nearby Roman military camp, likely as a result of the collapse of Roman state structures. We reconstructed multigenerational pedigrees and, using a novel approach to infer ancestry of unsampled relatives, inferred immediate intermarriage between incoming and local groups, with a distinct tendency for men from former Roman background marrying women of northern descent. Burial proximity correlates strongly with kinship, in some cases spanning six generations. These communities were organized around small family units, exhibited loosely patrilineal or bilateral descent patterns, practiced reproductive monogamy, and avoided close-kin marriages. Such practices reflect broader transformations in family structures that began during the Late Roman period, were transferred to small agrarian societies in the Early Medieval period, and continued to shape European societies. By the 7th century, ongoing admixture had shaped genetic diversity patterns into those resembling Central Europe today.

genetics↗

bioRxiv2024

Deep Multimodal Representations and Classification of First-Episode Psychosis via Live Face Processing

Schizophrenia is a severe psychiatric disorder associated with a wide range of cognitive and neurophysiological dysfunctions and long-term social difficulties. In this paper, we test the hypothesis that integration of multiple simultaneous acquisitions of neuroimaging, behavioral, and clinical information will be better for prediction of early psychosis than unimodal recordings. We propose a novel framework to investigate the neural underpinnings of the early psychosis symptoms (that can develop into Schizophrenia with age) using multimodal acquisitions of neural and behavioral recordings including functional near-infrared spectroscopy (fNIRS) and electroencephalography (EEG), and facial features. Our data acquisition paradigm is based on live face-to-face interaction in order to study the neural correlates of social cognition in first-episode psychosis (FEP). We propose a novel deep representation learning framework, Neural-PRISM, for learning joint multimodal compressed representations combining neural as well as behavioral recordings. These learned representations are subsequently used to describe, classify, and predict the severity of early psychosis in patients, as measured by the Positive and Negative Syndrome Scale (PANSS) and Global Assessment of Functioning (GAF) scores. We found that incorporating joint multimodal representations from fNIRS and EEG along with behavioral recordings enhances classification between typical controls and FEP individuals. Additionally, our results suggest that geometric and topological features such as curvatures and path signatures of the embedded trajectories of brain activity enable detection of discriminatory neural characteristics in early psychosis.

neuroscience↗

bioRxiv2024

Toll-like receptor 3 orchestrates a conserved mechanism of heart regeneration

The humans heart responds to tissue damage with persistent fibrotic scarring. Unlike humans, zebrafish can repair cardiac injury and re-grow heart tissue throughout life. Recently, Toll-like receptor 3 (Tlr3) was identified as an important mediator of cardiac regeneration in neonatal mice. However, no functional analysis of tlr3 knock-out mutant zebrafish in respect to cardiac regeneration has yet been performed. We hypothesize that TLR3 signalling plays a central, conserved role in driving cardiac regeneration upon injury. Therefore, we focused on tlr3 mediated cardiac regeneration in zebrafish, ultimately discovering an evolutionary conserved mechanism of heart repair. Using histological, behavioural, and RNA-Sequencing analysis, we uncovered a conserved mechanism of tlr3 mediated cardiac repair after myocardial injury. Upon myocardial cryoinjury subjection, survival is decreased in tlr3-/- fish as compared to wildtype controls. Tlr3-/- zebrafish fail to recruit immune cells to the injured ventricle, resulting in impaired DNA repair and transcriptional reprogramming of cardiomyocytes. Mechanistically, we uncover an evolutionary conserved mechanism of tlr3 activation in fibroblasts promoting monocyte migration towards an injured ventricular area. Our data reveal tlr3 as a novel therapeutic target to promote cardiac regeneration. Every experiment including human participants has been approved by the ethics committee of the Medical University of Innsbruck (Ref. Nr.: 1262/2023). All experiments including the use of laboratory animals have been approved by the federal ministry of education, science, and research of Austria (Ref. Nr.: 2020-0.345.504).

molecular biology↗

bioRxiv2023

The human genome's vocabulary as proposed by the DNA language model GROVER

Large Language Models (LLMs) on natural language have achieved a level of performance that allows the generation of coherent and syntactically correct text. DNA sequence of genomes follows rules similar to natural language, but a distinguishing factor is the absence of a concept analogous to words. We established byte-pair tokenization on the human genome and trained a foundation language model called GROVER ("Genome Rules Obtained Via Extracted Representations") to select the optimal vocabulary with a custom fine-tuning task of next-k-mer prediction. We thus defined a dictionary of words/tokens in the human genome that best carries the information content for DNA language models. Analyzing GROVERs learned representations, we observed that token embeddings primarily encode information related to their frequency, sequence content, and length. Some tokens are almost exclusively localized in repeats, while the vast majority widely distributes over the genome. The model also learns context and lexical ambiguity. Average embeddings of genomic regions relate to functional genomics annotation and thus indicate that GROVER has learned these structures purely from the contextual relationships of tokens. That we can extract functional annotations from the genome, purely based on sequence representation to the trained model, highlights the extent of information content encoded by the sequence. This is supported by fine-tuning tasks on genome biology with questions of promoter identity and protein-DNA binding. GROVER learns sequence context, a sense for grammatical structures and language rules in the genome. This knowledge can be extracted and used to compose a grammar book for the code of life.

genomics↗

bioRxiv2023

Distinguishing word identity and sequence context in DNA language models

Transformer-based large language models (LLMs) are very suited for biological sequence data, because the structure of protein and nucleic acid sequences show many analogies to natural language. Complex relationships in biological sequence can be learned, although there may not be a clear concept of words, because they can be generated through tokenization. Training is subsequently performed for masked token prediction. With this strategy, the models learn both the token sequence identity and a larger sequence context. We developed a framework to interrogate what a model learns, which is both relevant for the interpretability of the model and to evaluate its potential for specific tasks. We used a DNA language model, which is trained on the human reference genome with a Bidirectional Encoder Representations from Transformers (BERT) model. In this model, tokens are defined with overlapping k-mers. To gain insight into the models learning, we interrogated how the model performs predictions, extracted token embeddings, and defined a fine-tuning benchmarking task to predict the next tokens of different sizes without overlaps. This task is very suited to evaluate different pretrained DNA language models, also called foundation models, since it does not interrogate specific genome biology, does not depend on the tokenization strategy, the size of the vocabulary, the dictionary, or the number of parameters used to train the model. Lastly, the task performs without leakage of information from token identity into the prediction task, which makes it particularly useful to evaluate the learning of sequence context. Through this assessment we discovered that the model with overlapping k-mers struggles to learn larger sequence context. Instead, the learned embeddings largely represent token sequence. Still, good performance is achieved for genome biology inspired fine-tuning tasks. Models with overlapping tokens may be used for tasks where a larger sequence context is of less relevance, but the token sequence directly represents the desired learning feature. This emphasizes the need to interrogate knowledge representation in biological large language models. Transparency is particularly important for biomedical use cases and an understanding of what the models are learning can be used to match the model to the desired task.

bioinformatics↗

bioRxiv2022

Mapping molecular determinants of Cav2.2 inhibition by RGK proteins and homologs in Xenopus oocytes

The CaV1 and CaV2 families of voltage-dependent calcium channels play a crucial role in neurotransmitter release, excitation-contraction and many other cellular processes. Comprised of the membrane pore-forming 1, intracellular {beta} and extracellular 2{delta} subunits, these channels have been targets for pharmacological intervention for decades. Physiological functions of CaV channels are attenuated by either constitutively or transiently bounds proteins in the cellular environment. The RGK (Rad, Gem, Rem, and Rem2) G-protein family potently inhibits CaV1 and CaV2 function in heterologous expression systems. RGK proteins bind to CaV{beta} and inhibit channel localization and activity by forming a ternary complex with CaV1. Here, we evaluated the influence of RGK proteins on CaV2.2 channels heterologously expressed in Xenopus oocytes. Both Gem and Rad showed no nucleotide dependency on its inhibitory function on CaV2.2. The G-domain and C-terminus could inhibit the CaV2.2 channel independently when co-expressed with channel subunits. Our results demonstrated that structural determinants in Gem, crucial for channel inhibition, lie within the 222-296 amino acid region containing both the partial G-domain and C-terminus as determined from chimeric CaV{beta}-Gem constructs. We expanded our mapping efforts and prepared various chimeras of Drosophila melanogaster (Dm) RGK sequences fused to CaV{beta} and showed that 22 residues in RGK2t and RGK3L C-terminal imparted complete CaV2.2 inhibition. Point mutations in the DmRGK C-terminus, conserved in mammalian RGK proteins, abrogated the CaV2.2 inhibition to a significant extent, pointing to a hot region in the extreme C-terminus for inhibition of CaV channels. Since RGK homologs are now recognized as physiological modulators in {beta}-adrenergic regulation of CaV channels, the relevance of this curious G-protein family deserves close examination.

physiology↗

bioRxiv2021

Domain General Processes for Interactive Touch

The nexus model of social processing proposes that the right temporal parietal junction (rTPJ) serves as a neural hub for cognitive social functions. We test the hypothesis that the rTPJ is a domain general region including somatosensory social functions. Neuroimaging findings and cross-brain coherence for right- and left-hand handclasps with real vs. simulated hands were consistent with the domain general model.

neuroscience↗

bioRxiv2020

Reconstitution of β-adrenergic regulation of CaV1.2: Rad-dependent and Rad-independent protein kinase A mechanisms.

IntroductionCardiac L-type voltage-gated CaV1.2 channels are crucial in physiological regulation of cardiac excitation-contraction coupling. Adrenergic modulation of CaV1.2 starts with activation of {beta}-adrenergic receptors (AR) and culminates in protein kinase A (PKA) - induced increase of calcium influx through CaV1.2 channels. To date, this cascade has never been fully reconstituted in heterologous systems; even partial reconstitution proved challenging and controversial. A recent study identified Rad, a calcium channel inhibitory protein, as an essential component of the adrenergic signaling cascade. We corroborated this finding, further characterized, and fully reconstituted, the complete {beta}-AR CaV1.2 modulation cascade in a heterologous expression system. ObjectiveOur primary goal was to heterologously reconstitute the complete {beta}-adrenergic cascade, and to investigate the role of Rad and additional molecular determinants in adrenergic regulation of cardiac CaV1.2. Methods and ResultsWe utilized the Xenopus oocyte heterologous expression system. We expressed CaV1.2 channel subunits, without or with Rad and {beta}1-AR or {beta}2-AR. To activate PKA, we injected cyclic AMP (cAMP) into the oocytes, or extracellularly applied isoproterenol (Iso) to stimulate {beta}-AR. Whole-cell Ba2+ currents served as readout. We find and distinguish between two distinct pathways of PKA modulation of CaV1.2: Rad-dependent (~80% of total) and Rad-independent. We separate the two mechanisms by showing distinct requirements for the cytosolic N- and distal C- termini of 1C and for the CaV{beta} subunit. Finally, for the first time, we reconstitute the complete pathway using agonist activation of either {beta}1-AR or {beta}2-AR. The reconstituted system reproduces the known features of {beta}-AR regulation in cardiomyocytes, such as a >2-fold increase in CaV1.2 current, a hyperpolarizing shift in activation curve, and a high constitutive activity of {beta}2-AR. ConclusionsThe adrenergic modulation of CaV1.2 is composed of two distinct pathways, Rad-independent and Rad-dependent. The latter contributes most of the {beta}-AR-induced enhancement of CaV1.2 activity, crucially depends on CaV{beta} subunit, and is differently regulated by {beta}1-AR and {beta}2-AR. The reconstitution of the full {beta}-AR cascade provides the means to address central unresolved issues related to roles of auxiliary proteins in the cascade, CaV1.2 isoforms, and will help to develop therapies for catecholamine-induced cardiac pathologies.

physiology↗

bioRxiv2018

The Ca2+ sensor protein CMI1 fine tunes root development, auxin distribution and responses

Signaling cross-talks between auxin, a regulator of plant development and Ca2+, a universal second messenger have been proposed to modulate developmental plasticity in plants. However, the underlying molecular mechanisms are largely unknown. Here we report that in Arabidopsis roots, auxin elicits specific Ca2+ signaling pattern that spatially coincide with the expression pattern of auxin-regulated genes. We identified the EF-hand protein CMI1 (Ca2+ sensor Modulator of ICR1) as an interactor of the ROP effector ICR1 (Interactor of Constitutively active ROP). CMI1 is monomeric in solution, changes its secondary structure at Ca2+ concentrations ranging from 10-9 to 10-8 M and its interaction with ICR1 is Ca2+ dependent, involving a conserved hydrophobic pocket. cmi1 mutants display an increased auxin response including shorter primary roots, longer root hairs, longer hypocotyls and altered lateral root formation while ectopic expression of CMI1 induces root growth arrest and reduced auxin responses at the root tip. When expressed alone, CMI1 is localized at the plasma membrane, the cytoplasm and in nuclei. Interaction of CMI1 and ICR1 results in exclusion of CMI1 from nuclei and suppression of the root growth arrest. CMI1 expression is directly upregulated by auxin while expression of auxin induced genes is enhanced in cmi1 concomitantly with repression of auxin induced Ca2+ increases in the lateral root cap and vasculature, indicating that CMI1 represses early auxin responses. Collectively, our findings identify a crucial function of Ca2+ signaling and CMI1 in root growth and suggest an auxin-Ca2+ regulatory feedback loop that fine tunes root development.

plant biology↗

bioRxiv2018

Rapid Therapeutic Recommendations in the Context of a Global Public Health Crisis using Translational Bioinformatics Approaches: A proof-of-concept study using Nipah Virus Infection

We live in a world of emerging new diseases and old diseases resurging in more aggressive forms. Drug development by pharmaceutical companies is a market-driven and costly endeavor, and thus it is often a challenge when drugs are needed for diseases endemic only to certain regions or which affect only a few patients. However, biomedical open data is accessible and reusable for reanalysis and generation of a new hypotheses and discovery. In this study, we leverage biomedical data and tools to analyze available data on Nipah Virus (NiV) infection. NiV infection is an emerging zoonosis that is transmissible to humans and is associated with high mortality rates. In this study, explored the application of computational drug repositioning and chemogenomic enrichment analyses using host transcriptome data to match drugs that could reverse the virus-induced gene signature. We performed analyses using two gene signatures: i) A previously published gene signature (n=34), and ii) a gene signature generated using the characteristic direction method (n= 5,533). Our predictive framework suggests that several drugs including FDA approved therapies like beclometasone, trihexyphenidyl, S-propranolol etc. could modulate the NiV infection induced gene signatures in endothelial cells. A target specific analysis of CXCL10 also suggests the potential application of Eldelumab, an investigative therapy for Crohns disease and ulcerative colitis, as a putative candidate for drug repositioning. To conclude, we also discuss challenges and opportunities in clinical trials (n-of-1 and adaptive trials) for repositioned drugs. Further follow-up studies including biochemical assays and clinical trials are required to identify effective therapies for clinical use. Our proof-of-concept study highlights that translational bioinformatics methods including gene expression analyses and computational drug repositioning could augment epidemiological investigations in the context of an emerging disease with no effective treatment.

microbiology↗