bims-pideca 2025-12-07 papers

bims-pideca

Biomed News

on Class IA PI3K signalling in development and cancer

Issue of 2025–12–07
twenty-six papers selected by
Ralitsa Radostinova Madsen, MRC-PPU

Androgen deprivation-mediated activation of AKT is enhanced in prostate cancer with TMPRSS2:ERG fusion.
Epithelial-mesenchymal cell state heterogeneity predetermines differential phospho-signaling responses to EGF stimulation.
Analysis of clinical, single cell, and spatial data from the Human Tumor Atlas Network (HTAN) with massively distributed cloud-based queries.
Reprogramming of cholesterol sensing in epithelial cells supports pancreatic inflammation.
A genome-scale single-cell CRISPRi map of trans gene regulation across human pluripotent stem cell lines.
mTORC1 signaling in group 2 innate lymphoid cells coordinates neuro-immune crosstalk in allergic lung inflammation.
Reconstructing Waddington's landscape from data.
EGFR suppression and drug-induced potentiation are widespread features of oncogenic RTK fusions.
Implications of genetic and epigenetic aberrations to the tumorigenicity of human pluripotent stem cells.
Analyzing qPCR data: Better practices to facilitate rigor and reproducibility.
Regulation of Insulin Transcytosis Across Endothelium in Metabolic Health and Disease.
PixlMap: A generalisable pixel classifier for cellular phenotyping in multiplex immunofluorescence images.
Joint profiling of cell morphology and gene expression during in vitro neurodevelopment.
Optimizing clinical monitoring and management guidelines for capivasertib in HR-positive/HER2-negative advanced breast cancer: expert opinion.
ERK builds a population of short-lived nascent adhesions that produce persistent edge protrusion and cell migration.
Standardized metrics for assessment and reproducibility of imaging-based spatial transcriptomics datasets.
PTPRJ-Targeting Peptide Agonist Induces Broad Cellular Signaling Perturbations and DNA Damage in Lung Cancer Cells.
mTOR signaling during T cell activation promotes cytokine production in T cells through 3' UTR-mediated translation control.
MCL1 modulates mTORC1 signaling to promote bioenergetics and tumorigenesis.
A guide to transcriptomic deconvolution in cancer.
A unified genetic perturbation language for human cellular programming.
Harnessing liquid biopsy to unveil RAS-MEK pathway somatic pathogenic variants in extracranial arterio-venous malformations.
Reversibility, regulation, and the community of development: the legacy of Sir John B. Gurdon.
KSR1 is a scaffold for the Hippo signaling pathway.
Multilayered regulation of TORC1 signaling by Ait1, Gcn2, and SEAC/GATOR during nitrogen limitation and starvation.
Altered PI3K/mTOR signaling within the forebrain leads to respiratory deficits in a mouse model of epilepsy.

J Clin Invest. 2025 Dec 01. pii: e192368. [Epub ahead of print]135(23):

Androgen deprivation-mediated activation of AKT is enhanced in prostate cancer with TMPRSS2:ERG fusion.

Fen Ma, Sen Chen, Luigi Cecchi, Betul Ersoy-Fazlioglu, Joshua W Russo, Seiji Arai, Seifeldin Awad, Carla Calagua, Fang Xie, Larysa Poluben, Olga Voznesensky, Anson T Ku, Fatima Karzai, Changmeng Cai, David J Einstein, Huihui Ye, Xin Yuan, Alex Toker, Mary-Ellen Taplin, Adam G Sowalsky, Steven P Balk.

  TMPRSS2:ERG gene fusion (T:E fusion) in prostate adenocarcinoma (PCa) puts ERG under androgen receptor-regulated (AR-regulated) TMPRSS2 expression. T:E fusion is associated with PTEN loss and is highly associated with decreased INPP4B expression, which together may compensate for ERG-mediated suppression of AKT signaling. We confirmed in PCa cells and a mouse PCa model that ERG suppresses IRS2 and AKT activation. In contrast, ERG downregulation did not increase INPP4B, suggesting its decrease is indirect and reflects selective pressure to suppress INPP4B function. Notably, INPP4B expression was decreased in PTEN-intact and PTEN-deficient T:E fusion tumors, suggesting selection for a nonredundant function. As ERG in T:E fusion tumors is AR regulated, we further assessed whether AR inhibition increases AKT activity in T:E fusion tumors. A T:E fusion-positive PDX had increased AKT activity in vivo and response to AKT inhibition in vitro after androgen deprivation. Moreover, two clinical trials of neoadjuvant AR inhibition prior to radical prostatectomy showed greater increases in AKT activation in the T:E fusion-positive versus -negative tumors. These findings indicate that AKT activation may mitigate the efficacy of AR-targeted therapy in T:E fusion PCa and that these patients may most benefit from combination therapy targeting AR and AKT.

Keywords:  Cell biology; Inositol phosphates; Oncogenes; Oncology; Prostate cancer

DOI:  https://doi.org/10.1172/JCI192368
Mol Biol Cell. 2025 Dec 03. mbcE25070349

Epithelial-mesenchymal cell state heterogeneity predetermines differential phospho-signaling responses to EGF stimulation.

Felix V Kohane, Chantelle Johnstone, Daniel P Neumann, Ihuan Gunawan, Tim Huang, Fatemeh Vafaee, Christine L Chaffer, John G Lock.

Understanding why isogenic cancer cells respond differently to equivalent oncogenic stimuli is vital for optimizing anticancer therapies. Emerging evidence suggests that pre-existing differences in cell state may modulate signaling responses to new stimuli, but the interplay of specific cell states and signals remains unclear. We investigated whether epithelial-mesenchymal (E/M) state, a major axis of cancer cell heterogeneity, influences signaling responses to epidermal growth factor (EGF), a critical oncogenic stimulus in non-small cell lung cancer (NSCLC). We imaged >64,000 A549 NSCLC cells labeled for DNA, F-actin and alternate signaling markers (p-AKT-S473, p-AKT-T308, p-ERK or p-S6) after acute stimulation. Quantitative single-cell morphological and spatial profiling defined a stimulus-invariant 'E/M state landscape' over which EGF signaling responses were compared. This revealed state-dependent differences in signal-activation magnitudes, dynamics and subcellular routing. AKT responses exhibited phosphosite- and compartment-specific dynamics across states, with epithelial cells showing strong, transient membrane-localized S473 and higher internalized T308, whereas mesenchymal cells displayed weaker but sustained nuclear and ruffle-localized S473. Regression-based computational multiplexing concurrently inferred all signaling responses per cell, mapping state-dependent divergence in multi-molecular signaling trajectories. E/M state thus predetermines distinctive spatiotemporal profiles of EGF-induced signaling, with implications for signaling functions and anti-signaling therapy responses across E/M state-diverse tumors.

DOI: https://doi.org/10.1091/mbc.E25-07-0349
Res Sq. 2025 Nov 17. pii: rs.3.rs-7769205. [Epub ahead of print]

Analysis of clinical, single cell, and spatial data from the Human Tumor Atlas Network (HTAN) with massively distributed cloud-based queries.

David Gibbs, Dar'ya Pozhidayeva, Yamina Katariya, Boris Aguilar, Kristen Anton, Clarisse Lau, William Longabaugh, Ino de Bruijn, Alex Lash, Milen Nikolov, Jennifer Altreuter, Ashley Clayton, Aditi Gopalan, Adam Taylor, Nikolaus Schultz, Ethan Cerami, Vésteinn Thorsson.

Cancer research increasingly relies on large-scale, multimodal datasets that capture the complexity of tumor ecosystems across diverse patients, cancer types, and disease stages. The Human Tumor Atlas Network (HTAN) generates such data, including single-cell transcriptomics, proteomics, and multiplexed imaging. However, the volume and heterogeneity of the data present challenges for researchers seeking to integrate, explore, and analyze these datasets at scale. To this end, HTAN developed a cloud-based infrastructure that transforms clinical and assay metadata into aggregate Google BigQuery tables, hosted through the Institute for Systems Biology Cancer Gateway in the Cloud (ISB-CGC). This infrastructure introduces two key innovations: (1) a provenance-based HTAN ID table that simplifies cohort construction and cross-assay integration, and (2) the novel adaptation of BigQuery's geospatial functions for use in spatial biology, enabling neighborhood and correlation analysis of tumor microenvironments. We demonstrate these capabilities through R and Python notebooks that highlight use cases such as identifying precancer and organ-specific sample cohorts, integrating multimodal datasets, and analyzing single-cell and spatial data. By lowering technical and computational barriers, this infrastructure provides a cost-effective and intuitive entry point for researchers, highlighting the potential of cloud-based platforms to accelerate cancer discoveries.

DOI: https://doi.org/10.21203/rs.3.rs-7769205/v1
Mol Metab. 2025 Dec 02. pii: S2212-8778(25)00199-1. [Epub ahead of print] 102292

Reprogramming of cholesterol sensing in epithelial cells supports pancreatic inflammation.

Giulia Milan, Olga A Mareninova, Marco Fantuz, Martina Spacci, Carlotta Paoli, Jerik A Pineda, Roberta Noè, Beatrice Calciolari, Roberto Zoncu, Anna S Gukovskaya, Alessandro Carrer.

  Pancreatitis is a common cause of hospitalization that necessitates attentive clinical management. Affected individuals are at risk for pancreatic cancer due to aberrant signaling and empowered cell plasticity. Yet, molecular and cellular dynamics that govern epithelial cell behavior in response to inflammation remain largely elusive. Here we found that inflammation induces Endoplasmic Reticulum-Associated Degradation protein (ERAD)-mediated downregulation of Niemann-Pick type C protein 1 (NPC1), which leads to the sequestration of free cholesterol within acinar cells' lysosomes. Reducing intra-pancreatic cholesterol levels through genetic ablation of Acly ameliorates cerulein-induced pancreatitis, while pharmacological targeting of NPC1 exacerbates tissue damage. Mechanistically, the accumulation of lysosomal cholesterol is sensed by the mechanistic Target of Rapamycin Complex 1 (mTORC1) that promotes metaplasia of pancreatic acinar cells, an event commonly associated to pancreatitis and tissue regeneration. Indeed, cholesterol supplementation or NPC1 inhibition facilitate acinar-to-ductal metaplasia (ADM) both ex vivo and in vivo, in an mTORC1-dependent manner. These results identify a metabolic/signaling axis driving the reprogramming of pancreatic epithelial cells in response to inflammation. This hinges on a nutrient sensing paradigm, previously documented exclusively in pathological conditions.

Keywords:  acinar-to-ductal metaplasia (ADM); cholesterol; lysosome; mTORC1; pancreatitis

DOI:  https://doi.org/10.1016/j.molmet.2025.102292
Cell Genom. 2025 Dec 01. pii: S2666-979X(25)00332-5. [Epub ahead of print] 101076

A genome-scale single-cell CRISPRi map of trans gene regulation across human pluripotent stem cell lines.

Claudia Feng, Elin Madli Peets, Yan Zhou, Luca Crepaldi, Sunay Usluer, Alistair Dunham, Jana M Braunger, Jing Su, Magdalena E Strauss, Daniele Muraro, Kimberly Ai Xian Cheam, Marc Jan Bonder, Edgar Garriga Nogales, Sarah Cooper, Andrew Bassett, Steven Leonard, Yong Gu, Bo Fussing, David Burke, Leopold Parts, Oliver Stegle, Britta Velten.

  Population-scale resources of genetic, molecular, and cellular information form the basis for understanding human genomes, charting the heritable basis of disease and tracing the effects of mutations. Pooled perturbation assays, probing the effect of many perturbations coupled with single-cell RNA sequencing (scRNA-seq) readout, are especially potent references for interpreting disease-linked mutations or gene-expression changes. However, the utility of existing maps has been limited by the comprehensiveness of perturbations conducted and the relevance of their cell-line context. Here, we present a genome-scale CRISPR interference perturbation map with scRNA-seq readout across many genetic backgrounds in human pluripotent cells. We map trans expression changes induced by knockdowns and characterize their variation across donors, with expression quantitative trait loci linked to higher genetic modulation of perturbation effects. This study pioneers population-scale CRISPR perturbations with high-dimensional readouts, which will fuel the future of effective modulation of cellular disease phenotypes.

Keywords:  CRISPR; CRISPRi; CROP-seq; Perturb-seq; eQTL; genome-scale Perturb-seq; human induced pluripotent stem cells; iPSCs; scRNA-seq

DOI:  https://doi.org/10.1016/j.xgen.2025.101076
Nat Commun. 2025 Nov 29.

mTORC1 signaling in group 2 innate lymphoid cells coordinates neuro-immune crosstalk in allergic lung inflammation.

Dongdi Wang, Lin Hu, Jinyu Chen, Jinxin Qiu, Yue Chen, Michelle Zhang, Linfeng Zhao, Xiaohui Su, Jiping Sun, Ju Qiu, Wei Tang, Wenyong Zhou, Lei Shen.

Group 2 innate lymphoid cells (ILC2) initiate pathologic type 2 inflammation in allergic asthma in response to diverse tissue-derived stimuli. However, the molecular mechanisms by which ILC2 cells integrate and respond to environmental signals are unclear. Here, we show in a mouse model that in allergic asthma, mechanistic target of rapamycin complex 1 (mTORC1) activation in lung ILC2 cells increases. Genetic ablation of Raptor, an obligatory component of mTORC1 complex, results in reduced IL-5 and IL-13 production in ILC2 cells and protects mice from allergic inflammation. Pharmacological inhibition of mTORC1 by rapamycin suppresses ILC2 activation and ameliorates allergic lung inflammation. Mechanistically, mTORC1 activation upregulates neuromedin U receptor 1 (NMUR1) expression through epigenetic reprogramming, which augments ILC2 activation in response to neuromedin U (NMU). However, our experiments suggest that NMUR1 is not an exclusive mediator of ILC2 activation downstream of mTORC1. In conclusion, our work reveals that in ILC2s, mTORC1 signaling coordinates neuro-immune crosstalk for optimal activation, and highlights mTORC1 as a potential therapeutic target for allergic asthma.

DOI: https://doi.org/10.1038/s41467-025-66683-y
Proc Natl Acad Sci U S A. 2025 Dec 09. 122(49): e2521762122

Reconstructing Waddington's landscape from data.

Dillon J Cislo, M Joaquina Delás, James Briscoe, Eric D Siggia.

  The development of a zygote into a functional organism requires that this single progenitor cell gives rise to numerous distinct cell types. Attempts to exhaustively tabulate the interactions within developmental signaling networks that coordinate these hierarchical cell fate transitions are difficult to interpret or fit to data. An alternative approach models the cellular decision-making process as a flow in an abstract landscape whose signal-dependent topography defines the possible developmental outcomes and the transitions between them. Prior applications of this formalism have built landscapes in low-dimensional spaces without explicit maps to gene expression. Here, we present a computational geometry framework for fitting dynamical landscapes directly to high-dimensional single-cell data. Our method models the time evolution of probability distributions in gene expression space, enabling landscape construction with minimal free parameters and precise characterization of dynamical features, including fixed points, unstable manifolds, and basins of attraction. We demonstrate the applicability of this framework to multicolor flow-cytometry and RNA-seq data. Applied to a stem cell system that models ventral neural tube patterning, we recover a family of morphogen-dependent landscapes whose valleys align with canonical neural progenitor types. Remarkably, simple linear interpolation between landscapes captures signaling dependence, and chaining landscapes together reveals irreversible behavior following transient morphogen exposure. Our method combines the interpretability of landscape models with a direct connection to data, providing a general framework for understanding and controlling developmental dynamics.

Keywords:  Morse–Smale; Waddington landscape; dimensional reduction; transcription profiling

DOI:  https://doi.org/10.1073/pnas.2521762122
bioRxiv. 2025 Nov 20. pii: 2025.11.19.689362. [Epub ahead of print]

EGFR suppression and drug-induced potentiation are widespread features of oncogenic RTK fusions.

Yuzhi Carol Gao, David Gonzalez-Martinez, Sofia Wissert, Hana Bader, Nidhi Sahni, Anh Le, Robert C Doebele, Lukasz J Bugaj.

Regulation of cancer cells by their environment contributes to tumorigenesis and drug response, though the extent to which the oncogenic state can alter a cell's perception of its environment is not clear. Prior studies found that EML4-ALK, a receptor tyrosine kinase (RTK) fusion oncoprotein, suppresses transmembrane receptor signaling through EGFR. Moreover, suppression was reversed with targeted ALK inhibition, thereby promoting survival and drug tolerance. Here we tested whether such modulation of EGFR was common among other RTK fusions, which collectively are found in ∼5% of all cancers. Using live- and fixed-cell microscopy in isogenic and patient-derived cell lines, we found that a wide variety of RTK fusions suppress transmembrane EGFR and sequester essential adaptor proteins in the cytoplasm, as evidenced by the localization of endogenous Grb2. Targeted therapies rapidly released Grb2 from sequestration and potentiated EGFR. Synthetic optogenetic analogs of RTK fusions confirmed that cytoplasmic sequestration of Grb2 was sufficient to suppress perception of extracellular EGF and could do so without driving signaling from the synthetic fusion itself, demonstrating that fusion signaling and suppression of EGFR could be functionally decoupled. Our study uncovers that a large number of RTK fusions simultaneously act as both activators and suppressors of signaling, the mechanisms of which could be exploited for new biomimetic therapies that enhance cell killing and suppress drug tolerance.

DOI: https://doi.org/10.1101/2025.11.19.689362
Adv Drug Deliv Rev. 2025 Nov 29. pii: S0169-409X(25)00228-5. [Epub ahead of print]228 115743

Implications of genetic and epigenetic aberrations to the tumorigenicity of human pluripotent stem cells.

Gal Keshet, Ivana Barbaric, Nissim Benvenisty.

  Human pluripotent stem cells (hPSCs) hold immense promise for cell replacement therapies due to their capacity to give rise to derivatives of the three embryonic germ layers and their ability to divide indefinitely in culture. Since their first derivation less than 30 years ago, multiple hPSC-derived cell products are already in clinical trials for a range of pathologies. Nevertheless, hPSCs also possess an intrinsic tumorigenic potential and have been shown to acquire recurrent genetic and epigenetic aberrations strongly associated with cancer initiation and progression. These properties cast doubt on the safety of hPSCs and raise concerns regarding their use for transplantation. In this review, we summarize the different kinds of genetic and epigenetic abnormalities repeatedly observed in hPSCs, how they emerge, and their potential implications for the tumorigenicity of hPSC-based products. We also discuss shared and unique abnormalities found in hPSCs derived from different sources. Finally, we suggest possible methods for reducing the occurrence of these aberrations and managing their effects once they arise.

Keywords:  Epigenetic aberrations; Genetic aberrations; Human pluripotent stem cells; Tumorigenecity

DOI:  https://doi.org/10.1016/j.addr.2025.115743
Biochem Biophys Rep. 2025 Dec;44 102356

Analyzing qPCR data: Better practices to facilitate rigor and reproducibility.

Thomas H Hampton, Lily Taub, Kiyoshi Ferreria-Fukutani, Bruce A Stanton, Todd A MacKenzie.

  Quantitative PCR (qPCR) remains a widely used, cost-effective method for RNA quantitation, yet many published studies inadequately comply with MIQE (Minimum Information for Publication of Quantitative Real-Time PCR Experiments) and FAIR (Findable, Accessible, Interoperable, Reproducible) data principles. Widespread reliance on the 2-ΔΔCT method often overlooks critical factors such as amplification efficiency variability and reference gene stability. Furthermore, the absence of raw data and analysis code limits the community's ability to evaluate potential biases and reproduce findings. Here, we primarily aim to encourage researchers to share raw qPCR fluorescence data along with detailed analysis scripts that start from raw input and produce final figures and statistical tests. Using our recently published dataset, we model the complete qPCR analytical workflow-from raw fluorescence curves through to differential expression-highlighting key decision points that can influence results. We provide fully documented R scripts illustrating how ANCOVA (Analysis of Covariance), a flexible multivariable linear modeling approach, generally offers greater statistical power and robustness compared to 2-ΔΔCT. Additionally, simulations support ANCOVA's applicability across diverse experimental conditions. We also demonstrate how general-purpose data repositories (e.g., figshare) and code repositories (e.g., GitHub) facilitate adherence to FAIR principles and promote transparency in qPCR research. Finally, we offer graphical examples that transparently depict both target and reference gene behavior within the same figure, enhancing interpretability. This work establishes practical resources and conceptual foundations to improve rigor, reproducibility, and openness in qPCR data analysis.Image 1.

Keywords:  Delta-delta CT; FAIR; MIQE; RDML; Reproducibility; Rigor; qPCR

DOI:  https://doi.org/10.1016/j.bbrep.2025.102356
Arterioscler Thromb Vasc Biol. 2025 Dec 04.

Regulation of Insulin Transcytosis Across Endothelium in Metabolic Health and Disease.

Chieko Mineo, Philip W Shaul.

  The delivery of insulin to the skeletal muscle has a major influence on glucose disposal in muscle, where 80% of total body glucose disposal occurs. The skeletal muscle microvascular endothelial cells play a critical role in peripheral insulin sensitivity through their regulation of insulin delivery. Recent advancements in methodologies have provided in-depth views of the molecular mechanisms by which the endothelial cells regulate the delivery process. However, how the cellular machinery is modulated under physiological or pathological conditions remains largely unexplored. Conditions with estrogen deficiency and obesity are 2 situations that are closely associated with peripheral insulin resistance and type 2 diabetes in humans. It is of great interest to determine whether and how endothelial control of insulin delivery impacts the development of metabolic dysregulation under these and other conditions. This review aims to provide an overview of the molecular mechanisms governing insulin delivery to the skeletal muscle. The available evidence will be presented that the transcytosis of insulin across the endothelial cell monolayer in skeletal muscle plays a critical role in muscle insulin delivery, thereby having a major impact on overall glucose homeostasis. In vivo investigations with manipulation of mechanisms in endothelial cells will be summarized, and the current knowledge gaps will be presented. Interrogation of the role of the endothelium in insulin transport provides a paradigm in which insights are being gained about cellular actions of insulin, molecular transport by endothelial cells, and the intricacies of glucose homeostasis.

Keywords:  endothelial cells; endothelium, vascular; hyperglycemia; insulin; muscle fibers, skeletal

DOI:  https://doi.org/10.1161/ATVBAHA.125.321545
PLoS One. 2025 ;20(12): e0317865

PixlMap: A generalisable pixel classifier for cellular phenotyping in multiplex immunofluorescence images.

Rachel L Baird, David Mason, Kai Rakovic, Fiona Ballantyne, Ian R Powley, Anastasia Georgakopoulou, Lucy Hillary, Thomas G Bird, Leah Officer-Jones, John Le Quesne.

Multiplexed methods for the detection of protein expression generate extremely data-rich images of intact tissue sections. These images are invaluable for the quantification and analysis of complex biology and biomarker development. However, their interpretation presents a considerable analytical challenge. Cell segmentation from images is a key bottleneck and a major focus of research activity in artificial intelligence. Most current methods depend initially on the use of a nuclear counterstain to identify nuclear boundaries, which is a relatively straightforward task. The cellular boundary is then assigned either by expansion of the nuclear outline, or by the use of membrane or cytoplasm-specific stains to delineate cell boundaries, or by some combination of the two. The task is critical, as inaccurate segmentation leads to information loss and data contamination from neighbouring cells. Increasingly sophisticated methods are being developed to address these issues, but each has its own shortcomings. We present an alternative method which is inspired by the fact that the assignation of a cellular phenotype 'by eye' does not depend upon the accurate identification of cell boundaries. We present an easy-to-use deep learning-based cellular phenotyping method which leverages this human capacity to assign phenotypes without segmenting the entire cell, and which can accurately phenotype cells based on nuclear segmentation alone. Using human ground truth annotations of entire cellular regions, we developed a classifier leveraging the U-Net architecture within a commercially available deep learning image analysis platform, but the principle is transferrable to any deep-learning framework. Crucially, training requires only a single example of each compartmental stain (nuclear/cytoplasmic/membranous). The resulting algorithm assigns class identities to cells with nuclear labelling alone, without the need for whole cell expansion. The method is highly novel, broadly generalisable, and comparable in accuracy to intensity-based phenotyping methods, bridging the gap between inaccurate cellular segmentation and accurate phenotype generation.

DOI: https://doi.org/10.1371/journal.pone.0317865
Elife. 2025 Dec 01. pii: e102578. [Epub ahead of print]14

Joint profiling of cell morphology and gene expression during in vitro neurodevelopment.

Adithi Sundaresh, Dimitri Meistermann, Riina Lampela, Zhiyu Yang, Rosa Woldegebriel, Andrea Ganna, Pau Puigdevall Costa, Helena Kilpinen.

  Differentiation of induced pluripotent stem cells (iPSCs) toward neuronal lineages has enabled diverse cellular models of human neurodevelopment and related disorders. Here, we jointly profiled neuronal morphology and gene expression at single-cell resolution across 60,000 iPSC-derived cortical neurons at three developmental time points with Cell Painting (CP) and single-cell RNA-sequencing (scRNA-seq). By modeling the relationship between morphological features and gene expression within our differentiation system, we annotated image-based features with biological functions and showed that while CP resolves broader neuronal classes than scRNA-seq, it complements transcriptomic data by quantifying the biological processes that drive neuronal differentiation over time, such as mitochondrial function and cell cycle. Further, we found that while over 60% of the cells resembled those seen in the fetal brain, 28% represented metabolically abnormal cell states and broader neuronal classes specific to in vitro cells. We show that iPSC-derived cortical neurons are nonetheless a relevant model for a range of brain-related complex traits, including schizophrenia and bipolar disorder, and that disease heritability can also be captured in the morphological feature space. Finally, we applied CP to iPSC-derived neural progenitors from patients with Kabuki syndrome, revealing morphological signatures of precocious differentiation and altered cell cycling. These results highlight the potential of multi-modal single-cell characterization to reveal complementary and disease-relevant cellular and molecular phenotypes.

Keywords:  cell painting; developmental disorders; genetics; genomics; human; iPSC; neurodevelopment; regenerative medicine; single-cell genomics; stem cells; transcriptomics

DOI:  https://doi.org/10.7554/eLife.102578
NPJ Breast Cancer. 2025 Dec 04.

Optimizing clinical monitoring and management guidelines for capivasertib in HR-positive/HER2-negative advanced breast cancer: expert opinion.

Neil M Iyengar, Joyce A O'Shaughnessy, Heather B Moore, Erika Hamilton, Komal L Jhaveri, Cynthia X Ma, Karen Tedesco, Monali Vasekar, Sarah Donahue, Hope S Rugo.

Capivasertib, an AKT inhibitor, approved in combination with fulvestrant for hormone receptor-positive, human epidermal growth factor receptor 2-negative advanced breast cancer with ≥1 PIK3CA, AKT1, and/or PTEN alterations, significantly improved progression-free survival in the CAPItello-291 phase 3 trial. However, capivasertib-associated adverse events of diarrhea, rash, and hyperglycemia may require proactive management. This article provides practical recommendations to support prevention and early intervention to optimize adherence and treatment outcomes.

DOI: https://doi.org/10.1038/s41523-025-00864-2
bioRxiv. 2025 Nov 17. pii: 2025.11.17.688923. [Epub ahead of print]

ERK builds a population of short-lived nascent adhesions that produce persistent edge protrusion and cell migration.

Andrew P Shepherd, Keith R Carney, Andrew Elliott, Sangyoon J Han, Michelle C Mendoza.

Cell migration is realized through the fast and persistent protrusion of a leading edge in the direction of movement. The actin and adhesion structures that build edge protrusions are integrated such that pro-migration signaling pathways must control both assemblies to induce protrusion. Understanding the contribution of adhesion regulation has been complicated by the inability to selectively assay the nascent adhesions that promote edge protrusion. Here, we dissect how the core RAS→RAF→MEK→ERK pathway's control of nascent adhesions contributes to edge protrusion and cell migration by targeting an ERK FRET biosensor to adhesions and quantifying ERK's spatial and temporal activity. We find that ERK is activated in the assembling, membrane-proximal region of nascent adhesions through adhesion scaffold paxillin, which interacts with the ERK activator MEK. Tracking nascent adhesion dynamics during cell migration showed that ERK promotes both nascent adhesion assembly and disassembly to create a population of nascent adhesions with short lifetimes. MEK inhibition is partially complemented by expression of a talin R8vvv mutant that increases the nascent adhesion population, demonstrating the significance of ERK's adhesion regulation for edge protrusion and migration persistence. These findings suggest that when new adhesions initiate, the ERK activation level dictates adhesion assembly and disassembly rates to specifically build nascent adhesions that rapidly turnover, an adhesion population that promotes protrusion persistence and migration.
Significance: Cell migration is essential for development, healing, and cancer spread. To move, cells need to build and break tiny structures called nascent adhesions, specifically at their edge that protrudes in the direction of movement. We discovered that ERK, a key signaling protein activated during development and cancer, helps create a fast-turning-over population of these adhesions, which supports steady movement. Using new biosensors and adhesion tracking, we showed ERK is active in the assembling part of the adhesion, where it promotes both assembly and disassembly. Increasing the nascent adhesion population helps cells move when ERK is blocked. This work reveals how ERK controls cell movement by balancing adhesion dynamics and introduces tools to study similar processes in other signaling pathways.

DOI: https://doi.org/10.1101/2025.11.17.688923
Nat Biotechnol. 2025 Dec 03.

Standardized metrics for assessment and reproducibility of imaging-based spatial transcriptomics datasets.

Jasmine T Plummer, Felipe Segato Dezem, David P Cook, Jiwoon Park, Luke Zhang, Yutian Liu, Maycon Marção, Hannah DuBose, Arjumand Wani, Kellie Wise, Michael Roach, Kate Harvey, Taopeng Wang, Kirk B Jensen, Natalia Morosini, Roberto De Gregorio, Alicia Alonso, Shauna Lee Houlihan, Robert E Schwartz, Erika Hissong, Catherine Snopkowski, Jeffrey L Wrana, Natalie Ryan, Lisa M Butler, George Church, Alexander Swarbrick, Christopher E Mason, Luciano G Martelotto.

Spatial transcriptomics lacks standardized metrics for evaluating imaging-based in situ hybridization technologies across sites. In this study, we generated the Spatial Touchstone (ST) dataset from six tissue types across several global sites with centralized sectioning, analyzed on both Xenium and CosMx platforms. These platforms were selected for their widespread use and distinct chemistries. We assessed reproducibility, sensitivity, dynamic ranges, signal-to-noise ratio, false discovery rates, cell type annotation and congruence with single-cell profiling. This study offers ST standardized operating procedures (STSOPs) and an open-source software, SpatialQM, enabling evaluation of samples across all technical metrics and direct imputation of cell annotations. The generated imaging-based spatial transcriptomics data repository comprises 254 spatial profiles, incorporating both public and newly generated ST datasets in a web-based application, which enables analysis and comparison of user data against an extensive collection of imaging-based datasets. Finally, we establish best practices and metrics to evaluate and integrate imaging-based multi-omics data from single cells into spatial transcriptomics to spatial proteomics.

DOI: https://doi.org/10.1038/s41587-025-02811-9
bioRxiv. 2025 Nov 17. pii: 2025.11.17.688709. [Epub ahead of print]

PTPRJ-Targeting Peptide Agonist Induces Broad Cellular Signaling Perturbations and DNA Damage in Lung Cancer Cells.

Sophie Rizzo, William S Hart, Rachel L Fetter, Bokai Song, Forest M White, Matthew J Lazzara, Damien Thévenin.

Receptor protein tyrosine phosphatases (RPTPs) are key regulators of cell signaling. However, their study and therapeutic targeting have been limited by the lack of known natural ligands or selective agonists, as well as an incomplete understanding of their structure-function relationships. Nonetheless, receptor homodimerization has been shown to suppress RPTP catalytic activity by restricting substrate access, offering a promising strategy for examining and modulating their function. Our previous work on PTPRJ, a member of the RPTP family, showed that its transmembrane domain regulates homodimerization, thereby controlling access to receptor tyrosine kinase (RTK) substrates and their phosphorylation levels. We also developed peptides that disrupt this dimerization, thereby inhibiting RTK phosphorylation and reducing cancer cell migration. These peptides were then engineered for selective, pH-sensitive insertion into the acidic tumor microenvironment to enhance efficacy while limiting off-target effects. Yet how broadly PTPRJ activation reshapes the phosphotyrosine landscape and whether those changes yield coherent cellular phenotypes remains unclear. In this study, we employed tyrosine phosphoproteomics, immunoblotting, immunofluorescence, and functional assays to assess the global impact of our lead peptide candidate, Hybrid 7, in A549 lung cancer cells that endogenously express PTPRJ. We find that Hybrid 7 decreases EGFR phosphorylation and selectively reduces phosphorylation across additional RTKs and motility adaptors, producing strong inhibition of EGF-driven migration and reduced proliferation. Hybrid 7 also elevates reactive oxygen species and DNA damage, and enforces CDK1-dependent G2/M arrest, indicating a primarily cytostatic, checkpoint-mediated response. These findings highlight the potential of RPTP-targeting peptides as valuable tools for dissecting RPTP function and as possible therapeutic agents capable of modulating key oncogenic pathways and inhibiting cancer progression.

DOI: https://doi.org/10.1101/2025.11.17.688709
Mol Cell. 2025 Dec 04. pii: S1097-2765(25)00901-3. [Epub ahead of print]85(23): 4452-4462.e5

mTOR signaling during T cell activation promotes cytokine production in T cells through 3' UTR-mediated translation control.

Anouk P Jurgens, Josephine Zwijnen, Antonia Bradarić, Floris P J van Alphen, Kaspar Bresser, Koos Rooijers, Arie J Hoogendijk, Branka Popović, Monika C Wolkers.

  T cells are key contributors to clearing our body of infected and malignant cells. During activation, T cells undergo profound translational alterations, and the evolutionarily and highly conserved kinase mammalian target of rapamycin (mTOR) is central in this process. It mediates T cell differentiation, homeostasis, and activation and promotes the production of pro-inflammatory cytokines. mTOR executes its translation activity through terminal oligopyrimidine (TOP) motifs located in the 5' untranslated region (5' UTR) of target genes. Here, we uncovered a distinct 3' UTR-mediated mechanism of mTOR signaling on cytokine production in T cells. Non-classical TOP motifs present in the cytokine 3' UTRs do not contribute to mTOR-mediated translation regulation. Rather, AU-rich elements (AREs) are required for mTOR-mediated cytokine production. Furthermore, we discovered that the RNA-binding protein DDX21 binds to 3' UTR AREs and confers mTOR-mediated translation control. In conclusion, we present a previously unappreciated ARE-dependent, 3' UTR-mediated mechanism that mTOR employs to regulate cytokine production.

Keywords:  3′ UTR; 3′ untranslated region; ARE-binding protein; DDX21; RBP; RNA-binding protein; T cell; cytokine regulation; mTOR; mammalian target of rapamycin; post-transcriptional regulation

DOI:  https://doi.org/10.1016/j.molcel.2025.11.005
Nat Commun. 2025 Dec 01. 16(1): 10841

MCL1 modulates mTORC1 signaling to promote bioenergetics and tumorigenesis.

Wentao Gui, Petr Paral, Bhavuk Dhamija, Eman Hagag, Martin Dusa, Jana Humajova, Pavla V Francova, Jan Kucka, Jan Pankrac, Caroline Schütz, Vasileios Armenis, Filippo Ferrucci, Mario Schubert, Kaomei Guan, Franziska Baenke, Daniel E Stange, Lorenz H Lehmann, Wolfram Weckwerth, Peter Mirtschink, Sofia Traikov, Belmonte Giuseppe, Clelia Miracco, Martin Bornhäuser, Saverio Minucci, Ludek Sefc, Libor Macurek, Mohamed Elgendy.

Myeloid cell leukemia-1 (MCL1) is among the most overexpressed proteins in tumors. MCL1 contributes to tumorigenesis by antagonizing apoptosis. However, apoptosis-unrelated functions are emerging. Screening an array of signaling switches identifies mTORC1 to be modulated by MCL1 but not by the anti-apoptotic Bcl-2 or Bcl-xL. mTORC1 is a central metabolic regulator. MCL1 impacts metabolism via modulating the expression of hexokinase 2 (HK2) in an mTORC1-dependent manner, which ultimately contributes to the tumor-promoting effects of MCL1. MCL1 inhibitors suppress mTORC1 in tumor cells but are associated with cardiotoxicity due to mTORC1 inhibition in the heart. Dietary leucine supplementation rescues mTORC1 signaling in the hearts of humanized Mcl-1 mice and greatly ameliorates the cardiotoxicity of MCL1 inhibitors. Taken together, here we describe tumor-promoting roles for MCL1 in regulating mTORC1 signaling and subsequently in bioenergetics, besides its role in antagonizing apoptosis, identifying MCL1 as a hinge of cell bioenergetics and survival.

DOI: https://doi.org/10.1038/s41467-025-66831-4
Nat Rev Cancer. 2025 Dec 02.

A guide to transcriptomic deconvolution in cancer.

Yaoyi Dai, Shuai Guo, Yidan Pan, Carla Castignani, Matthew D Montierth, Peter Van Loo, Wenyi Wang.

Cancer tissues are heterogeneous mixtures of tumour, stromal and immune cells, where each component comprises multiple distinct cell types and/or states. Mapping this heterogeneity and understanding the unique contributions of each cell type to the tumour transcriptome is crucial for advancing cancer biology, yet high-throughput expression profiles from tumour tissues only represent combined signals from all cellular sources. Computational deconvolution of these mixed signals has emerged as a powerful approach to dissect both cellular composition and cell-type-specific expression patterns. Here, we provide a comprehensive guide to transcriptomic deconvolution, specifically tailored for cancer researchers, presenting a systematic framework for selecting and applying deconvolution methods, considering the unique complexities of tumour tissues, data availability and method assumptions. We detail 43 deconvolution methods and outline how different approaches serve distinctive applications in cancer research: from understanding tumour-immune surveillance to identifying cancer subtypes, discovering prognostic biomarkers and characterizing spatial tumour architecture. By examining the capabilities and limitations of these methods, we highlight emerging trends and future directions, particularly in addressing tumour cell plasticity and dynamic cell states.

DOI: https://doi.org/10.1038/s41568-025-00886-9
bioRxiv. 2025 Nov 20. pii: 2025.11.20.689421. [Epub ahead of print]

A unified genetic perturbation language for human cellular programming.

Austin Hartman, Oliver Takacsi-Nagy, Courtney Kernick, Nicole E Theberath, Johnathan Lu, Lujing Wu, Michelle Mantilla, Siddhesh Mittra, Alison McClellan, Nicole Johnson, Lina Mohamad, Lesly Castillo-Colin, Farzad Hoque, Alexander Eapen, Andy Chen, Laura M Moser, Trini Rogando, Anabella Hernandez, Katherine Santostefano, Ansuman T Satpathy, Theodore L Roth.

Evolution simultaneously and combinatorially explores complex genetic changes across perturbation classes, including gene knockouts, knockdowns, overexpression, and the creation of new genes from existing domains. Separate technologies are capable of genetic perturbations at scale in human cells, but these methods are largely mutually incompatible. Here we present CRISPR-All, a unified genetic perturbation language for programming of any major type of genetic perturbation simultaneously, in any combination, at genome scale, in primary human cells. This is enabled by a standardized molecular architecture for each major perturbation class, development of a functional syntax for combining arbitrary numbers of elements across classes, and linkage to unique single cell compatible barcodes. To facilitate use, CRISPR-All converts high level descriptions of desired complex genetic changes into a single DNA sequence that can rewire genomic programs within a cell. Using the CRISPR-All language allowed for head-to-head functional comparisons across perturbation types in a comprehensive analysis of all previously identified genetic enhancements of human CAR-T cells. Combining CRISPR-All programs with single cell RNA sequencing revealed a greater diversity of phenotypic states, including improved functional performance, only accessible through distinct perturbation classes. Finally, CRISPR-All combinatorial genome scale screening of up to four distinct perturbations simultaneously revealed additive functional improvements in human T cells accessible only through iterative multiplexing of modifications across perturbation classes. CRISPR-All enables exploration of a combinatorial genetic perturbation space, which may be impactful for biological and clinical applications.

DOI: https://doi.org/10.1101/2025.11.20.689421
Commun Med (Lond). 2025 Dec 05. 5(1): 508

Harnessing liquid biopsy to unveil RAS-MEK pathway somatic pathogenic variants in extracranial arterio-venous malformations.

Kartik D Bhatia, Dianne Sylvester, Bhavna Padhye, Caroline Bateman, David Lord, Geoffrey McCowage, Luciano Dalla-Pozza, Timothy Harrington, Brendan Steinfort, Mark Dexter, Sarah Josephi-Taylor, Russell Dale, Smadar Kahana-Edwin.

BACKGROUND: Arterio-venous malformations (AVMs) have been associated with somatic genetic variants in the RAS-MEK pathway, generating interest in the role of MEK inhibitors. However, open biopsy for molecular characterisation carries a potentially life-threatening bleeding risk.
METHODS: We utilized liquid biopsy from the AVM efferent draining vein in 10 children and young adults (female 7, age range 7.3-22.2 years) with extracranial AVMs to identify the underlying somatic variants.
RESULTS: Here we show identification of somatic mosaic variants in 8 of 10 patients. There were no procedural complications. Two patients with spinal arteriovenous metameric syndrome (Cobb syndrome) demonstrated somatic activating KRAS variants. Two additional patients had somatic in-frame deletion-insertion variants of HRAS. Three patients had other variants involving the RAS-MEK pathway and one within PIK3CA. Five patients have commenced molecularly directed pharmacotherapy leading to reduced disability. Variant allele frequency was inversely correlated with sampling distance from the nidus. One patient who underwent sampling at two separate embolization sessions demonstrated a higher variant allele frequency in the sample obtained after embolization was completed.
CONCLUSIONS: Our study further supports the role of the RAS-MEK pathway in AVM pathophysiology. In addition, we demonstrate the effective use of liquid biopsy to genotype extracranial AVMs in children. Finally, we provide insights into how the localized high-pressure high-flow conditions within the AVM distinctly shape cell free DNA fragmentation patterns.

DOI: https://doi.org/10.1038/s43856-025-01174-1
Nat Commun. 2025 Dec 04. 16(1): 10899

Reversibility, regulation, and the community of development: the legacy of Sir John B. Gurdon.

Richard P Halley-Stott, Eva Hörmanseder, Jerome Jullien, Vincent Pasque.

John B. Gurdon approached biology with characteristic clarity and patience, asking not only what happens in development, but how much of a molecule is present, and for how long. His nuclear-transfer experiments revealed that cellular identity is not fixed but can be reset, reshaping both biology and medicine. Through studies of oocytes, translational control, and the community effect, he showed that stability in living systems arises from persistence and interaction. His influence reached far beyond the bench: through generosity and curiosity, he nurtured a community of scientists who, like their mentor, enjoy asking how a cell knows what to be.

DOI: https://doi.org/10.1038/s41467-025-67023-w
Commun Biol. 2025 Dec 01. 8(1): 1725

KSR1 is a scaffold for the Hippo signaling pathway.

Samar Sayedyahossein, Mohammed Rizwan Babu Sait, Zhigang Li, Riddhi Banerjee, Andy Tran, Louise Thines, Mehdi Karimi, Mehrnoosh Bahmani, Naomi Mishan, Pooya Borzou, Sergio A Hassan, David B Sacks.

The evolutionarily conserved Hippo signaling pathway regulates organ size and tissue homeostasis. Yes-associated protein (YAP) functions as a transcriptional co-activator and is a critical downstream effector of the Hippo signaling pathway. Altered crosstalk with oncogenic signaling pathways contributes to YAP dysregulation in cancer. Kinase Suppressor of Ras 1 (KSR1) scaffolds the Ras cascade. Some of the functions of the Ras and Hippo pathways in regulating cellular processes are similar. Nevertheless, the potential intersection of Ras and Hippo signaling has not been explored. Here, we identify KSR1 as a previously unrecognized scaffold of the Hippo pathway. We demonstrate that KSR1 constitutively binds to YAP and MST1 and forms a complex with LATS1. Moreover, KSR1 modulates YAP protein levels and its transcriptional activity, at least in part through the RhoA/actin axis. Our findings provide insight into the role of KSR1 as a scaffold of the Hippo signaling that could yield novel therapeutics.

DOI: https://doi.org/10.1038/s42003-025-09009-4
Nat Commun. 2025 Nov 29.

Multilayered regulation of TORC1 signaling by Ait1, Gcn2, and SEAC/GATOR during nitrogen limitation and starvation.

Cristina M Padilla, Jeaho Lim, Austin A Lipinski, Paul R Langlais, Andrew P Capaldi.

The Target of Rapamycin kinase Complex I (TORC1) is a central hub in the cell growth and metabolic control network of eukaryotes. How its upstream regulators cooperate to tune signaling across environmental conditions remains unclear. Here, we combine phosphoproteomics, TORC1 activity assays, and targeted genetic perturbations to dissect TORC1 regulation in Saccharomyces cerevisiae during transitions from a high-quality nitrogen source (glutamine) to a low-quality nitrogen source (proline), and on to complete nitrogen starvation. In proline medium, Ait1 and Gcn2 attenuate TORC1 activity, establishing a partially inhibited "Low Nitrogen Adaptive" state marked by extensive metabolic reprogramming without growth arrest. In contrast, during nitrogen starvation, SEAC, Ait1, and Gcn2 cooperate to drive TORC1 into a fully inhibited state, triggering widespread dephosphorylation of its downstream targets and entry into quiescence. Our results define a multilayered regulatory circuit that governs graded TORC1 control-a design likely conserved across eukaryotes.

DOI: https://doi.org/10.1038/s41467-025-66907-1
eNeuro. 2025 Dec 01. pii: ENEURO.0292-25.2025. [Epub ahead of print]

Altered PI3K/mTOR signaling within the forebrain leads to respiratory deficits in a mouse model of epilepsy.

Patrick Woller, Vamshidhar Singidi, McKenzie Rice, Durgesh Tiwari, Christina Gross, Steven A Crone.

People with epilepsy may experience sudden death due to respiratory failure through mechanisms that are currently not well understood. Epilepsy causing mutations are thought to elicit seizures due to altered function of forebrain circuits, yet breathing is controlled largely by the brainstem. To investigate how altered forebrain activity could impact breathing, we examined respiratory and seizure phenotypes in a mouse epilepsy model with a forebrain-specific deletion of the phosphatase and tensin homolog (PTEN) gene (PTEN-cKO). Using chronic diaphragm electromyography (EMG) and cortical electroencephalography (EEG), we monitored PTEN-cKO mice (6 males and 4 females) and control littermates (6 males and 3 females) continuously from pre-seizure onset through end-stage disease. PTEN-cKO mice develop spontaneous seizures that progress in frequency with age, accompanied by gradual changes in respiratory function, even during interictal periods. As seizure burden increases, PTEN-cKO mice experience an increased frequency of interictal apneas, slowing of respiratory rhythm, prolongation of inspiratory bursts, and elevation of inspiratory effort. All animals experienced a terminal apnea prior to cardiac arrest. These findings demonstrate that PTEN deletion in the forebrain disrupts the control of breathing and leads to terminal respiratory failure.Significance Statement Epilepsy typically alters forebrain function, but whether this alone is sufficient to alter the control of breathing, particularly in the absence of seizures, remains unclear. Here, we show in a spontaneous seizure model that forebrain-specific mutation of PTEN leads to progressive respiratory abnormalities initially manifesting as increased interictal apneas, then progressing to slower, deeper breathing and ultimately terminating in respiratory failure. These findings suggest that forebrain circuit dysfunction in epilepsy can disrupt respiratory control, potentially increasing the risk of sudden unexpected death in epilepsy (SUDEP).

DOI: https://doi.org/10.1523/ENEURO.0292-25.2025