Biology subjects

Milner, A.

Publications and source records attributed to Milner, A..

3 recordsLinked to original sources

bioRxiv2025

Glucagon-like peptide-1 receptor C-terminal tail phosphorylation determines signalling responses from pancreatic β-cells

The glucagon-like peptide-1 receptor (GLP-1R) is a class B G protein-coupled receptor that regulates glucose homeostasis by promoting cAMP production and insulin secretion in pancreatic {beta}-cells. Although receptor phosphorylation is known to influence GLP-1R signalling, its functional relevance in {beta}-cells remains incompletely understood. Here, we investigated the role of three C-terminal serine doublets (Ser441/442, Ser444/445, Ser451/452) in the human receptor using site-directed mutagenesis in INS-1 832/3 {beta}-cells deleted for endogenous GLP-1R expression. GLP-1 stimulation induced serine phosphorylation of wild-type GLP-1R, which was abolished in all individual doublet as well as triple doublet phospho-null mutants. Basal phosphorylation was detected in wild-type receptors under unstimulated conditions, suggesting dynamic cycling between phosphorylated and unphosphorylated states. All mutant receptors trafficked normally to the plasma membrane, but the triple alanine doublet (AA) mutant exhibited reduced GLP-1-induced internalisation and significantly decreased plasma membrane diffusion under vehicle conditions, indicating altered receptor dynamics even in the absence of agonist. Importantly, the Ser451/452 doublet was required for GRK2 recruitment to the active GLP-1R. Functionally, the triple AA mutant showed impaired GLP-1-induced cAMP generation with minimal change in {beta}-arrestin 2 recruitment, resulting in biased signalling. The cAMP defect was partially rescued by phosphodiesterase 4 (PDE4) inhibition with rolipram, implicating altered PDE engagement in the defective cAMP response. These findings demonstrate that both basal and agonist-induced GLP-1R phosphorylation are critical for receptor signalling in {beta}-cells, revealing a previously unappreciated role for basal phosphorylation in shaping receptor behaviour and establishing a positive link between GLP-1R phosphorylation and {beta}-cell signalling competence.

cell biology↗

bioRxiv2025

D- and L-Lactate enhance intestinal barrier function via activation of an apical HCAR1/Giα pathway in a human colonic epithelial cell model

The stereoisomers of lactate, L- and D- are not only metabolic substrates but also signalling molecules, capable of activating and signalling through its G protein-coupled receptor, Hydroxycarboxylic acid receptor 1 (HCAR1). These stereoisomers are both produced by the gut microbiota at millimolar concentrations creating a physiological environment for lactate-sensing unique to the gut yet, poorly understood. Here we identify a role for D-/L-lactate on intestinal barrier function. A human colonic epithelial cell model, Caco2, activated Gi signalling in response to both L- and D-lactate, although L-lactate exhibited a more potent and rapid Gi signal profile. When differentiated, apically but not basally treated D-/L-lactate enhanced tight junctions and reduced cell permeability, consistent with the apical localization of HCAR1. This improved barrier function occurred in a Gi-dependent manner. In addition, apical lactate rescued the reduced intestinal barrier function induced by lipopolysaccharides. This work highlights the potential for D-/L-lactate supplementation in improving gut health. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=113 SRC="FIGDIR/small/685091v1_ufig1.gif" ALT="Figure 1"> View larger version (29K): org.highwire.dtl.DTLVardef@28fa82org.highwire.dtl.DTLVardef@56a75borg.highwire.dtl.DTLVardef@10578c6org.highwire.dtl.DTLVardef@3c3cd2_HPS_FORMAT_FIGEXP M_FIG C_FIG

molecular biology↗

bioRxiv2024

Assessing the use of RIVPACS-derived invertebrate taxonomic predictions for river management

The River Invertebrate Prediction and Classification System (RIVPACS) is used widely in freshwater management to set targets for macroinvertebrate ecological health based on the expected scores of metrics such as WHPT or LIFE in the absence of anthropogenic stressors. An underutilised capability of RIVPACS-type models is the capability to predict expected macroinvertebrate community composition, which could function as a novel management metric for river health. We present a novel Monte-Carlo simulation approach that generates simulated expected communities for Englands rivers based on RIVPACS predictions. This allows for assessments of macroinvertebrate health using similarity calculations between observed and expected communities. We assess 10-year trends in similarity between 2010 and 2019 at 4172 sites in England, and contrast these trends with WHPT ASPT O/E trends in the same period. Similarity scores include both Chi-Squared and Hellinger methods, to prioritise rare and common species, respectively. We find that whilst most sites (63.3%) showed improvement in WHPT ASPT O/E in this period, most sites showed declines in similarity for Chi-Squared and Hellinger O/E (51.1% and 58.8%, respectively). We identified three case study regions showing contrasting trends and illustrate how the new RIVPACS-derived similarity calculations can track meaningful shifts in composition associated with water quality and multiple stressors including invasive species. RIVPACS-derived similarity calculations potentially provide a sensitive and practical management metric to assess ecosystem health, although further work is required to understand the composition of communities in changing environments with clear changes in stressor regimes.

ecology↗