Biology subjects

Minchin, J. E.

Publications and source records attributed to Minchin, J. E..

3 recordsLinked to original sources

bioRxiv2025

In vivo CRISPR screening identifies regulators of hyperplastic and hypertrophic adipose remodelling in zebrafish

Adipose tissues exhibit a remarkable capacity to expand, regress, and remodel in response to energy status. The cellular mechanisms underlying adipose remodelling are central to metabolic health. Hypertrophic remodelling - characterised by the enlargement of existing adipocytes - is associated with insulin resistance, type 2 diabetes, and cardiovascular disease. In contrast, hyperplastic remodelling - in which new adipocytes are generated - is linked to improved metabolic outcomes. Despite its clinical importance, the regulation of hypertrophic and hyperplastic adipose morphology remains poorly understood. Here, we integrate human transcriptomic data with a quantitative CRISPR-imaging platform in zebrafish to identify regulators of adipose morphology. We developed an image-based phenotyping pipeline that captures lipid droplet size, number, and spatial patterning, and applied generalised additive modelling to quantify hyperplastic versus hypertrophic morphology signatures. Using this platform, we conducted an F0 CRISPR screen targeting 25 candidate genes and identified three that induced hypertrophic morphology (txnipa, mmp14b and foxp1b) and an additional candidate that altered total adiposity (kazna). For functional validation, we generated stable loss-of-function alleles for both zebrafish foxp1 paralogues. Spatial analysis along the anterior-posterior axis revealed that foxp1b mutants display developmental hypertrophy but profoundly blunted adaptive responses to high-fat diet ([~]68% reduction across all spatial zones), while foxp1a mutants show normal baseline morphology but disrupted spatial patterning of diet-induced hypertrophy. Together, these findings establish a scalable CRISPR-imaging platform for in vivo genetic screening of adipose morphology, and reveal distinct roles for Foxp1 paralogues in developmental patterning and adaptive responses to dietary challenge in adipose tissue.

developmental biology↗

bioRxiv2025

Human genetic studies and zebrafish models identify Plxna4 as a regulator of adiposity, somatic growth, and feeding behaviours

Obesity is a major public health crisis, affecting billions worldwide and increasing the risk of metabolic and cardiovascular diseases. While lifestyle factors play a role, genetic variation is a key determinant of both obesity susceptibility and the efficacy of treatment strategies. Recent studies have implicated the Semaphorin 3 signalling pathway in obesity; however, specific roles for pathway components remain largely unexplored. Here, we focus on Class A Plexins and their potential contributions to body weight regulation. Using large-scale genetic association data, we identified that rare, predicted loss-of-function mutations in PLXNA4 were associated with body mass index (BMI) in females. Furthermore, common variant analysis revealed that genetic variation at PLXNA4 was linked to BMI, height, and various neuropsychiatric disorders. To investigate the biological role of Plxna4, we generated zebrafish plxna4 loss-of-function mutants, which exhibited an 85-92% reduction in Plxna4 protein. Despite appearing morphologically normal, mutant zebrafish at juvenile stages were shorter, had increased body fat levels relative to size-matched wild-type siblings, and displayed hypertrophic subcutaneous adipose tissue. Feeding assays revealed that plxna4 mutants consumed more food than wild-type siblings and exhibited food-stimulated hyperactivity, characterised by increased swimming speed, higher speed variability, and frequent high-speed bursts. Together, these findings demonstrate a conserved role for Plxna4 in regulating feeding behaviour and body fat levels, providing new insights into the genetic basis of obesity and warranting further studies to elucidate the molecular mechanisms underlying these effects.

developmental biology↗

bioRxiv2022

Loss of Mfn1 but not Mfn2 enhances adipogenesis

ObjectiveA biallelic missense mutation in mitofusin 2 (MFN2) causes multiple symmetric lipomatosis and partial lipodystrophy, implicating disruption of mitochondrial fusion or interaction with other organelles in adipocyte differentiation, growth and/or survival. In this study, we aimed to document the impact of loss of mitofusin 1 (Mfn1) or 2 (Mfn2) on adipogenesis in cultured cells. MethodsWe characterised adipocyte differentiation of wildtype (WT), Mfn1-/- and Mfn2-/- mouse embryonic fibroblasts (MEFs) and 3T3-L1 preadipocytes in which Mfn1 or 2 levels were reduced using siRNA. ResultsMfn1-/- MEFs displayed striking fragmentation of the mitochondrial network, with surprisingly enhanced propensity to differentiate into adipocytes, as assessed by lipid accumulation, expression of adipocyte markers (Plin1, Fabp4, Glut4, Adipoq), and insulin-stimulated glucose uptake. RNA sequencing revealed a corresponding pro-adipogenic transcriptional profile including Pparg upregulation. Mfn2-/- MEFs also had a disrupted mitochondrial morphology, but in contrast to Mfn1-/- MEFs they showed reduced expression of adipocyte markers and no increase in insulin-stimulated glucose uptake. Mfn1 and Mfn2 siRNA mediated knockdown studies in 3T3-L1 adipocytes generally replicated these findings. ConclusionsLoss of Mfn1 but not Mfn2 in cultured pre-adipocyte models is pro-adipogenic. This suggests distinct, non-redundant roles for the two mitofusin orthologues in adipocyte differentiation.

cell biology↗