BACKGROUND:Insulin deficiency (ID) causes severe metabolic defects and death if untreated, while insulin therapy does not fully restore metabolic homeostasis. Leptin therapy corrects metabolic abnormalities and promotes survival in rodents unable to produce insulin, suggesting the existence of insulin-independent glucoregulatory mechanisms. METHODS:We use mice with diphtheria toxin-induced pancreatic β-cell ablation, resulting in severe insulin deficiency. We assess hepatic translation by polysome profiling, ribosome profiling (Ribo-seq), and RNA sequencing. To identify leptin-responsive hepatic factors, we compare the hepatic translatome during intracerebroventricular leptin treatment and following leptin withdrawal. Regenerating islet-derived protein 3 alpha (Reg3α) is subsequently overexpressed in the liver of ID mice to assess its metabolic effects. RESULTS:ID suppresses hepatic mechanistic target of rapamycin complex 1 (mTORC1) signaling and global protein synthesis while extensively remodeling the hepatic translatome. Translation of anabolic and glucose-metabolism pathways is reduced, whereas transcripts involved in lipid metabolism are selectively enhanced. Leptin treatment markedly increases hepatic Reg3α translation. Hepatic Reg3α overexpression significantly improves hyperglycemia in ID mice without altering insulin-stimulated AKT phosphorylation in key metabolic tissues. These findings identify hepatic translational rewiring as an important feature of ID and Reg3α as an insulin-independent glucoregulatory factor.

The transcription factor c-Fos plays an important role in hepatic metabolism; however, its role in metabolic dysfunction-associated steatotic liver disease (MASLD) and hepatocellular carcinoma (HCC) is unclear. Here, we show that hepatic c-Fos is induced by insulin after feeding and suppressed by glucagon during fasting in chow-fed mice. In lean mice, adenovirus-mediated c-Fos ectopic expression in the liver is sufficient to cause insulin resistance. In diet-induced obesity or after ectopic expression in hepatocytes, c-Fos promotes MASLD progression by altering PPAR signaling and fatty acid metabolism pathways. Mechanistically, c-Fos drives glycolysis, stress-associated MAPK, and insulin-related PI3K-Akt signaling, exacerbating metabolic dysregulation. In HCC, c-Fos expression correlates with PI3K-Akt, MAPK, and calcium signaling pathways activation. Moreover, c-Fos siRNA knockdown in human liver cancer cells reduces proliferation and increases apoptosis under lipotoxic or ER stress conditions. These findings identify c-Fos as a critical mediator of liver steatosis progression, linking hepatocyte signaling and metabolic reprogramming to liver dysfunction and tumorigenesis. ### Competing Interest Statement The authors have declared no competing interest. European Research Council, GA817940 Fund for Scientific Research, https://ror.org/03q83t159, 35112672, 40007740, 40007402 The ULB Foundation China Scholarship Council

In January 2005, Cell Metabolism released its very first issue, and this year we are proud to celebrate 20 years of publishing broadly impactful metabolic research. In this Voices series, we highlight individuals who, like Cell Metabolism, were just starting out in the metabolism field when they published in the pages of the first issues of this new journal. Over the past 20 years, these authors have continued their journeys in metabolism research, developing their own careers and their own laboratories, and they continue to be authors with us today. It is an honor to hear from these investigators and feel that the journal was a part of their own scientific journeys, and their careers part of ours.

The importance of immunometabolism in the development of metabolic diseases is clear. Yet, how certain metabolic disorders, such as insulin deficiency (ID), influence immune cell function, and vice versa, is poorly understood. Also, therapeutic strategies to harness the interplay between immune cells and metabolism are lacking. Here, we observe that ID rearranges the immune landscape of the liver, causing a decrease of T cells and an increase of the Kupffer cells, accompanied by a shift in the transcriptional signature and polarization of the latter. Treating ID mice with the protein S100A9 rescues the polarization and lipid-related changes caused by ID in the Kupffer cells, and, through them, rescues hypertriglyceridemia and hyperketonemia in a TLR4-dependent manner. Additionally, S100A9 acts on other immune niches to increase glucose uptake in skeletal muscle, improving hyperglycemia. In summary, our findings pinpoint the S100A9-TLR4 axis as a new tool to harness immune cells for improving ID-related metabolic dysfunction.

The transcription factor c-Fos plays a key role in liver metabolism, stress responses, and carcinogenesis. Here, the role of hepatic c-Fos in the pathophysiology of metabolic dysfunction-associated steatotic liver disease and hepatocellular carcinoma (HCC) is investigated. In chow-fed mice, hepatic c-Fos is induced by insulin after feeding and suppressed by glucagon during fasting. Adenovirus-mediated hepatic c-Fos ectopic expression is sufficient to induce insulin resistance in chow-fed mice. In models of diet-induced obesity and inducible hepatocyte-specific Fos-expressing mice, elevated c-Fos expression is associated with transcriptomic changes in PPAR signaling and fatty acid metabolism pathways. Mechanistically, ectopic c-Fos expression enhances glycolysis and activates stress-related MAPK and insulin-related PI3K-Akt signaling, which can contribute to metabolic dysregulation. In HCC, persistent c-Fos expression correlates with activation of PI3K-Akt, MAPK, and calcium signaling pathways. Functional studies show that c-Fos knockdown reduces proliferation and restores apoptotic sensitivity in HCC cells under lipotoxic or endoplasmic reticulum stress conditions. These findings identify c-Fos as a transcriptional regulator responsive to metabolic and hormonal cues, with potential roles in liver metabolic dysfunction and tumorigenesis.

Immunometabolism contributes to the development of metabolic diseases. Yet, how certain metabolic disorders, such as insulin deficiency (ID), influence immune cell function is poorly understood. Here, we observe that ID rearranges the immune landscape of the liver, causing a decrease in T cells and an increase in Kupffer cells, accompanied by a shift in the transcriptome and polarization of the latter. Treating ID mice with the protein S100A9 rescues the polarization and lipid-related changes caused by ID in the KCs, and rescues hypertriglyceridemia and hyperketonemia in a TLR4-dependent manner. Additionally, S100A9 acts on other immune niches to increase glucose uptake in skeletal muscle, improving hyperglycemia. In summary, the S100A9-TLR4 axis is a new tool to harness immune cells for improving ID-related metabolic dysfunction. ### Competing Interest Statement G.R. and R.C. are co-founders, directors, and stockholders of Diatheris SA. G.R. and R.C. are inventors on patent applications related to S100A9 protein. All other authors declare that they have no competing interests. Innosuisse Swiss Innovation Agency, https://ror.org/05a2bhn71, 104.549 IP-LS Innobooster, GRS-061/23 Fondation pour la recherche sur le diabète Foundation Valery Swiss National Science Foundation, 184767, 219229, 214870 Leona M. and Harry B. Helmsley Charitable Trust, 2405-06952 DiaGen Association

Type 1 diabetes mellitus (T1DM) is characterized by insulin deficiency leading to hyperglycemia and several metabolic defects. Insulin therapy remains the cornerstone of T1DM management, yet it increases the risk of life-threatening hypoglycemia and the development of major comorbidities. Here, we report an insulin signaling–independent pathway able to improve glycemic control in T1DM rodents. Co-treatment with recombinant S100 calcium-binding protein A9 (S100A9) enabled increased adherence to glycemic targets with half as much insulin and without causing hypoglycemia. Mechanistically, we demonstrate that the hyperglycemia-suppressing action of S100A9 is due to a Toll-like receptor 4–dependent increase in glucose uptake in specific skeletal muscles (i.e., soleus and diaphragm). In addition, we found that T1DM mice have abnormal systemic inflammation, which is resolved by S100A9 therapy alone (or in combination with low insulin), hence uncovering a potent anti-inflammatory action of S100A9 in T1DM. In summary, our findings reveal the S100A9-TLR4 skeletal muscle axis as a promising therapeutic target for improving T1DM treatment.

Supplementary Figure S1 from A Secreted Form of ADAM9 Promotes Carcinoma Invasion through Tumor-Stromal Interactions

Neddylation is a post-translational mechanism that adds a ubiquitin-like protein, namely neural precursor cell expressed developmentally downregulated protein 8 (NEDD8). Here, we show that neddylation in mouse liver is modulated by nutrient availability. Inhibition of neddylation in mouse liver reduces gluconeogenic capacity and the hyperglycemic actions of counter-regulatory hormones. Furthermore, people with type 2 diabetes display elevated hepatic neddylation levels. Mechanistically, fasting or caloric restriction of mice leads to neddylation of phosphoenolpyruvate carboxykinase 1 (PCK1) at three lysine residues-K278, K342, and K387. We find that mutating the three PCK1 lysines that are neddylated reduces their gluconeogenic activity rate. Molecular dynamics simulations show that neddylation of PCK1 could re-position two loops surrounding the catalytic center into an open configuration, rendering the catalytic center more accessible. Our study reveals that neddylation of PCK1 provides a finely tuned mechanism of controlling glucose metabolism by linking whole nutrient availability to metabolic homeostasis.

OBJECTIVE:p63 is a transcription factor within the p53 protein family that has key roles in development, differentiation and prevention of senescence, but its metabolic actions remain largely unknown. Herein, we investigated the physiological role of p63 in glucose metabolism.DESIGN:We used cell lines and mouse models to genetically manipulate p63 in hepatocytes. We also measured p63 in the liver of patients with obesity with or without type 2 diabetes (T2D).RESULTS:We show that hepatic p63 expression is reduced on fasting. Mice lacking the specific isoform TAp63 in the liver (p63LKO) display higher postprandial and pyruvate-induced glucose excursions. These mice have elevated SIRT1 levels, while SIRT1 knockdown in p63LKO mice normalises glycaemia. Overexpression of TAp63 in wild-type mice reduces postprandial, pyruvate-induced blood glucose and SIRT1 levels. Studies carried out in hepatocyte cell lines show that TAp63 regulates SIRT1 promoter by repressing its transcriptional activation. TAp63 also mediates the inhibitory effect of insulin on hepatic glucose production, as silencing TAp63 impairs insulin sensitivity. Finally, protein levels of TAp63 are reduced in obese persons with T2D and are negatively correlated with fasting glucose and homeostasis model assessment index.CONCLUSIONS:These results demonstrate that p63 physiologically regulates glucose homeostasis.

Supplementary Figure S2 from A Secreted Form of ADAM9 Promotes Carcinoma Invasion through Tumor-Stromal Interactions

The histone deacetylase sirtuin 6 (SIRT6) has been endowed with anti-cancer capabilities in many tumor types. Here, we investigate the impact of SIRT6-overexpression (SIRT6-OE) in Delta16HER2 mice, which are a bona fide model of HER2-positive breast cancer. After an initial delay in the tumor onset, SIRT6-OE induces a more aggressive phenotype of Delta16HER2 tumors promoting the formation of higher number of tumor foci and metastases than controls. This phenotype of SIRT6-OE tumors is associated with cancer stem cell (CSC)-like features and tumor dormancy, and low senescence and oxidative DNA damage. Accordingly, a sub-set of HER2-positive breast cancer patients with concurrent SIRT6-OE has a significant poorer relapse-free survival (RFS) probability than patients with low expression of SIRT6. ChIP-seq, RNA-seq and RT-PCR experiments indicate that SIRT6-OE represses the expression of the T-box transcription factor 3 ( Tbx3 ) by deacetylation of H3K9ac. Accordingly, loss-of-function mutations of TBX3 or low TBX3 expression levels are predictive of poor prognosis in HER2-positive breast cancer patients. Our work indicates that high levels of SIRT6 are indicative of poor prognosis and high risk of metastasis in HER2-positive breast cancer and suggests further investigation of TBX3 as a downstream target of SIRT6 and co-marker of poor-prognosis. Our results point to a breast cancer subtype-specific effect of SIRT6 and warrant future studies dissecting the mechanisms of SIRT6 regulation in different breast cancer subtypes.

Unrestrained ketogenesis leads to life-threatening ketoacidosis whose incidence is high in patients with diabetes. While insulin therapy reduces ketogenesis this approach is sub-optimal. Here, we report an insulin-independent pathway able to normalize diabetic ketogenesis. By generating insulin deficient male mice lacking or re-expressing Toll-Like Receptor 4 (TLR4) only in liver or hepatocytes, we demonstrate that hepatic TLR4 in non-parenchymal cells mediates the ketogenesis-suppressing action of S100A9. Mechanistically, S100A9 acts extracellularly to activate the mechanistic target of rapamycin complex 1 (mTORC1) in a TLR4-dependent manner. Accordingly, hepatic-restricted but not hepatocyte-restricted loss of Tuberous Sclerosis Complex 1 (TSC1, an mTORC1 inhibitor) corrects insulin-deficiency-induced hyperketonemia. Therapeutically, recombinant S100A9 administration restrains ketogenesis and improves hyperglycemia without causing hypoglycemia in diabetic mice. Also, circulating S100A9 in patients with ketoacidosis is only marginally increased hence unveiling a window of opportunity to pharmacologically augment S100A9 for preventing unrestrained ketogenesis. In summary, our findings reveal the hepatic S100A9-TLR4-mTORC1 axis in non-parenchymal cells as a promising therapeutic target for restraining diabetic ketogenesis.

p53 regulates several signaling pathways to maintain the metabolic homeostasis of cells and modulates the cellular response to stress. Deficiency or excess of nutrients causes cellular metabolic stress, and we hypothesized that p53 could be linked to glucose maintenance. We show here that upon starvation hepatic p53 is stabilized by O -GlcNAcylation and plays an essential role in the physiological regulation of glucose homeostasis. More specifically, p53 binds to PCK1 promoter and regulates its transcriptional activation, thereby controlling hepatic glucose production. Mice lacking p53 in the liver show a reduced gluconeogenic response during calorie restriction. Glucagon, adrenaline and glucocorticoids augment protein levels of p53, and administration of these hormones to p53 deficient human hepatocytes and to liver-specific p53 deficient mice fails to increase glucose levels. Moreover, insulin decreases p53 levels, and over-expression of p53 impairs insulin sensitivity. Finally, protein levels of p53, as well as genes responsible of O- GlcNAcylation are elevated in the liver of type 2 diabetic patients and positively correlate with glucose and HOMA-IR. Overall these results indicate that the O -GlcNAcylation of p53 plays an unsuspected key role regulating in vivo glucose homeostasis.

Reduced activation of energy metabolism increases adiposity inhumans and other mammals. Thus, exploring dietary and molecular mechanisms ableto improve energy metabolism is of paramount medical importance, as suchmechanisms can be leveraged as a therapy for obesity and related disorders.Here, we show that a designer protein-deprived dietenriched in free essential amino acids can i) promote the brown fatthermogenic program and fatty acid oxidation, ii) stimulate uncouplingprotein 1 (UCP1)-independent respiration in subcutaneous white fat, iii) changethe gut microbiota composition, and iv) prevent and reverse obesity anddysregulated glucose homeostasis in multiple mouse models, prolonging the healthylifespan. These effects are independent of unbalanced amino acid ratio,energy consumption, and intestinal calorie absorption. A brown fat-specificactivation of the mechanistic target of rapamycin complex 1 seems involved in thediet-induced beneficial effects, as also strengthened by in vitro experiments. Hence, our results suggest that brown andwhite fat may be targets of specific amino acids to control UCP1-dependentand -independent thermogenesis, thereby contributingto the improvement of metabolic health.

The discovery and administration of exogenous insulin has revolutionised diabetes treatment and continues, almost 100 years on, to be the basis for the management of insulin deficiency. However, insulin therapy still has potentially life-threatening side effects such as hypoglycaemia and increased risk of cardiovascular disease. So far, improvements in insulin therapy have focused mainly on modulating its pharmacokinetic and pharmacodynamic properties and improving delivery methods, while variations in the insulin sensitivity of peripheral tissues has received relatively little attention. Notably, tissue insulin sensitivity has been shown to vary considerably around the clock, which could contribute greatly to the effect (and risk of side effects) of a given dose of insulin. Recent evidence suggests that photic inputs regulate diurnal variations in the insulin sensitivity of metabolically relevant tissues via a previously unrecognised mechanism involving the ventromedial hypothalamic nucleus. Therefore, understanding the mechanisms underlying photic control of insulin action is of paramount medical importance. In addition, considering “when” (i.e., the time of day) could assist in deciding “how much” insulin should be administered and hence could aid the fine-tuning of insulin dosage, lowering the risk of side effects, and improving the quality of life of patients with insulin deficiencs.

Skeletal muscle promotes metabolic balance by regulating glucose uptake and the stimulation of multiple interorgan crosstalk. We show here that the catalytic activity of Vav2, a Rho GTPase activator, modulates the signaling output of the IGF1- and insulin-stimulated phosphatidylinositol 3-kinase pathway in that tissue. Consistent with this, mice bearing a Vav2 protein with decreased catalytic activity exhibit reduced muscle mass, lack of proper insulin responsiveness and, at much later times, a metabolic syndrome-like condition. Conversely, mice expressing a catalytically hyperactive Vav2 develop muscle hypertrophy and increased insulin responsiveness. Of note, while hypoactive Vav2 predisposes to, hyperactive Vav2 protects against high fat diet-induced metabolic imbalance. These data unveil a regulatory layer affecting the signaling output of insulin family factors in muscle.

Cancer therapy is limited, in part, by lack of specificity. Thus, identifying molecules that are selectively expressed by, and relevant for, cancer cells is of paramount medical importance. Here, we show that peptidyl-prolyl-cis-trans-isomerase (PPIase) FK506-binding protein 10 (FKBP10)-positive cells are present in cancer lesions but absent in the healthy parenchyma of human lung. FKBP10 expression negatively correlates with survival of lung cancer patients, and its downregulation causes a dramatic diminution of lung tumor burden in mice. Mechanistically, our results from gain- and loss-of-function assays show that FKBP10 boosts cancer growth and stemness via its PPIase activity. Also, FKBP10 interacts with ribosomes, and its downregulation leads to reduction of translation elongation at the beginning of open reading frames (ORFs), particularly upon insertion of proline residues. Thus, our data unveil FKBP10 as a cancer-selective molecule with a key role in translational reprogramming, stem-like traits, and growth of lung cancer.