Biology subjects

Huckstep, H.

Publications and source records attributed to Huckstep, H..

4 recordsLinked to original sources

bioRxiv2026

Personalized phosphoproteomics establish mTORC1 as a regulator of exercise-induced insulin sensitization in human skeletal muscle

Exercise enhances skeletal muscle insulin sensitivity, but the signaling mechanisms responsible are poorly understood. Understanding them may open new therapeutic avenues for individuals with limited exercise capacity. Here, we used rapamycin to inhibit mTORC1 in combination with exercise and insulin stimulation in healthy men. A single dose of rapamycin enhanced the insulin-sensitizing effect of exercise by 53% on average compared to placebo. Responses varied widely across individuals (-40% to 218%), and we leveraged this variance through personalized phosphoproteomics to map the mTORC1-dependent signaling network in skeletal muscle. This identified the protein kinase MKNK2 as a candidate downstream effector, which we then targeted for functional validation. Pharmacological inhibition of MKNK2 with eFT508 in insulin-clamped mice reduced both whole-body and skeletal muscle insulin sensitivity, confirming a functional role for MKNK2 activity in muscle glucose uptake. We then used eFT508 in ex vivo incubated human skeletal muscle to map the signaling network downstream of MKNK2, identifying the translational initiator eIF4G1 as a further regulatory node. Together, these findings indicate that exercise-induced insulin sensitization is actively constrained by a negative feedback pathway running from mTORC1 through the translational regulators MKNK2 and eIF4G1, raising the possibility that rapid translation of unidentified target proteins contributes to fine-tuning glucose uptake.

physiology↗

bioRxiv2026

High-throughput screening in hiPSC-cardiac models reveals cardiomyocyte-specific cell cycle regulatory mechanisms

Introductory ParagraphMultiple regulatory mechanisms govern cardiomyocyte proliferation including epigenetic modifications, metabolism and mechanical load. However, it is unclear whether such mechanisms can be pharmacologically targeted to induce cardiomyocyte proliferation without affecting other cell types. Here, we develop a dual-reporter (TNNT2eGFP; PCNAmScarlet-I) and a high-throughput image-based pipeline in human induced pluripotent stem cell (hiPSC)-derived cardiomyocytes, with counter screening in non-myocytes, to identify compounds that selectively promote cardiomyocyte proliferation without affecting other cell types. We identify the PIM kinase inhibitor GDC-0339 as a cardiomyocyte-selective pro-proliferative compound. GDC-0339 induced proliferation of hiPSC-derived cardiomyocytes without activity in non-myocytes, non-cardiac fibroblasts or epithelial cells. Phosphoproteomic profiling of GDC-0339 in cardiomyocytes and non-cardiac fibroblasts revealed a cardiomyocyte-specific mechanism of action involving sarcomere disassembly via remodelling of the F-actin cytoskeleton and metabolic reprogramming to anaerobic metabolism via Pyruvate Dehydrogenase Kinases (PDKs). Thus, we uncover cardiomyocyte-specific mechanisms governing the cell cycle that are potentially druggable.

cell biology↗

bioRxiv2026

Ageing blunts the phospho-proteomic response to resistance exercise in humans

Ageing blunts the adaptive growth response in human skeletal muscle. To define the molecular processes underlying this impairment, we performed unbiased analysis of the phosphoproteome and total proteome in healthy young and old human skeletal muscle following acute resistance exercise (ResEX) and essential amino acid (EAA) ingestion. Ageing led to global suppression of the growth-induced global phosphoproteome, despite intact mTORC1 activation in old muscle. Our results identify widespread effects of ageing on skeletal muscle growth and highlight novel pathways for therapeutic development in aged skeletal muscle.

physiology↗

bioRxiv2025

Expression of the Close homolog of L-1 and embigin identifies activated myofibroblasts.

AbstractFibrosis is driven by the emergence of myofibroblasts, which are the primary producers of the extracellular matrix proteins that form fibrotic lesions. Despite this critical role, detecting myofibroblasts remains challenging due to the paucity of selective markers. We therefore screened for novel myofibroblast-specific markers, discovering that the expression of the close homolog of L1 (ChL-1) and embigin (Emb) distinguish activated myofibroblasts from their quiescent precursors. We report that ChL-1+/Emb+ fibroblasts: (1) emerge during cardiac inflammation, (2) have elevated expression of collagens and inflammatory factors and (3) localise to fibrotic zones - consistent with activated myofibroblasts. Mechanistically, Chl1+/Emb+ myofibroblasts differentiate from resident fibroblasts which upregulate these markers in response to proinflammatory cytokines, such as IL-1 and IL-17. Moreover, we show that embigin could be exploited to target antibody-based therapies to myofibroblasts and confirm this protein as a conserved marker of activated fibroblasts in multiple tissues and settings. Collectively, these findings identify ChL-1 and embigin as novel myofibroblast surface-markers that could be used to identify, enumerate and target this pathogenic population.

cell biology↗