Biology subjects

Foretz, M.

Publications and source records attributed to Foretz, M..

13 recordsLinked to original sources

bioRxiv2026

SIX1-dependent myofiber typology and metabolism controls muscle hypertrophy

The different types of muscle fibres respond in a specific way to hypertrophy or atrophy. The mechanisms underlying these heterogeneous adaptations remain poorly understood. Using single-nucleus RNA sequencing, we propose that fast glycolytic fibres show genetic limitations to hypertrophy induced by mechanical overload. We show that a prior fibre transition, achieved by reducing SIX1 protein expression (hypomorphism), enhances and accelerates overload-induced hypertrophy, bypassing the genetic limitations of fast glycolytic fibres. In contrast and unexpectedly, Six1 knockout in myofibers abolished overload-induced hypertrophy and instead caused atrophy of IIb/IIx fibers, despite the induction of a strong slow oxidative phenotype. In particular, Six1 deletion leads to metabolic defects caused by inhibition of glycolysis, AMPK and mitochondrial biogenesis. Our findings highlight the critical role of SIX1/AMPK/glycolysis-dependent aerobic metabolism in muscle growth and suggest that fibre type transitions, coupled with preserved metabolic function, may optimise hypertrophic responses.

physiology↗

bioRxiv2025

Common and distinct roles of AMPKγ isoforms in small-molecule activator-stimulated glucose uptake in mouse skeletal muscle

ObjectiveSmall-molecule activators targeting the allosteric drug and metabolite (ADaM) site of AMPK enhance insulin-independent glucose uptake in skeletal muscle and lower glucose in preclinical models of hyperglycemia. The regulatory AMPK{gamma} subunit plays a central role in energy sensing. While the skeletal muscle-selective {gamma}3 isoform is essential for AMP/ZMP-induced glucose uptake, it is dispensable for ADaM site-binding activators. We hypothesized that the predominant {gamma}1 isoform is required for ADaM site activator-stimulated glucose uptake in skeletal muscle. MethodsSingle-nucleus RNA sequencing (snRNA-seq) was performed on mouse and human skeletal muscle mapping AMPK subunit isoform distribution across resident cell types. To determine {gamma} isoform-specific requirements for activator-stimulated glucose uptake, skeletal muscle-specific inducible AMPK{gamma}1/{gamma}3 double knockout (im{gamma}1-/-/{gamma}3-/-) and single knockout (im{gamma}1-/- and im{gamma}3-/-) mice were generated. Ex vivo glucose uptake was measured following treatment with AICAR (AMP-mimetic) or MK-8722 (ADaM site activator), and in vivo MK-8722-induced blood glucose lowering was assessed. ResultssnRNA-seq revealed distinct AMPK isoform distribution: {gamma}1 was ubiquitously expressed, whereas {gamma}3 was enriched in glycolytic myofibers in both mouse and human skeletal muscle. Ex vivo, glucose uptake stimulated by either AICAR or MK-8722 was abolished in im{gamma}1-/-/{gamma}3-/- muscle, and MK-8722-induced blood glucose lowering was significantly blunted in vivo. AICAR but not MK-8722-stimulated muscle glucose uptake was abolished in im{gamma}3-/-, whereas both activators fully retained effects on glucose uptake and glucose lowering in im{gamma}1-/- mice. ConclusionsWhile {gamma}1 predominates in stabilizing the AMPK2{beta}2{gamma}1 complex, it is dispensable for AMPK activator-stimulated glucose uptake in skeletal muscle, whether mediated via the nucleotide-binding or ADaM site.

biochemistry↗

bioRxiv2025

Mechanism and cellular actions of the potent AMPK inhibitor BAY-3827

Inhibition of AMP-activated protein kinase (AMPK) is under increasing investigation for its therapeutic potential in many diseases, including certain cancers. However, existing AMPK- inhibitors available as tool compounds are largely limited to compound C/dorsomorphin and SBI-0206965, both of which suffer from poor selectivity and off-target effects. Here we describe the structure-based molecular insights and cellular actions of a recently identified potent AMPK inhibitor, BAY-3827. Kinase selectivity profiling and sequence analyses of kinases that are highly or weakly inhibited by BAY-3827 uncovered key conserved residues involved in its inhibitory mechanism. A 2.5 [A] co-crystal structure of the AMPK kinase domain (KD)-BAY-3827 complex and comparison with known KD-inhibitor structures, revealed an overlapping site in the ATP-binding pocket and an C helix-out conformation. A distinct feature of the BAY-3827-bound state is the formation of a disulfide bridge between the D helix Cys106 and the activation loop residue Cys174. This bridge appears to stabilize the activation loop such that Asn162 repositions the DFG motif Phe158 toward the C-terminal kinase lobe, displacing His137 and disrupting the regulatory spine, thereby promoting an inactive state. In hepatocytes, 2.5-5 M BAY-3827, but not the structurally resembling inactive BAY-974, fully blocked AMPK activator (MK-8722)-mediated phosphorylation of ACC1 and corresponding inhibition of lipogenesis. Unbiased transcriptome analysis in MK- 8722-treated wild-type and AMPK-null hepatocytes revealed that 5 M BAY-3827 downregulated >30% of MK-8722-stimulated AMPK-dependent genes. Based on its greater selectivity and potency substantiated by comprehensive structural and cellular investigations, BAY-3827 is a powerful tool to delineate AMPK functions. One-sentence summaryWe provide the mechanism of action of the potent and selective AMPK inhibitor BAY-3827, which blocks AMPK-dependent cellular functions.

biochemistry↗

bioRxiv2024

Control of alveolar bone development, homeostasis, and socket healing by salt inducible kinases

Alveolar bone supports and anchors teeth. The parathyroid hormone-related protein (PTHrP) pathway plays a key role in alveolar bone biology. Salt inducible kinases (SIKs) are important downstream regulators of PTH/PTHrP signaling in the appendicular skeleton where SIK inhibition increases bone formation and trabecular bone mass. However, the function of these kinases in alveolar bone remains unknown. Here, we report a critical role for SIK2/SIK3 in alveolar bone development, homeostasis, and socket healing after tooth extraction. Inducible SIK2/SIK3 deletion led to dramatic alveolar bone defects without changes in tooth eruption. Ablating these kinases impairs alveolar bone formation due to disrupted osteoblast maturation, a finding associated with ectopic periostin expression by fibrous cells in regions of absent alveolar bone at steady state and following molar extraction. Distinct phenotypic consequences of SIK2/SIK3 deletion in appendicular versus craniofacial bones prompted us to identify a specific transcriptomic signature in alveolar versus long bone osteoblasts. Thus, SIK2/SIK3 deletion illuminates a key role for these kinases in alveolar bone biology and highlights the emerging concept that different osteoblast subsets utilize unique genetic programs. Summary statementSIK2/SIK3 deletion in alveolar bone reduces bone formation and mass by impairing osteoblast maturation, unlike in long bones, where it increases bone formation and mass.

developmental biology↗

bioRxiv2023

Exercise training adaptations in liver glycogen and glycerolipids require hepatic AMP-activated protein kinase in mice

Regular exercise elicits adaptations in glucose and lipid metabolism that allow the body to meet energy demands of subsequent exercise bouts more effectively and mitigate metabolic diseases including fatty liver. Energy discharged during the acute exercise bouts that comprise exercise training may be a catalyst for liver adaptations. During acute exercise, liver glycogenolysis and gluconeogenesis are accelerated to supply glucose to working muscle. Lower liver energy state imposed by gluconeogenesis and related pathways activates AMP-activated protein kinase (AMPK), which conserves ATP partly by promoting lipid oxidation. This study tested the hypothesis that AMPK is necessary for liver glucose and lipid adaptations to training. Liver-specific AMPK12 knockout (LAKO) and wild type (WT) mice completed sedentary and exercise training protocols. Liver nutrient fluxes were quantified at rest or during acute exercise following training. Liver metabolites and molecular regulators of metabolism were assessed. Training increased liver glycogen in WT mice, but not in LAKO mice. The inability to increase glycogen led to lower glycogenolysis, glucose production, and circulating glucose during acute exercise in trained LAKO mice. Deletion of AMPK12 attenuated training-induced declines in liver diacylglycerides. In particular, training lowered the concentration of unsaturated and elongated fatty acids comprising diacylglycerides in WT mice, but not in LAKO mice. Training increased liver triacylglycerides and the desaturation and elongation of fatty acids in triacylglycerides of LAKO mice. These lipid responses were independent of differences in tricarboxylic acid cycle fluxes. In conclusion, AMPK is required for liver training adaptations that are critical to glucose and lipid metabolism. NEW & NOTEWORTHYThis study shows that the energy sensor and transducer, AMP-activated protein kinase, is necessary for an exercise training-induced: i) increase in liver glycogen that is necessary for accelerated glycogenolysis during exercise, ii) decrease in liver glycerolipids independent of TCA cycle flux, and iii) decline in the desaturation and elongation of fatty acids comprising liver diacylglycerides. The mechanisms defined in these studies have implications for use of regular exercise or AMPK-activators in patients with fatty liver.

physiology↗

bioRxiv2022

AMPK deficiency in smooth muscles causes persistent pulmonary hypertension after birth and premature death

We address a paradox, that AMPK may facilitate hypoxic pulmonary vasoconstriction and its deficiency precipitate pulmonary hypertension. Here we show that AMPK-1/2 deficiency in smooth muscles promotes persistent pulmonary hypertension of the newborn. Accordingly, dual AMPK-1/2 deletion in smooth muscles causes premature death of mice after birth, associated with increased muscularization and remodeling throughout the pulmonary arterial tree, reduced alveolar numbers and alveolar membrane thickening, but with no edema. Spectral Doppler ultrasound indicates pulmonary hypertension and attenuated hypoxic pulmonary vasoconstriction. Age-dependent right ventricular pressure elevation, dilation and reduced cardiac output was also evident. KV1.5 potassium currents of pulmonary arterial myocytes are markedly smaller under normoxia, which is known to facilitate pulmonary hypertension. Mitochondrial fragmentation and reactive oxygen species accumulation is also evident. Importantly, there is no evidence of systemic vasculopathy or hypertension in these mice. Moreover, hypoxic pulmonary vasoconstriction is attenuated by AMPK-1 or AMPK-2 deletion without triggering pulmonary hypertension.

pathology↗

bioRxiv2022

Metformin improves glycemia independently of skeletal muscle AMPK via enhanced intestinal glucose clearance

Metformin is an inexpensive oral anti-hyperglycemic agent used worldwide as a first-choice drug for the prevention of type 2 diabetes mellitus (T2DM). Although current view suggests that metformin exerts its anti-hyperglycemic effect by lowering hepatic glucose production, it has been proposed that metformin also reduce hyperglycemia by increasing glucose uptake in skeletal muscle via activation of AMP-activated protein kinase (AMPK). Herein, we demonstrate in lean and diet-induced obese (DIO) male and female mouse models that the anti-hyperglycemic effect of metformin occurs independently of muscle AMPK, and instead relies on elevated intestinal glucose clearance. Furthermore, we report that the AMPK activity is elevated in skeletal muscle from patients with T2DM following chronic metformin treatment, but this is not associated with enhanced peripheral insulin sensitivity. These results argue against existing paradigms and emphasize the non-essential role of muscle AMPK but important role of the intestine for the anti-hyperglycemic effect of metformin.

physiology↗

bioRxiv2022

Prolactin-induced AMPK stabilizes alveologenesis and lactogenesis through regulation of STAT5 signaling

AMP-activated protein kinase (AMPK) is an evolutionarily conserved serine/threonine kinase that regulates energy homeostasis at cellular and organismal levels. It has been shown to affect several steps of breast cancer progression in a context-dependent manner. However, its role in normal mammary gland development and physiology remains ill-explored. Here, we show that AMPK expression and activity increased within murine mammary epithelia from puberty to pregnancy with highest levels during lactation, and then declined during involution. In ex vivo cultures of mammary epithelial cells (MECs) in organotypic scaffolds, treatment with lactogenic hormone prolactin (PRL) enhanced AMPK expression and activity. To understand the role of AMPK on mammary morphogenesis in vivo, we generated mice with conditional knockout of AMPK isoforms 1 and 2 (AMPK KO) in MECs. AMPK KO mammary glands showed accelerated alveolar development with increased epithelial content of both luminal and myoepithelial lineages, suggestive of hyperproliferation. AMPK KO mice also showed elevated beta-casein expression during pregnancy and lactation. These observations were phenocopied upon treatment of ex vivo cultivated wild-type MECs with a cognate AMPK inhibitor. AMPK null MECs showed increased phosphorylated STAT5 which is known to drive alveologenesis downstream of prolactin signaling. Our study identifies a novel interplay between AMPK and PRL-STAT5 signaling that determines mammary alveologenesis and differentiation.

developmental biology↗

bioRxiv2021

Serotonin regulates hepcidin expression via a gut-liver axis

Iron is essential to key biological processes of all living organisms. Proper iron levels must be maintained to meet biological needs and prevent toxicity. Given the central role played by the hormone hepcidin in systemic iron homeostasis, extensive research has sought to identify regulators of its expression. Diverse evidence shows the gut to be an essential sensor and regulator of iron homeostasis, independently of other known hepcidin regulators, including bone marrow signals. Here we identify gut-derived serotonin as a key physiological factor in hepcidin regulation. In response to hypoxia, serotonin synthesized and secreted by enterochromaffin cells can act beyond the gut to repress hepcidin expression in the liver, through a 5-HT2B receptor-dependent pathway. Bone marrow transplant experiments clearly indicate the gut is responsible for hepcidin repression. This regulatory system appears to be conserved in humans: a significant negative correlation exists between hepcidin and serotonin levels in the serum of healthy individuals. Our findings imply hepcidin regulation by serotonin is a physiological process, and modulation of the gut serotonergic system may have broad therapeutic implications.

physiology↗

bioRxiv2021

Dual targeting of salt inducible kinases and CSF1R uncouples bone formation and bone resorption

Bone formation and resorption are typically coupled, such that the efficacy of anabolic osteoporosis treatments may be limited by bone destruction. The multi-kinase inhibitor YKL-05-099 potently inhibits salt inducible kinases (SIKs) and may represent a promising new class of bone anabolic agents. Here we report that YKL-05-099 increases bone formation in hypogonadal female mice without increasing bone resorption. Postnatal mice with inducible, global deletion of SIK2 and SIK3 show increased bone mass, increased bone formation, and, distinct from the effects of YKL-05-099, increased bone resorption. No cell-intrinsic role of SIKs in osteoclasts was noted. In addition to blocking SIKs, YKL-05-099 also binds and inhibits CSF1R, the receptor for the osteoclastogenic cytokine M-CSF. Modeling reveals that YKL-05-099 binds to SIK2 and CSF1R in a similar manner. Dual targeting of SIK2/3 and CSF1R induces bone formation without concomitantly increasing bone resorption and thereby may overcome limitations of most current anabolic osteoporosis therapies.

physiology↗

bioRxiv2020

Myeloid deletion and therapeutic activation of AMP-activated protein kinase (AMPK) do not alter atherosclerosis in male or female mice

ObjectiveThe dysregulation of myeloid-derived cell metabolism can drive atherosclerosis. AMP-activated protein kinase (AMPK) controls various aspects of macrophage dynamics and lipid homeostasis, which are important during atherogenesis. Approach and ResultsWe aimed to clarify the role of myeloid-specific AMPK signaling by using LysM-Cre to drive the deletion of both the 1 and 2 catalytic subunits (MacKO), in male and female mice made acutely atherosclerotic by PCSK9-AAV and Western diet-feeding. After 6 weeks of Western diet feeding, half received daily injection of either the AMPK activator, A-769662 or a vehicle control for a further 6 weeks. After 12 weeks, myeloid cell populations were not different between genotype or sex. Similarly, aortic sinus plaque size, lipid staining and necrotic area were not different in male and female MacKO mice compared to their littermate floxed controls. Moreover, therapeutic intervention with A-769662 had no effect. There were no differences in the amount of circulating total cholesterol or triglyceride, and only minor differences in the levels of inflammatory cytokines between groups. Finally, CD68+ area or markers of autophagy showed no effect of either lacking AMPK signaling or systemic AMPK activation. ConclusionsOur data suggest that while defined roles for each catalytic AMPK subunit have been identified, global deletion of myeloid AMPK signaling does not significantly impact atherosclerosis. Moreover, we show that intervention with the first-generation AMPK activator, A-769662, was not able to stem the progression of atherosclerosis. Highlights- The deletion of both catalytic subunits of AMPK in myeloid cells has no significant effect on the progression of atherosclerosis in either male or female mice - Therapeutic delivery of a first-generation AMPK activator (A-769662) for the last 6 weeks of 12-week study had no beneficial effect in either male or female mice - Studying total AMPK deletion may mask specific effects of each isoform and highlights the need for targeted disruption of AMPK phosphorylation sites via knock-in mutations, rather than the traditional "sledgehammer" knockout approach

biochemistry↗

bioRxiv2019

The hypoxic ventilatory response is facilitated by the activation of Lkb1-AMPK signalling pathways downstream of the carotid bodies

We recently demonstrated that the role of the AMP-activated protein kinase (AMPK), a ubiquitously expressed enzyme that governs cell-autonomous metabolic homeostasis, has been extended to system-level control of breathing and thus oxygen and energy (ATP) supply to the body. Here we assess the contribution to the hypoxic ventilatory response (HVR) of two upstream kinases that govern the activities of AMPK. Lkb1, which activates AMPK in response to metabolic stress and CaMKK2 which mediates the alternative Ca2+-dependent mechanism of AMPK activation. HVRs remained unaffected in mice with global deletion of the CaMKK2 gene. By contrast, HVRs were markedly attenuated in mice with conditional deletion of LKB1 in catecholaminergic cells, including carotid body type I cells and brainstem respiratory networks. In these mice hypoxia evoked hypoventilation, apnoea and Cheyne-Stokes-like breathing, rather than hyperventilation. Attenuation of HVRs, albeit less severe, was also conferred in mice carrying [~]90% knockdown of Lkb1 expression. Carotid body afferent input responses were retained following either [~]90% knockdown of Lkb1 or AMPK deletion. In marked contrast, LKB1 deletion virtually abolished carotid body afferent discharge during normoxia, hypoxia and hypercapnia. We conclude that Lkb1 and AMPK, but not CaMKK2, facilitate HVRs at a site downstream of the carotid bodies.

physiology↗

bioRxiv2018

AMPK promotes induction of a tumor suppressor FLCN through activation of TFEB independently of mTOR

AMP-activated protein kinase (AMPK) is a central energy sensor and master regulator of energy homeostasis. AMPK not only elicits acute metabolic responses, but also promotes metabolic reprogramming and adaptions in the long-term through regulation of specific transcription factors/co-activators. We performed a whole-genome transcriptome profiling in wild-type and AMPK-deficient mouse embryonic fibroblasts (MEF) and mouse primary hepatocytes that had been treated with two distinct classes of small-molecule AMPK activators, namely 5-aminoimidazole-4-carboxamide ribonucleotide (AICAR) or 991. This led to the identification of distinct compound-dependent gene expression signatures and to the discovery of several AMPK-regulated genes, including folliculin (Flcn), a gene encoding for a tumor suppressor and nutrient sensor. Gene set enrichment and pathway analyses identified the lysosomal pathway and the associated transcription factor EB (TFEB) as key transcriptional mediator responsible for AMPK-dependent gene expression changes. AMPK-induced Flcn expression was abolished in TFEB/TFE3 double knockout MEF and the promoter activity of Flcn was profoundly reduced when its putative TFEB-binding site was mutated. Mechanistically, we have found that AMPK promotes the dephosphorylation and nuclear localization of TFEB independently of mTOR activity.\n\nCollectively, we identified the AMPK-TFEB-FLCN axis as a potential key regulator for cellular and metabolic homeostasis. Moreover, data from zebrafish with physiologically and pharmacologically activated AMPK confirmed the AMPK-TFEB-FLCN cascade in vivo.

molecular biology↗