Many cancer therapies achieve durable control without complete tumor eradication, suggesting that disrupting tumor organization may be more critical than killing cells. We propose that effective treatments converge by destabilizing the tumor's Group Phenotypic Composition (GPC), the functional and spatial organization of interacting cell populations. When this organization collapses, tumors lose coherence. This perspective provides a unifying framework for designing therapies targeting tumor-level dynamics rather than cell number alone.

Methionine is indispensable for cell proliferation, stem cell maintenance, and epigenetic regulation, three processes that are central to embryonic development. Here, we assessed the consequences of short-term dietary methionine restriction (MR) on mouse embryonic organ growth. In comparison with growth of the liver and heart which was unaffected, MR for 5 days led to a severe reduction in neuronal production and neocortex growth. Progenitor cohort labeling revealed a time-dependent sensitivity to MR and cell cycle analysis indicated that progenitors are stalled in S/G2 phases following MR. Unexpectedly, neuronal production was completely rescued at birth when switching the dam back to control diet, uncovering a mechanism of catch-up growth. We used imaging mass cytometry to probe metabolic and epigenetic markers in neural progenitors following MR and during catch-up growth. Altogether, our data uncover a reversible state of quiescence in S/G2 which is metabolically distinct from G0 quiescence and associated with efficient catch-up growth of the neocortex.

Pyruvate metabolism defects lead to severe neuropathies such as the Leigh syndrome (LS) but the molecular mechanisms underlying neuronal cell death remain poorly understood. Here, we unravel a connection between pyruvate metabolism and the regulation of the epitranscriptome that plays an essential role during brain development. Using genetically engineered mouse model and primary neuronal cells, we identify the transcription factor E4F1 as a key coordinator of AcetylCoenzyme A (AcCoA) production by the pyruvate dehydrogenase complex (PDC) and its utilization as an essential co-factor by the Elongator complex to acetylate tRNAs at the wobble position uridine 34 (U34). E4F1-mediated direct transcriptional regulation of Dlat and Elp3, two genes encoding key subunits of the PDC and of the Elongator complex, respectively, ensures proper translation fidelity and cell survival in the central nervous system (CNS) during mouse embryonic development. Furthermore, analysis of PDH-deficient cells highlight a crosstalk linking the PDC to ELP3 expression that is perturbed in LS patients.

Methionine -an essential amino acid that has to be provided by nutrition- and its metabolite S-Adenosyl methionine (SAM) are indispensable for cell proliferation, stem cell maintenance and epigenetic regulation [1][1]–[5][2], three processes that are central to embryonic development [6][3]. Previous studies using chronic dietary restriction of methyl donors prior to and during gestation indicated that methionine restriction (MR) is detrimental to the development or growth of the neocortex [7][4],[8][5], however, the consequences of acute MR have not been extensively studied. Here, we designed a dietary MR regime coinciding with the neurogenic phases of neocortex development in the mouse. Our results indicate that dietary MR for 5 days leads to a severe reduction in neocortex growth and neuronal production. In comparison, growth of the liver and heart was unaffected, highlighting an organ-specific response to MR which was also observed at the cellular and molecular levels. Progenitor cohort labeling revealed a time-dependent sensitivity to MR and cell cycle analyses indicated that after 5 days of MR, progenitors are stalled in the S/G2 phases. Unexpectedly, neocortex growth reduction induced after 5 days of MR is completely rescued at birth when switching the dam back to control diet for the remaining of gestation, uncovering a mechanism of catch-up growth. Using multiplexed imaging we probed metabolic and epigenetic markers following MR and during catch-up growth and show that pyruvate metabolism is rewired in progenitors. Altogether, our data uncover a transient state of quiescence in G2/S which is metabolically distinct from G0 quiescence and associated with efficient catch-up growth. More globally, our study highlights both the extreme sensitivity of the developing neocortex to acute dietary changes and its remarkable plasticity. ### Competing Interest Statement The authors have declared no competing interest. [1]: #ref-1 [2]: #ref-5 [3]: #ref-6 [4]: #ref-7 [5]: #ref-8

Introduction et objectifs Depuis sa création en 2008, le Réseau d’histologie expérimentale de Montpellier (RHEM) assure la traçabilité de tous les échantillons issus des modèles pré-cliniques générés par les chercheurs de la région. Pour valoriser cette collection biologique précieuse, le RHEM a créé, à l’image des biobanques humaines, une biobanque dédiée aux tissus de modèles précliniques, « Biological tissue collection » (BTC).Cette BTC vise à permettre aux chercheurs, en évitant de répliquer des expérimentations animales déjà réalisées, de (1) promouvoir une recherche éthique et responsable (règle des 3R), en réduisant ainsi le nombre d’animaux utilisés, (2) gagner du temps et de l’argent, par l’utilisation de lames histologiques issues de blocs déjà générés, (3) raffiner après l’expérimentation initiale, c’est-à-dire exploiter au mieux les résultats obtenus, via des collaborations scientifiques. Matériels et méthodes Le RHEM trace tous les échantillons issus de modèles pré-cliniques, ainsi que les données associées à ces blocs FFPE (fixés au formol et inclus en paraffine). Nous avons développé un portail web (BTC) permettant d’interroger cette biobanque (https://btc.rhem.cnrs.fr/). Résultats La BTC, ouverte depuis mars 2023, permet ainsi l’interrogation à distance, par la communauté scientifique française et internationale, de la biobanque contenant un peu moins de 30 000 blocs issus majoritairement de modèles murins dont 45 % proviennent du système digestif et 13 % du système lymphoïde. Cette interrogation se fait sur la base de divers critères (espèces, gène, âge, sexe, organe) dans le but d’obtenir des lames histologiques pour pouvoir réaliser des colorations, des immunomarquages, des analyses spatiales… Conclusion Cet outil unique de partage public offre aux chercheurs de nouvelles possibilités d’étudier l’expression de biomarqueurs dans des tissus de modèles de maladies humaines difficilement accessibles ou coûteux tels que des modèles de cancer, d’arthrose, ou d’atteinte infectieuse, cardiovasculaire ou centrale.

Cutaneous melanoma is an invasive cancer with a worldwide annual death toll of 57,000 (Arnold et al., JAMA Dermatol 2022). In a metastatic state, surgical interventions are not curative and must be coupled with targeted therapy, or immunotherapy. However, resistance appears almost systematically and late-stage prognosis can remain poor. The complexity to eradicate melanoma stems from its plasticity; these cancer cells continually adapt to the tumor microenvironment, which leads to treatment resistance. Our primary assumption is that therapeutic resistance relies in part on a series of non-genetic transitions including changes in the metabolic states of these cancer cells. The 3D spatial distribution of blood vessels that are sources of nutrition and oxygen that drive this metabolic status is an important variable for understanding zoning aspects of this adaptation process. Using Whole Slide Images (WSI) of melanoma tumors from Patient-Derived Xenograft (PDX) mouse models, we build 3D vascular models to help predict and understand the metabolic states of cancer cells within the tumor. Our 3D reconstruction pipeline was based on PDX tumor samples sectioned over 2mm depth and stained with Hematoxylin and Eosin (H&E). The pipeline involves three primary steps, including 2D vessel segmentation using Deep Learning, intensity- and affine-based image registration, and 3D reconstruction using interpolation and 3D rendering (allowing for better interaction with biologists, pathologists, and clinicians). The originality of our computer-assisted pipeline is its capability to (a) deal with sparse data (i.e., not all tissue sections were readily available), and (b) adapt to a multitude of WSI-related challenges (e.g., epistemic uncertainty, extended processing times due to WSI scale, etc.). We posit both our 3D reconstruction pipeline, quantitative results of the major stages of the process, and a detailed illustration of the challenges faced, presenting resolutions to improve the pipeline's efficiency.

Cutaneous melanoma is a highly invasive tumor and, despite the development of recent therapies, most patients with advanced metastatic melanoma have a poor clinical outcome. The most frequent mutations in melanoma affect the BRAF oncogene, a protein kinase of the MAPK signaling pathway. Therapies targeting both BRAF and MEK are effective for only 50% of patients and, almost systematically, generate drug resistance. Genetic and non-genetic mechanisms associated with the strong heterogeneity and plasticity of melanoma cells have been suggested to favor drug resistance but are still poorly understood. Recently, we have introduced a novel mathematical formalism allowing the representation of the relation between tumor heterogeneity and drug resistance and proposed several models for the development of resistance of melanoma treated with BRAF/MEK inhibitors. In this paper, we further investigate this relationship by using a new computational model that copes with multiple cell states identified by single cell mRNA sequencing data in melanoma treated with BRAF/MEK inhibitors. We use this model to predict the outcome of different therapeutic strategies. The reference therapy, referred to as “continuous” consists in applying one or several drugs without disruption. In “combination therapy”, several drugs are used sequentially. In “adaptive therapy” drug application is interrupted when the tumor size is below a lower threshold and resumed when the size goes over an upper threshold. We show that, counter-intuitively, the optimal protocol in combination therapy of BRAF/MEK inhibitors with a hypothetical drug targeting cell states that develop later during the tumor response to kinase inhibitors, is to treat first with this hypothetical drug. Also, even though there is little difference in the timing of emergence of the resistance between continuous and adaptive therapies, the spatial distribution of the different melanoma subpopulations is more zonated in the case of adaptive therapy.

E4F1 is essential for early embryonic mouse development and for controlling the balance between proliferation and survival of actively dividing cells. We previously reported that E4F1 is essential for the survival of murine p53-deficient cancer cells by controlling the expression of genes involved in mitochondria functions and metabolism, and in cell-cycle checkpoints, including CHEK1, a major component of the DNA damage and replication stress responses. Here, combining ChIP-Seq and RNA-Seq approaches, we identified the transcriptional program directly controlled by E4F1 in Human Triple-Negative Breast Cancer cells (TNBC). E4F1 binds and regulates a limited list of direct target genes (57 genes) in these cells, including the human CHEK1 gene and, surprisingly, also two other genes encoding post-transcriptional regulators of the ATM/ATR-CHK1 axis, namely, the TTT complex component TTI2 and the phosphatase PPP5C, that are essential for the folding and stability, and the signaling of ATM/ATR kinases, respectively. Importantly, E4F1 also binds the promoter of these genes in vivo in Primary Derived Xenograft (PDX) of human TNBC. Consequently, the protein levels and signaling of CHK1 but also of ATM/ATR kinases are strongly downregulated in E4F1-depleted TNBC cells resulting in a deficiency of the DNA damage and replicative stress response in these cells. The E4F1-depleted cells fail to arrest into S-phase upon treatment with the replication-stalling agent Gemcitabine, and are highly sensitized to this drug, as well as to other DNA-damaging agents, such as Cisplatin. Altogether, our data indicate that in breast cancer cells the ATM/ATR-CHK1 signaling pathway and DNA damage-stress response are tightly controlled at the transcriptional and post-transcriptional level by E4F1.

The p53 pathway is functionally inactivated in most, if not all, human cancers. The p53 protein is a central effector of numerous stress-related molecular cascades. p53 controls a safeguard mechanism that prevents accumulation of abnormal cells and their transformation by regulating DNA repair, cell cycle progression, cell death, or senescence. The multiple cellular processes regulated by p53 were more recently extended to the control of metabolism and many studies support the notion that perturbations of p53-associated metabolic activities are linked to cancer development, as well as to other pathophysiological conditions including aging, type II diabetes, and liver disease. Although much less documented than p53 metabolic activities, converging lines of evidence indicate that other key components of this tumor suppressor pathway are also involved in cellular metabolism through p53-dependent as well as p53-independent mechanisms. Thus, at least from a metabolic standpoint, the p53 pathway must be considered as a non-linear pathway, but the complex metabolic network controlled by these p53 regulators and the mechanisms by which their activities are coordinated with p53 metabolic functions remain poorly understood. In this review, we highlight some of the metabolic pathways controlled by several central components of the p53 pathway and their role in tissue homeostasis, metabolic diseases, and cancer.

Growing evidence supports the importance of the p53 tumor suppressor in metabolism but the mechanisms underlying p53-mediated control of metabolism remain poorly understood. Here, we identify the multifunctional E4F1 protein as a key regulator of p53 metabolic functions in adipocytes. While E4F1 expression is upregulated during obesity, E4f1 inactivation in mouse adipose tissue results in a lean phenotype associated with insulin resistance and protection against induced obesity. Adipocytes lacking E4F1 activate a p53-dependent transcriptional program involved in lipid metabolism. The direct interaction between E4F1 and p53 and their co-recruitment to the Steaoryl-CoA Desaturase-1 locus play an important role to regulate monounsaturated fatty acids synthesis in adipocytes. Consistent with the role of this E4F1-p53-Steaoryl-CoA Desaturase-1 axis in adipocytes, p53 inactivation or diet complementation with oleate partly restore adiposity and improve insulin sensitivity in E4F1-deficient mice. Altogether, our findings identify a crosstalk between E4F1 and p53 in the control of lipid metabolism in adipocytes that is relevant to obesity and insulin resistance.

Background: The TP53 gene is one of the most commonly inactivated tumor suppressors in human cancers. p53 functions during cancer progression have been linked to a variety of transcriptional and non-transcriptional activities that lead to the tight control of cell proliferation, senescence, DNA repair, and cell death. However, converging evidence indicates that p53 also plays a major role in metabolism in both normal and cancer cells. Scope of review: We provide an overview of the current knowledge on the metabolic activities of wild type (WT) p53 and highlight some of the mechanisms by which p53 contributes to whole body energy homeostasis. We will also pinpoint some evidences suggesting that deregulation of p53-associated metabolic activities leads to human pathologies beyond cancer, including obesity, diabetes, liver, and cardiovascular diseases. Major conclusions: p53 is activated when cells are metabolically challenged but the origin, duration, and intensity of these stresses will dictate the outcome of the p53 response. p53 plays pivotal roles both upstream and downstream of several key metabolic regulators and is involved in multiple feedback-loops that ensure proper cellular homeostasis. The physiological roles of p53 in metabolism involve complex mechanisms of regulation implicating both cell autonomous effects as well as autocrine loops. However, the mechanisms by which p53 coordinates metabolism at the organismal level remain poorly understood. Perturbations of p53-regulated metabolic activities contribute to various metabolic disorders and are pivotal during cancer progression. (C) 2019 The Authors. Published by Elsevier GmbH.

Single-cell transcriptomics offers unprecedented opportunities to infer the ligand-receptor (LR) interactions underlying cellular networks. We introduce a new, curated LR database and a novel regularized score to perform such inferences. For the first time, we try to assess the confidence in predicted LR interactions and show that our regularized score outperforms other scoring schemes while controlling false positives. SingleCellSignalR is implemented as an open-access R package accessible to entry-level users and available from https://github.com/SCA-IRCM. Analysis results come in a variety of tabular and graphical formats. For instance, we provide a unique network view integrating all the intercellular interactions, and a function relating receptors to expressed intracellular pathways. A detailed comparison of related tools is conducted. Among various examples, we demonstrate SingleCellSignalR on mouse epidermis data and discover an oriented communication structure from external to basal layers.

BACKGROUND:The ubiquitin proteasome pathway is involved in the pathogenesis of psoriasis and proteasome subunits are increased in lesional psoriatic skin. Recent works have highlighted that proteasome levels can be regulated through modulation of proteasome assembly notably by the proteasome maturation protein POMP.OBJECTIVES:To investigate whether proteasome assembly and POMP expression are modified in psoriatic skin.METHODS:Proteasome assembly as well as expression of proteasome regulators were assessed in non-lesional and lesional psoriatic skin using native gel electrophoresis and western blots respectively. The protein and mRNA expression levels of POMP were compared by western blots, immunohistochemistry and quantitative polymerase chain reaction. The role of POMP in keratinocyte proliferation and differentiation was assessed by silencing POMP gene expression by RNA interference in human immortalized keratinocyte HaCaT cells.RESULTS:Both 20S and 26S proteasomes (and their respective proteolytic activities) as well as the main proteasome regulators are increased in lesional psoriatic skin. POMP binds to 20S precursor complexes and is overexpressed in lesional epidermal psoriatic skin, supporting that POMP-mediated proteasome assembly is increased in psoriatic skin. POMP silencing inhibited HaCaT cell proliferation and induced apoptosis through the inhibition of the proteasome assembly. Moreover POMP partial depletion decreased the expression of the differentiation markers keratin 10 and involucrin during the [Ca2+]-induced HaCaT cells differentiation.CONCLUSION:Altogether these results establish a potential role for POMP and proteasome assembly in psoriasis pathogenesis.

The ubiquitin-proteasome pathway is involved in the pathogenesis of psoriasis and proteasome subunits are increased in lesional psoriatic skin. Recent works have highlighted that proteasome levels can be regulated through modulation of proteasome assembly notably by the proteasome maturation protein POMP. We investigated whether proteasome assembly and POMP expression are modified in psoriatic skin. Proteasome assembly as well as expression of proteasome regulators were assessed in non-lesional and lesional psoriatic skin using native gel electrophoresis and western blots, respectively. The protein and mRNA expression levels of POMP were compared by western blots, immunohistochemistry and qPCR. The role of POMP in keratinocyte proliferation and differentiation was assessed by silencing POMP gene expression by RNA interference in HaCaT keratinocytes. We show that both 20S and 26S proteasomes (and their respective proteolytic activities) as well as the main proteasome regulators (PA28αβ, PA28γ, PA200 and 19S complex) are increased in lesional psoriatic skin. POMP binds to 20S precursor complexes and is overexpressed in lesional epidermal psoriatic skin, supporting that POMP-mediated proteasome assembly is increased in psoriatic skin. POMP silencing inhibited HaCaT cell proliferation and induced apoptosis through the inhibition of proteasome assembly. Moreover POMP partial depletion decreased the expression of the differentiation markers keratin 10 and involucrin during the calcium - induced HaCaT cells differentiation. Altogether these results suggest a role for POMP and proteasome assembly in psoriasis pathogenesis.

L’activité protéolytique et l’expression des sous-unités des protéasomes sont augmentées dans l’épiderme psoriasique. Nos précédents résultats étaient en faveur d’une régulation post-transcriptionnelle de cette expression. Notre objectif était d’étudier l’assemblage du protéasome et l’expression d’une protéine chaperonne clé de l’assemblage du protéasome, la protéine de maturation du protéasome (POMP), dans la peau psoriasique et au cours de la différenciation kératinocytaire. Les expressions des protéasomes et de POMP étaient évaluées par IHC, WB et RT-PCR. L’assemblage du protéasome était étudié par électrophorèse PAGE (conditions non dénaturantes) à partir de lysats de peaux psoriasiques ou de kératinocytes. L’expression de POMP était modulée avec des ARN interférents dans des lignées kératinocytaires HaCat. Nous montrons que tous les sous-types (30S, 26S, 20S) et tous les régulateurs (PA28, PA200, 19S) des protéasomes sont surexprimés dans la peau lésée où les protéasomes 26S sont proportionnellement plus augmentés que les 20S (en faveur d’un assemblage accru) et où POMP est surexprimée et préférentiellement associée à des précurseurs du protéasome. L’inhibition de POMP a un effet anti-proliferatif et pro-apoptotique via l’inhibition de l’assemblage du protéasome dans les lignées kératinocytaires. Cette inhibition diminue l’expression de keratine 10 et d’involucrine au cours de la différenciation calcium-induite des HaCat. Ces résultats indiquent que l’augmentation de l’assemblage, de l’activité et de la quantité des protéasomes dans l’épiderme psoriasique lésé est liée (au moins en partie) à la surexpression de POMP. De plus, POMP peut réguler la prolifération et la différenciation kératinocytaire. Un ciblage thérapeutique de POMP en agissant à la fois sur les protéasomes et les immunoprotéasomes aurait une action anti-inflammatoire et anti-proliférative potentiellement intéressante et adaptée au traitement du psoriasis.

The multifunctional protein E4 transcription factor 1 (E4F1) is an essential regulator of epidermal stem cell (ESC) maintenance. Here, we found that E4F1 transcriptionally regulates a metabolic program involved in pyruvate metabolism that is required to maintain skin homeostasis. E4F1 deficiency in basal keratinocytes resulted in deregulated expression of dihydrolipoamide acetyltransferase (Dlat), a gene encoding the E2 subunit of the mitochondrial pyruvate dehydrogenase (PDH) complex. Accordingly, E4f1 knock-out (KO) keratinocytes exhibited impaired PDH activity and a redirection of the glycolytic flux toward lactate production. The metabolic reprogramming of E4f1 KO keratinocytes associated with remodeling of their microenvironment and alterations of the basement membrane, led to ESC mislocalization and exhaustion of the ESC pool. ShRNA-mediated depletion of Dlat in primary keratinocytes recapitulated defects observed upon E4f1 inactivation, including increased lactate secretion, enhanced activity of extracellular matrix remodeling enzymes, and impaired clonogenic potential. Altogether, our data reveal a central role for Dlat in the metabolic program regulated by E4F1 in basal keratinocytes and illustrate the importance of PDH activity in skin homeostasis.