Biology subjects

Walker, H.

Publications and source records attributed to Walker, H..

8 recordsLinked to original sources

bioRxiv2025

Functional architecture of cardiac TF regulatory landscapes in control of mammalian heart development

Congenital heart disease (CHD), the most common human birth defect, often results from disruptions in gene regulatory networks (GRNs) that control cardiac lineage specification and cell type identity during heart development. A conserved core set of cardiac transcription factors (TFs) orchestrates these processes through combinatorial interactions that are cell type-specific and tightly regulated across space and time. However, the genomic enhancer architecture that integrates upstream effectors to establish precise cardiac TF dosage and downstream transcriptional output remains largely unresolved. Here, we assessed the functional necessity of five developmental heart enhancer modules previously linked to the regulation of Gata4 and Hand2, core cardiac TFs exhibiting overlapping roles in myocardial and endocardial development. While individual enhancer deletions in mouse embryonic hearts revealed a surprising degree of transcriptional resilience, a subset of Gata4 enhancers proved indispensable for embryonic progression in a genetically compromised background. To achieve higher precision in cardiac cell type-specific enhancer prediction, we applied single-nucleus multiome profiling, enabling the delineation of cardiac cistromes underlying heart morphogenesis. By integrating this resource with deep learning applications, site-directed transgenesis, and chromatin conformation modeling, we mapped the cardiac enhancer repertoire and regulatory signatures that orchestrate Hand2 dynamics across distinct cardiac compartments and lineages. Genome editing further revealed an essential role for the Hand2-upstream regulatory interval (H2-URI) in transcriptional control of endocardial lineage effectors and, consequently, trabecular network formation and cardiac cushion patterning. Together our findings highlight substantial resilience in the cis-regulatory architectures governing cardiac TF dynamics and demonstrate that combinatorial integration of upstream lineage identities across modular enhancer landscapes establishes the cardiac cell type-specific programs driving heart morphogenesis. These results advance the reconstruction of cardiac GRNs and enhance the functional interpretation of CHD-associated variants.

developmental biology↗

bioRxiv2025

Collagen I promotes cancer cell survival via amino acid import and mTORC1 activation

Invasive breast and pancreatic cancer cells thrive within a collagen I-rich, poorly perfused extracellular matrix (ECM) network, necessitating robust metabolic adaptation to endure nutrient deficiency, such as glucose starvation. Here we demonstrate that collagen I is critical for the survival and growth of breast and pancreatic cancer cells, by maintaining their anabolic status. Mechanistically, collagen I promotes 2{beta}1 integrin-dependent mammalian target of rapamycin complex 1 (mTORC1) activation and drives the membrane localisation of the LAT1-4F2hc amino acid transporter. This process ensures a sustained intracellular essential amino acid supply, further fuelling mTORC1 activity and limiting autophagy. This collagen I-driven pathway is essential for cancer cell survival under nutrient stress, as inhibiting the activity of 2{beta}1 integrin or the LAT1-4F2hc transporter significantly reduces cell growth and invasion in both 2D and 3D models. Finally, the clinical relevance of these transporters is underscored by the significant upregulation of LAT1-4F2hc expression in basal-like breast and pancreatic cancer patients, correlating with poor prognosis and drug resistance. Collectively, our findings highlight that targeting the LAT1-4F2hc transporter might represent a highly promising therapeutic strategy to limit cancer cell growth and invasion in highly fibrotic and nutrient-deprived tumours.

cancer biology↗

bioRxiv2025

Impaired Spatiotemporal Encoding of Social Behavior and Anxiety in the Prefrontal Cortex of Mice Lacking ASD-Risk Gene Shank3.

The prefrontal cortex is a central regulator of complex behaviors, including social interaction and anxiety-related behaviors. The prefrontal cortex encodes these behaviors using heterogeneous groups of neurons, or ensembles, which collectively process inputs and communicate with distributed brain regions. We examined whether loss of the Autism-risk gene Shank3 alters the recruitment of neurons encoding socioemotional behavior collectively, or if abnormal activity during specific behaviors might affect functionally or anatomically defined populations of neurons. To do this, we combined spatially-resolved microendoscopic calcium imaging across the prefrontal microcircuit with functionally defined labeling of neurons as control and mutant mice engaged in social interaction or anxiety-provoking behaviors. We then utilized a non-biased transcriptomic method to identify neurons activated by social interactions. We show that the recruitment of heterogeneous neuronal populations are altered in a cell type and spatially dependent manner by loss of Shank3, with impaired recruitment of behavior-specific activity patterns within superficial, but not deeper aspects of the prefrontal cortex.

neuroscience↗

bioRxiv2025

Co-release of opposing signaling molecules from cortical neurons controls the escalation and release of aggression

Neuropeptides exert broad effects across the brain to influence behavior. However, the mechanism by which the brain uses neuropeptide signaling to exert top-down control over complex behavioral responses remains poorly understood. Prolonged social isolation induces a distinct internal state that results in sweeping changes to behavior, including increased aggression. Here, we find that isolation-induced aggression activates Tachykinin-2 expressing (Tac2+) neurons in the mouse medial prefrontal cortex (mPFC). Genetic characterization of Tac2+ cells in the mPFC reveals them to be a population of unexplored GABAergic neurons. In-depth behavioral analyses combined with in vivo recordings of neural activity demonstrate that mPFC Tac2+ neurons are tuned to distinct phases of isolation-induced aggression: investigatory behaviors that escalate to attack (aggression escalation) and attack itself (aggression release). Loss-of-function perturbations targeting the release of either Neurokinin B (NkB), the stimulatory peptide encoded by Tac2, or the inhibitory neurotransmitter, GABA, reveal that these signaling molecules exert dissociable control over aggression escalation and release. These findings identify distinct roles for opposing signaling molecules co-released from the same population of cells. This suggests a surprising neurochemical mechanism by which neuropeptidergic populations in the prefrontal cortex exert top-down influence over complex behavior via the co-release of signaling molecules with opposing actions but coordinated impacts on behavior.

neuroscience↗

bioRxiv2024

Tiny amphibious insects use tripod gait for seamless transition across land, water, and duckweed

Insects exhibit remarkable adaptability in their locomotive strategies across diverse environments, a crucial trait for foraging, survival, and predator avoidance. Microvelia, tiny 2-3 mm insects that adeptly walk on water surfaces, exemplify this adaptability by using the alternating tripod gait in both aquatic and terrestrial terrains. These insects commonly inhabit low-flow ponds and streams cluttered with natural debris like leaves, twigs, and duckweed. Using high-speed imaging and pose-estimation software, we analyze Microvelia spp.s movement across water, sandpaper (simulating land), and varying duckweed densities (10%, 25%, and 50% coverage). Our results reveal Microvelia maintain consistent joint angles and strides of their upper and hind legs across all duckweed coverages, mirroring those seen on sandpaper. Microvelia adjust the stride length of their middle legs based on the amount of duckweed present, decreasing with increased duckweed coverage and at 50% duckweed coverage, their middle legs strides closely mimic their strides on sandpaper. Notably, Microvelia achieve speeds up to 56 body lengths per second on water, nearly double those observed on sandpaper and duckweed (both rough, frictional surfaces), highlighting their higher speeds on low friction surfaces such as the waters surface. This study highlights Microvelias ecological adaptability, setting the stage for advancements in amphibious robotics that emulate their unique tripod gait for navigating complex terrains.

animal behavior and cognition↗

bioRxiv2023

Distinct transcriptional programs define a heterogeneous neuronal ensemble for social interaction

Reliable representations of information regarding complex behaviors including social interactions require the coordinated activity of heterogeneous cell types within distributed brain regions. Activity in the medial prefrontal cortex is critical in regulating social behavior, but our understanding of the specific cell types which comprise the social ensemble has been limited by available mouse lines and molecular tagging strategies which rely on the expression of a single marker gene. Here we sought to quantify the heterogeneous neuronal populations which are recruited during social interaction in parallel in a non-biased manner and determine how distinct cell types are differentially active during social interactions. We identify distinct populations of prefrontal neurons activated by social interaction by quantification of immediate early gene (IEG) expression in transcriptomically clustered neurons. This approach revealed variability in the recruitment of different excitatory and inhibitory populations within the medial prefrontal cortex. Furthermore, evaluation of the populations of IEGs recruited following social interaction revealed both cell-type and region-specific transcriptional programs, suggesting that reliance on a single molecular marker is insufficient to quantify activation across all cell types. Our findings provide a comprehensive description of cell-type specific transcriptional programs invoked by social interactions and reveal new insights into the heterogeneous neuronal populations which compose the social ensemble.

neuroscience↗

bioRxiv2022

A single amino acid transporter controls the uptake of priming-inducing beta-amino acids and the associated trade-off between induced resistance and plant growth.

Selected beta-amino acids, such as beta-aminobutyric acid (BABA) and R-beta-homoserine (RBH), can prime plants for resistance against broad-spectrum diseases. Here, we describe a genome-wide screen of fully annotated Arabidopsis T-DNA insertion lines for impaired in RBH-induced immunity (iri) against the downy mildew pathogen Hyaloperonospora arabidopsidis, yielding 104 lines that were partially affected and 4 lines that were completely impaired in RBH-induced resistance. The iri1-1 mutant phenotype could be confirmed by an independent T-DNA insertion in the same gene, encoding the high-affinity amino acid transporter LHT1. Using uptake experiments with IRI1/LHT1-expressing yeast cells and mass spectrometry-based quantification of RBH and BABA in leaves of mutant and over-expression lines of IRI1/LHT1, we demonstrate that IRI1/LHT1 acts as the main transporter for cellular uptake and systemic distribution of RBH and BABA. Subsequent characterisation of mutant and over-expression lines of IRI1/LHT1 for induced resistance and growth responses revealed that the level of IRI1/LHT1 expression determines the trade-off between induced resistance and plant growth by RBH and BABA.

plant biology↗

bioRxiv2021

The extracellular matrix promotes breast cancer cell growth under amino acid starvation by promoting tyrosine catabolism

Breast and pancreatic tumours are embedded in a collagen I-rich extracellular matrix (ECM) network, where nutrients are scarce due to limited blood flow and elevated tumour growth. Metabolic adaptation is required for cancer cells to endure these conditions. Here, we demonstrated that the presence of ECM supported the growth of invasive breast cancer cells, but not non-transformed mammary epithelial cells, and pancreatic cancer cells under amino acid starvation, through a mechanism that required macropinocytosis-dependent ECM uptake. Importantly, we showed that this behaviour was acquired during carcinoma progression. ECM internalisation, followed by lysosomal degradation, contributed to the upregulation of the intracellular levels of several amino acids, most notably tyrosine and phenylalanine. This resulted in elevated tyrosine catabolism on ECM under starvation, leading to increased fumarate levels, potentially feeding into the tricarboxylic acid cycle. Interestingly, this pathway was required for ECM-dependent cell growth under amino acid starvation, as the knockdown of p-hydroxyphenylpyruvate hydroxylase-like protein (HPDL), the third enzyme of the pathway, opposed cell growth on ECM in both 2D and 3D systems, without affecting cell proliferation on plastic. Finally, high HPDL expression correlated with poor prognosis in breast and pancreatic cancer patients. Collectively, our results highlight that the ECM in the tumour microenvironment represents an alternative source of nutrients to support cancer cell growth, by regulating phenylalanine and tyrosine metabolism.

cancer biology↗