Biology subjects

Howden, A. J. M.

Publications and source records attributed to Howden, A. J. M..

4 recordsLinked to original sources

bioRxiv2025

Single cell proteomic analysis defines discrete neutrophil functional states in human glioblastoma

Neutrophils are vital innate immune cells shown to infiltrate glioblastomas, however we currently lack the molecular understanding of their functional states within the tumour niche. Given that neutrophils are known to display a prominent discordance between mRNA and protein abundance, we developed ultra-sensitive mini-bulk and single cell proteomic (SCP) workflows to study the heterogeneity of peripheral blood and tumour associated neutrophils (TAN) from patients with glioblastoma. Mini-bulk analysis enabled a deeper protein coverage of circulating immature, mature and TAN populations, defining signatures of maturity and demonstrating that TANs resemble mature circulating neutrophils. Analysis of the SCP data resulted in the detection of >1,100 proteins from a single TAN providing a detailed characterization of neutrophil subsets in glioblastoma. Our approach shows evidence of pathogenic and anti-tumorigenic clusters and discovers cell states invisible to scRNAseq, opening new opportunities to selectively target pro-tumoural neutrophil states.

immunology↗

bioRxiv2024

LPS stimulates dynamic changes in B cell metabolism to promote proliferation

Naive B cells exit quiescence and enter a proliferative state upon activation, ultimately differentiating into antibody-secreting or memory B cells. Toll-like receptor (TLR) ligands, such as lipopolysaccharide (LPS), can serve as physiological stimuli to initiate this transition. Using quantitative proteomics, we show that TLR4 engagement induces metabolic reprogramming in murine B cells, increasing the expression of amino acid transporters and cholesterol biosynthetic enzymes. The amino acid transporter SLC7A5 is markedly upregulated following LPS stimulation, and conditional deletion of Slc7a5 impairs B cell proliferation, underscoring its essential role in B cell activation. LPS also elevates intracellular cholesterol levels, and inhibition of the rate-limiting enzyme HMG-CoA reductase blocks proliferation. This effect was mediated by a dual requirement for cholesterol metabolism and protein prenylation downstream of HMG-CoA reductase. Notably, this was not unique to TLR4 signalling but is also observed in B cells activated via TLR7, TLR9, CD40, or the B cell receptor. Together, these findings reveal that metabolic rewiring, including amino acid uptake and cholesterol metabolism, is an essential feature of B cell activation and proliferation.

immunology↗

bioRxiv2024

Decoding Cortical Circuits: Synaptic Signatures and Disease Vulnerabilities of Layer 5 Pyramidal Neuron Types

Cortical layer 5 (L5) intratelencephalic (IT) and pyramidal tract (PT) neurons are embedded in distinct information processing pathways. The morphology, connectivity, electrophysiological properties, and role in behavior of these neurons have been extensively analyzed. However, the molecular composition of their synapses remains largely uncharacterized. Here, we dissect the protein composition of the excitatory postsynaptic compartment of L5 neurons in intact somatosensory circuits, using an optimized proximity biotinylation workflow with subsynaptic resolution. We find distinct synaptic signatures of L5 IT and PT neurons that are defined by proteins regulating synaptic organization and transmission, including cell-surface proteins (CSPs), neurotransmitter receptors and ion channels. In addition, we find a differential vulnerability to disease, with a marked enrichment of autism risk genes in the synaptic signature of L5 IT neurons compared to PT neurons. Our results align with human studies and suggest that the excitatory postsynaptic compartment of L5 IT neurons is notably susceptible in autism. Together, our analysis sheds light on the proteins that regulate synaptic organization and function of L5 neuron types and contribute to their susceptibility in disease. Our approach is versatile and can be broadly applied to other neuron types to create a protein-based, synaptic atlas of cortical circuits.

neuroscience↗

bioRxiv2022

The Immunological Proteome Resource

The Immunological Proteome Resource (ImmPRes; http://immpres.co.uk/) is an open access public resource integrating proteomic data generated by large-scale mass-spectrometry analysis of murine hematopoietic populations. The initial focus is T lymphocytes and how their proteomes are shaped by immune activation, environment, and intracellular signalling pathways with an aim to expand it to B cells and innate immune cells. It is a multidisciplinary effort between immunology and mass spectrometry-based labs with the objective to help define an in-depth high-quality map of immune cell proteomes. Maintaining data reproducibility and integrity are a priority within the resource, thus there is an in-depth protocols section explaining in detail the sample processing and the mass spectrometry-based analysis. ImmPRes provides open access to proteomic datasets covering a wide range of murine leukocyte populations with analysis of copy numbers per cell of > 10,000 proteins, enabling new understanding of lymphocyte phenotypes. All data is accessible via a simple graphical interface that supports easy interrogation of the data and options to download data summaries and raw data files.

immunology↗