bims-pideca Biomed News
on Class IA PI3K signalling in development and cancer
Issue of 2024–11–17
29 papers selected by
Ralitsa Radostinova Madsen, MRC-PPU
  1. Vertical inhibition of p110α/AKT and N-cadherin enhances treatment efficacy in PIK3CA-aberrated ovarian cancer cells.
  2. Parallel phosphoproteomics and metabolomics map the global metabolic tyrosine phosphoproteome.
  3. Low-affinity ligands of the epidermal growth factor receptor are long-range signal transmitters in collective cell migration of epithelial cells.
  4. Shear stress-stimulated AMPK couples endothelial cell mechanics, metabolism, and vasodilation.
  5. Bilateral regulation of EGFR activity and local PI(4,5)P2 dynamics in mammalian cells observed with superresolution microscopy.
  6. Directed evolution of engineered virus-like particles with improved production and transduction efficiencies.
  7. The cellular dogma.
  8. The Bi-Steric Inhibitor RMC-5552 Reduces mTORC1 Signaling and Growth in Lymphangioleiomyomatosis.
  9. Molecular Spies in Action: Genetically Encoded Fluorescent Biosensors Light up Cellular Signals.
  10. Engineered PsCas9 enables therapeutic genome editing in mouse liver with lipid nanoparticles.
  11. Transparency, bias, and reproducibility across science: a meta-research view.
  12. Bridging the Gap from Proteomics Technology to Clinical Application: Highlights from the 68th Benzon Foundation Symposium.
  13. Clinical functional proteomics of intercellular signalling in pancreatic cancer.
  14. A non-canonical role of somatic Cyclin D/CYD-1 in oogenesis and in maintenance of reproductive fidelity, dependent on the FOXO/DAF-16 activation state.
  15. Systems-level reconstruction of kinase phosphosignaling networks regulating endothelial barrier integrity using temporal data.
  16. Transcriptional regulators ensuring specific gene expression and decision-making at high TGFβ doses.
  17. The pro-oncogenic noncanonical activity of a RAS•GTP:RanGAP1 complex facilitates nuclear protein export.
  18. Protein phosphatase-1 regulates the binding of filamin C to FILIP1 in cultured skeletal muscle cells under mechanical stress.
  19. The massed-spaced learning effect in non-neural human cells.
  20. Proteomic and functional comparison between human induced and embryonic stem cells.
  21. Vascular lesions: Hemangioma or venous malformation?
  22. scPair: Boosting single cell multimodal analysis by leveraging implicit feature selection and single cell atlases.
  23. Rediscovering publicly available single-cell data with the DISCO platform.
  24. Enhanced Gαq Signaling in TSC2-deficient Cells Is Required for Their Neoplastic Behavior.
  25. Cultivating the next generation of leaders : How postdocs, principal investigators and institutes can nurture and select for leadership competencies.
  26. A comprehensive human embryo reference tool using single-cell RNA-sequencing data.
  27. TPC1 regulates melanoma tumourigenesis via mTORC1 and TFEB.
  28. Reconstructing the regulatory programs underlying the phenotypic plasticity of neural cancers.
  29. Unveiling RACK1: a key regulator of the PI3K/AKT pathway in prostate cancer development.
  1. Mol Oncol. 2024 Nov 14.
    Vertical inhibition of p110α/AKT and N-cadherin enhances treatment efficacy in PIK3CA-aberrated ovarian cancer cells.
    Shibo Zhang, Hei Ip Hong, Victor C Y Mak, Yuan Zhou, Yiling Lu, Guanglei Zhuang, Lydia W T Cheung.
      Phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit alpha [PIK3CA, encoding PI3Kalpha (also known as p110α)] is one of the most commonly aberrated genes in human cancers. In serous ovarian cancer, PIK3CA amplification is highly frequent but PIK3CA point mutation is rare. However, whether PIK3CA amplification and PIK3CA driver mutations have the same functional impact in the disease is unclear. Here, we report that both PIK3CA amplification and E545K mutation are tumorigenic. While the protein kinase B (AKT) signaling axis was activated in both E545K knock-in cells and PIK3CA-overexpressing cells, the mitogen-activated protein kinase 3/1 (ERK1/2) pathway was induced selectively by E545K mutation but not PIK3CA amplification. Intriguingly, AKT signaling in these PIK3CA-aberrated cells increased transcriptional coactivator YAP1 (YAP) Ser127 phosphorylation and thereby cytoplasmic YAP levels, which in turn increased cell migration through Ras-related C3 botulinum toxin substrate 1 (RAC1) activation. In addition to the altered YAP signaling, AKT upregulated N-cadherin expression, which also contributed to cell migration. Pharmacological inhibition of N-cadherin reduced cell migratory potential. Importantly, co-targeting N-cadherin and p110α/AKT caused additive reduction in cell migration in vitro and metastases formation in vivo. Together, this study reveals the molecular pathways driven by the PIK3CA aberrations and the exploitable vulnerabilities in PIK3CA-aberrated serous ovarian cancer cells.
    Keywords:  N‐cadherin; PI3K; PIK3CA; cytoplasmic YAP; p110α; serous ovarian cancer
    DOI:  https://doi.org/10.1002/1878-0261.13761
  2. Proc Natl Acad Sci U S A. 2024 Nov 19. 121(47): e2413837121
    Parallel phosphoproteomics and metabolomics map the global metabolic tyrosine phosphoproteome.
    Alissandra L Hillis, Tigist Tamir, Grace E Perry, John M Asara, Jared L Johnson, Tomer M Yaron, Lewis C Cantley, Forest M White, Alex Toker.
      Tyrosine phosphorylation of metabolic enzymes is an evolutionarily conserved posttranslational modification that facilitates rapid and reversible modulation of enzyme activity, localization, or function. Despite the high abundance of tyrosine phosphorylation events detected on metabolic enzymes in high-throughput mass spectrometry-based studies, functional characterization of tyrosine phosphorylation sites has been limited to a subset of enzymes. Since tyrosine phosphorylation is dysregulated across human diseases, including cancer, understanding the consequences of metabolic enzyme tyrosine phosphorylation events is critical for informing disease biology and therapeutic interventions. To globally identify metabolic enzyme tyrosine phosphorylation events and simultaneously assign functional significance to these sites, we performed parallel phosphoproteomics and polar metabolomics in nontumorigenic mammary epithelial cells (MCF10A) stimulated with epidermal growth factor (EGF) in the absence or presence of the EGF receptor inhibitor erlotinib. We performed an integrated analysis of the phosphoproteomic and metabolomic datasets to identify tyrosine phosphorylation sites on metabolic enzymes with functional consequences. We identified two previously characterized (pyruvate kinase muscle isozyme, phosphoglycerate mutase 1) and two uncharacterized (glutathione S-transferase Pi 1, glutamate dehydrogenase 1) tyrosine phosphorylation sites on metabolic enzymes with purported functions based on metabolomic analyses. We validated these hits using a doxycycline-inducible CRISPR interference system in MCF10A cells, in which target metabolic enzymes were depleted with simultaneous reexpression of wild-type, phosphomutant, or phosphomimetic isoforms. Together, these data provide a framework for identification, prioritization, and characterization of tyrosine phosphorylation sites on metabolic enzymes with functional significance.
    Keywords:  EGFR; cancer metabolism; metabolomics; phosphotyrosine; proteomics
    DOI:  https://doi.org/10.1073/pnas.2413837121
  3. Cell Rep. 2024 Nov 14. pii: S2211-1247(24)01337-8. [Epub ahead of print]43(11): 114986
    Low-affinity ligands of the epidermal growth factor receptor are long-range signal transmitters in collective cell migration of epithelial cells.
    Eriko Deguchi, Shuhao Lin, Daiki Hirayama, Kimiya Matsuda, Akira Tanave, Kenta Sumiyama, Shinya Tsukiji, Tetsuhisa Otani, Mikio Furuse, Alexander Sorkin, Michiyuki Matsuda, Kenta Terai.
      Canonical epidermal growth factor (EGF) receptor (EGFR) activation involves the binding of seven EGFR ligands (EGFRLs); however, their extracellular dynamics remain elusive. Here, employing fluorescent probes and a tool for triggering ectodomain shedding, we show that epiregulin (EREG), a low-affinity EGFRL, rapidly and efficiently activates EGFR in Madin-Darby canine kidney (MDCK) epithelial cells and mouse epidermis. During collective cell migration, EGFR and extracellular signal-regulated kinase (ERK) activation waves propagate in an a disintegrin and metalloprotease 17 (ADAM17) sheddase- and EGFRL-dependent manner. Upon induced EGFRL shedding, low-affinity ligands EREG and amphiregulin (AREG) mediate faster and broader ERK waves than high-affinity ligands. Tight/adherens junction integrity is essential for ERK activation propagation, suggesting that tight intercellular spaces prefer the low-affinity EGFRLs for efficient signal transmission. In EREG-deficient mice, ERK wave propagation and cell migration were impaired during skin wound repair. We additionally show that heparin-binding EGF-like growth factor (HBEGF) primarily promotes surrounding cell motility. Our findings underscore the pivotal role of low-affinity EGFRLs in rapid intercellular signal transmission.
    Keywords:  ADAM17; CP: Cell biology; EGFR ligand; EREG; ERK activation wave; FRET; chemogenetics; collective cell migration; transgenic mice
    DOI:  https://doi.org/10.1016/j.celrep.2024.114986
  4. J Cell Sci. 2024 Nov 08. pii: jcs.262232. [Epub ahead of print]
    Shear stress-stimulated AMPK couples endothelial cell mechanics, metabolism, and vasodilation.
    Nicholas M Cronin, Logan W Dawson, Kris A DeMali.
      Endothelial cells respond to mechanical force by stimulating cellular signaling, but how these pathways are linked to elevations in cell metabolism and whether metabolism supports the mechanical response remains poorly understood. Here, we show that the application of force to endothelial cells stimulates VE-cadherin to activate liver kinase B1 (LKB1) and AMP-activated protein kinase (AMPK), a master regulator of energy homeostasis. VE-cadherin stimulated AMPK increases eNOS activity and localization to the plasma membrane, reinforcement of the actin cytoskeleton and cadherin adhesion complex, and glucose uptake. We present evidence for the increase in metabolism being necessary to fortify the adhesion complex, actin cytoskeleton, and cellular alignment. Together these data extend the paradigm for how mechanotransduction and metabolism are linked to include a connection to vasodilation, thereby providing new insight into how diseases involving contractile, metabolic, and vasodilatory disturbances arise.
    Keywords:  Cell-Cell Adhesion; Mechanotransduction; VE-Cadherin
    DOI:  https://doi.org/10.1242/jcs.262232
  5. Elife. 2024 Nov 08. pii: e101652. [Epub ahead of print]13
    Bilateral regulation of EGFR activity and local PI(4,5)P2 dynamics in mammalian cells observed with superresolution microscopy.
    Mitsuhiro Abe, Masataka Yanagawa, Michio Hiroshima, Toshihide Kobayashi, Yasushi Sako.
      Anionic lipid molecules, including phosphatidylinositol-4,5-bisphosphate (PI(4,5)P2), are implicated in the regulation of epidermal growth factor receptor (EGFR). However, the role of the spatiotemporal dynamics of PI(4,5)P2 in the regulation of EGFR activity in living cells is not fully understood, as it is difficult to visualize the local lipid domains around EGFR. Here, we visualized both EGFR and PI(4,5)P2 nanodomains in the plasma membrane of HeLa cells using super-resolution single-molecule microscopy. The EGFR and PI(4,5)P2 nanodomains aggregated before stimulation with epidermal growth factor (EGF) through transient visits of EGFR to the PI(4,5)P2 nanodomains. The degree of coaggregation decreased after EGF stimulation and depended on phospholipase Cγ, the EGFR effector hydrolyzing PI(4,5)P2. Artificial reduction in the PI(4,5)P2 content of the plasma membrane reduced both the dimerization and autophosphorylation of EGFR after stimulation with EGF. Inhibition of PI(4,5)P2 hydrolysis after EGF stimulation decreased phosphorylation of EGFR-Thr654. Thus, EGFR kinase activity and the density of PI(4,5)P2 around EGFR molecules were found to be mutually regulated.
    Keywords:  EGFR; cell biology; epidermal growth factor receptor; human; membrane lipid; phosphatidylinositol-4,5-bisphosphate; single-molecule imaging; super-resolution microscopy
    DOI:  https://doi.org/10.7554/eLife.101652
  6. Nat Biotechnol. 2024 Nov 13.
    Directed evolution of engineered virus-like particles with improved production and transduction efficiencies.
    Aditya Raguram, Meirui An, Paul Z Chen, David R Liu.
      Engineered virus-like particles (eVLPs) are promising vehicles for transient delivery of proteins and RNAs, including gene editing agents. We report a system for the laboratory evolution of eVLPs that enables the discovery of eVLP variants with improved properties. The system uses barcoded guide RNAs loaded within DNA-free eVLP-packaged cargos to uniquely label each eVLP variant in a library, enabling the identification of desired variants following selections for desired properties. We applied this system to mutate and select eVLP capsids with improved eVLP production properties or transduction efficiencies in human cells. By combining beneficial capsid mutations, we developed fifth-generation (v5) eVLPs, which exhibit a 2-4-fold increase in cultured mammalian cell delivery potency compared to previous-best v4 eVLPs. Analyses of v5 eVLPs suggest that these capsid mutations optimize packaging and delivery of desired ribonucleoprotein cargos rather than native viral genomes and substantially alter eVLP capsid structure. These findings suggest the potential of barcoded eVLP evolution to support the development of improved eVLPs.
    DOI:  https://doi.org/10.1038/s41587-024-02467-x
  7. Cell. 2024 Nov 14. pii: S0092-8674(24)01211-X. [Epub ahead of print]187(23): 6421-6423
    The cellular dogma.
    Stephen R Quake.
      In this essay, I will put forth what I see as a major conceptual challenge for biology in the next decade, one that is inspired by Crick's Central Dogma: understanding information flow in the cell in the most general sense.
    DOI:  https://doi.org/10.1016/j.cell.2024.10.029
  8. Am J Respir Cell Mol Biol. 2024 Nov 12.
    The Bi-Steric Inhibitor RMC-5552 Reduces mTORC1 Signaling and Growth in Lymphangioleiomyomatosis.
    Jilly F Evans, Owen A Ledwell, Yan Tang, Ryan Rue, Alexander R Mukhitov, Rémi Diesler, Susan M Lin, Swaroop V Kanth, Maria C Basil, Edward Cantu, Elizabeth P Henske, Vera P Krymskaya.
      Mutations in the Tuberous Sclerosis Complex (TSC) genes result in the hyperactivation of the mechanistic/mammalian target of rapamycin 1 (mTORC1) growth pathway in mesenchymal pulmonary cells. Rapamycin (SirolimusTM), a naturally occurring macrolide, is the only therapeutic approved for women with lymphangioleiomyomatosis (LAM), a progressive, destructive lung disease caused by TSC gene mutations and mTORC1 hyperactivation. However, on cessation of the drug, lung function decline continues. We demonstrated here that pulmonary LAM cancer stem-like cells (SLS) most highly expressed the eukaryotic translation initiation factor 4E (eIF4E)-dependent translation initiation genes. We also showed that the eukaryotic initiation factor 4E-binding protein 1 (4E-BP1) gene has the lowest expression in these cells, indicating that the 4E-BP1/eIF4E ratio in LAM SLS cells favors unrestrained eIF4E oncogenic mRNA translation. The bi-steric mTORC1-selective compound RMC-5552 prevented growth of LAM-associated fibroblasts (LAFs) and phosphorylation of proteins in the ribosomal protein S6K1/ribosomal protein S6 (S6K1/S6) and 4E-BP1/eIF4E translation mTORC1-driven pathways, whereas rapamycin only blocked the S6K/S6 axis. Rapamycin inhibition of LAF growth was rapidly reversed, but RMC-5552 inhibition was more durable. RMC-5552, through its potential to eradicate LAM cancer SLS cells, may have therapeutic benefit in LAM and other diseases with mTORC1 hyperactivity.
    Keywords:  RMC-5552 bi-steric inhibitor; TSC2; lymphangioleiomyomatosis; mTORC1; rapamycin
    DOI:  https://doi.org/10.1165/rcmb.2024-0242OC
  9. Chem Rev. 2024 Nov 13.
    Molecular Spies in Action: Genetically Encoded Fluorescent Biosensors Light up Cellular Signals.
    Anneliese M M Gest, Ayse Z Sahan, Yanghao Zhong, Wei Lin, Sohum Mehta, Jin Zhang.
      Cellular function is controlled through intricate networks of signals, which lead to the myriad pathways governing cell fate. Fluorescent biosensors have enabled the study of these signaling pathways in living systems across temporal and spatial scales. Over the years there has been an explosion in the number of fluorescent biosensors, as they have become available for numerous targets, utilized across spectral space, and suited for various imaging techniques. To guide users through this extensive biosensor landscape, we discuss critical aspects of fluorescent proteins for consideration in biosensor development, smart tagging strategies, and the historical and recent biosensors of various types, grouped by target, and with a focus on the design and recent applications of these sensors in living systems.
    DOI:  https://doi.org/10.1021/acs.chemrev.4c00293
  10. Nat Commun. 2024 Nov 07. 15(1): 9173
    Engineered PsCas9 enables therapeutic genome editing in mouse liver with lipid nanoparticles.
    Dmitrii Degtev, Jack Bravo, Aikaterini Emmanouilidi, Aleksandar Zdravković, Oi Kuan Choong, Julia Liz Touza, Niklas Selfjord, Isabel Weisheit, Margherita Francescatto, Pinar Akcakaya, Michelle Porritt, Marcello Maresca, David Taylor, Grzegorz Sienski.
      Clinical implementation of therapeutic genome editing relies on efficient in vivo delivery and the safety of CRISPR-Cas tools. Previously, we identified PsCas9 as a Type II-B family enzyme capable of editing mouse liver genome upon adenoviral delivery without detectable off-targets and reduced chromosomal translocations. Yet, its efficacy remains insufficient with non-viral delivery, a common challenge for many Cas9 orthologues. Here, we sought to redesign PsCas9 for in vivo editing using lipid nanoparticles. We solve the PsCas9 ribonucleoprotein structure with cryo-EM and characterize it biochemically, providing a basis for its rational engineering. Screening over numerous guide RNA and protein variants lead us to develop engineered PsCas9 (ePsCas9) with up to 20-fold increased activity across various targets and preserved safety advantages. We apply the same design principles to boost the activity of FnCas9, an enzyme phylogenetically relevant to PsCas9. Remarkably, a single administration of mRNA encoding ePsCas9 and its guide formulated with lipid nanoparticles results in high levels of editing in the Pcsk9 gene in mouse liver, a clinically relevant target for hypercholesterolemia treatment. Collectively, our findings introduce ePsCas9 as a highly efficient, and precise tool for therapeutic genome editing, in addition to the engineering strategy applicable to other Cas9 orthologues.
    DOI:  https://doi.org/10.1038/s41467-024-53418-8
  11. J Clin Invest. 2024 Nov 15. pii: e181923. [Epub ahead of print]134(22):
    Transparency, bias, and reproducibility across science: a meta-research view.
    John Pa Ioannidis.
      N/A.
    DOI:  https://doi.org/10.1172/JCI181923
  12. Mol Cell Proteomics. 2024 Nov 08. pii: S1535-9476(24)00167-1. [Epub ahead of print] 100877
    Bridging the Gap from Proteomics Technology to Clinical Application: Highlights from the 68th Benzon Foundation Symposium.
    Vincent Albrecht, Johannes Müller-Reif, Thierry M Nordmann, Andreas Mund, Lisa Schweizer, Philipp E Geyer, Lili Niu, Juanjuan Wang, Frederik Post, Marc Oeller, Andreas Metousis, Annelaura Bach Nielsen, Medini Steger, Nicolai J Wewer Albrechtsen, Matthias Mann.
      The 68th Benzon Foundation Symposium brought together leading experts to explore the integration of mass spectrometry (MS)-based proteomics and artificial intelligence in revolutionizing personalized medicine. This report highlights key discussions on recent technological advances in MS-based proteomics, including improvements in sensitivity, throughput, and data analysis. Particular emphasis was placed on plasma proteomics and its potential for biomarker discovery across various diseases. The symposium addressed critical challenges in translating proteomic discoveries to clinical practice, including standardization, regulatory considerations and the need for robust 'business cases' to motivate adoption. Promising applications were presented in areas such as cancer diagnostics, neurodegenerative diseases, and cardiovascular health. The integration of proteomics with other omics technologies and imaging methods was explored, showcasing the power of multi-modal approaches in understanding complex biological systems. Artificial intelligence emerged as a crucial tool for the acquisition of large-scale proteomic datasets, extracting meaningful insights, and enhancing clinical decision-making. By fostering dialogue between academic researchers, industry leaders in proteomics technology, and clinicians, the symposium illuminated potential pathways for proteomics to transform personalized medicine, advancing the cause of more precise diagnostics and targeted therapies.
    DOI:  https://doi.org/10.1016/j.mcpro.2024.100877
  13. Nature. 2024 Nov 13.
    Clinical functional proteomics of intercellular signalling in pancreatic cancer.
    Peiwu Huang, Weina Gao, Changying Fu, Min Wang, Yunguang Li, Bizhu Chu, An He, Yuan Li, Xiaomei Deng, Yehan Zhang, Qian Kong, Jingxiong Yuan, Hebin Wang, Yu Shi, Dong Gao, Renyi Qin, Tony Hunter, Ruijun Tian.
      Pancreatic ductal adenocarcinoma (PDAC) has an atypical, highly stromal tumour microenvironment (TME) that profoundly contributes to its poor prognosis1. Here, to better understand the intercellular signalling between cancer and stromal cells directly in PDAC tumours, we developed a multidimensional proteomic strategy called TMEPro. We applied TMEPro to profile the glycosylated secreted and plasma membrane proteome of 100 human pancreatic tissue samples to a great depth, define cell type origins and identify potential paracrine cross-talk, especially that mediated through tyrosine phosphorylation. Temporal dynamics during pancreatic tumour progression were investigated in a genetically engineered PDAC mouse model. Functionally, we revealed reciprocal signalling between stromal cells and cancer cells mediated by the stromal PDGFR-PTPN11-FOS signalling axis. Furthermore, we examined the generic shedding mechanism of plasma membrane proteins in PDAC tumours and revealed that matrix-metalloprotease-mediated shedding of the AXL receptor tyrosine kinase ectodomain provides an additional dimension of intercellular signalling regulation in the PDAC TME. Importantly, the level of shed AXL has a potential correlation with lymph node metastasis, and inhibition of AXL shedding and its kinase activity showed a substantial synergistic effect in inhibiting cancer cell growth. In summary, we provide TMEPro, a generically applicable clinical functional proteomic strategy, and a comprehensive resource for better understanding the PDAC TME and facilitating the discovery of new diagnostic and therapeutic targets.
    DOI:  https://doi.org/10.1038/s41586-024-08225-y
  14. PLoS Genet. 2024 Nov 15. 20(11): e1011453
    A non-canonical role of somatic Cyclin D/CYD-1 in oogenesis and in maintenance of reproductive fidelity, dependent on the FOXO/DAF-16 activation state.
    Umanshi Rautela, Gautam Chandra Sarkar, Ayushi Chaudhary, Debalina Chatterjee, Mohtashim Rosh, Aneeshkumar G Arimbasseri, Arnab Mukhopadhyay.
      For the optimal survival of a species, an organism coordinates its reproductive decisions with the nutrient availability of its niche. Thus, nutrient-sensing pathways like insulin-IGF-1 signaling (IIS) play an important role in modulating cell division, oogenesis, and reproductive aging. Lowering of the IIS leads to the activation of the downstream FOXO transcription factor (TF) DAF-16 in Caenorhabditis elegans which promotes oocyte quality and delays reproductive aging. However, less is known about how the IIS axis responds to changes in cell cycle proteins, particularly in the somatic tissues. Here, we show a new aspect of the regulation of the germline by this nutrient-sensing axis. First, we show that the canonical G1-S cyclin, Cyclin D/CYD-1, regulates reproductive fidelity from the uterine tissue of wild-type worms. Then, we show that knocking down cyd-1 in the uterine tissue of an IIS receptor mutant arrests oogenesis at the pachytene stage of meiosis-1 in a DAF-16-dependent manner. We observe activated DAF-16-dependent deterioration of the somatic gonadal tissues like the sheath cells, and transcriptional de-regulation of the sperm-to-oocyte switch genes which may be the underlying reason for the absence of oogenesis. Deleting DAF-16 releases the arrest and leads to restoration of the somatic gonad but poor-quality oocytes are produced. Together, our study reveals the unrecognized cell non-autonomous interaction of Cyclin D/CYD-1 and FOXO/DAF-16 in the regulation of oogenesis and reproductive fidelity.
    DOI:  https://doi.org/10.1371/journal.pgen.1011453
  15. NPJ Syst Biol Appl. 2024 Nov 16. 10(1): 134
    Systems-level reconstruction of kinase phosphosignaling networks regulating endothelial barrier integrity using temporal data.
    Ling Wei, John D Aitchison, Alexis Kaushansky, Fred D Mast.
      Phosphosignaling networks control cellular processes. We built kinase-mediated regulatory networks elicited by thrombin stimulation of brain endothelial cells using two computational strategies: Temporal Pathway Synthesizer (TPS), which uses phosphoproteomics data as input, and Temporally REsolved KInase Network Generation (TREKING), which uses kinase inhibitor screens. TPS and TREKING predicted overlapping barrier-regulatory kinases connected with unique network topology. Each strategy effectively describes regulatory signaling networks and is broadly applicable across biological systems.
    DOI:  https://doi.org/10.1038/s41540-024-00468-9
  16. Life Sci Alliance. 2025 Jan;pii: e202402859. [Epub ahead of print]8(1):
    Transcriptional regulators ensuring specific gene expression and decision-making at high TGFβ doses.
    Laura Hartmann, Panajot Kristofori, Congxin Li, Kolja Becker, Lorenz Hexemer, Stefan Bohn, Sonja Lenhardt, Sylvia Weiss, Björn Voss, Alexander Loewer, Stefan Legewie.
      TGFβ-signaling regulates cancer progression by controlling cell division, migration, and death. These outcomes are mediated by gene expression changes, but the mechanisms of decision-making toward specific fates remain unclear. Here, we combine SMAD transcription factor imaging, genome-wide RNA sequencing, and morphological assays to quantitatively link signaling, gene expression, and fate decisions in mammary epithelial cells. Fitting genome-wide kinetic models to our time-resolved data, we find that most of the TGFβ target genes can be explained as direct targets of SMAD transcription factors, whereas the remainder show signs of complex regulation, involving delayed regulation and strong amplification at high TGFβ doses. Knockdown experiments followed by global RNA sequencing revealed transcription factors interacting with SMADs in feedforward loops to control delayed and dose-discriminating target genes, thereby reinforcing the specific epithelial-to-mesenchymal transition at high TGFβ doses. We identified early repressors, preventing premature activation, and a late activator, boosting gene expression responses for a sufficiently strong TGFβ stimulus. Taken together, we present a global view of TGFβ-dependent gene regulation and describe specificity mechanisms reinforcing cellular decision-making.
    DOI:  https://doi.org/10.26508/lsa.202402859
  17. Nat Cancer. 2024 Nov 11.
    The pro-oncogenic noncanonical activity of a RAS•GTP:RanGAP1 complex facilitates nuclear protein export.
    Brajendra K Tripathi, Nicole H Hirsh, Xiaolan Qian, Marian E Durkin, Dunrui Wang, Alex G Papageorge, Ross Lake, Yvonne A Evrard, Adam I Marcus, Suresh S Ramalingam, Mary Dasso, Karen H Vousden, James H Doroshow, Kylie J Walters, Douglas R Lowy.
      Canonical RAS signaling, including PI3K/AKT- and RAF/MEK-dependent activities, results mainly from RAS•GTP interaction with its effectors at the plasma membrane. Here, we identified a fundamental, oncogenic, noncanonical RAS•GTP activity that increases XPO1-dependent export of nuclear protein cargo into the cytoplasm and is independent of PI3K/AKT and RAF/MEK signaling. This RAS-dependent step acts downstream from XPO1 binding to nuclear protein cargo and is mediated by a perinuclear protein complex between RAS•GTP and RanGAP1 that facilitates hydrolysis of Ran•GTP to Ran•GDP, which promotes release of nuclear protein cargo into the cytoplasm. The export of nuclear EZH2, which promotes cytoplasmic degradation of the DLC1 tumor suppressor protein, is a biologically important component of this pro-oncogenic activity. Conversely, preventing nuclear protein export contributes to the antitumor activity of KRAS inhibition, which can be further augmented by reactivating the tumor suppressor activity of DLC1 or potentially combining RAS inhibitors with other cancer treatments.
    DOI:  https://doi.org/10.1038/s43018-024-00847-5
  18. Sci Rep. 2024 11 09. 14(1): 27348
    Protein phosphatase-1 regulates the binding of filamin C to FILIP1 in cultured skeletal muscle cells under mechanical stress.
    Thomas Kokot, Johannes P Zimmermann, Anja N Schwäble, Lena Reimann, Anna L Herr, Nico Höfflin, Maja Köhn, Bettina Warscheid.
      The actin-binding protein filamin c (FLNc) is a key mediator in the response of skeletal muscle cells to mechanical stress. In addition to its function as a structural scaffold, FLNc acts as a signaling adaptor which is phosphorylated at S2234 in its mechanosensitive domain 20 (d20) through AKT. Here, we discovered a strong dephosphorylation of FLNc-pS2234 in cultured skeletal myotubes under acute mechanical stress, despite high AKT activity. We found that all three protein phosphatase 1 (PP1) isoforms are part of the FLNc d18-21 interactome. Enzymatic assays demonstrate that PP1 efficiently dephosphorylates FLNc-pS2234 and in vitro and in cells upon PP1 activation using specific modulators. FLNc-pS2234 dephosphorylation promotes the interaction with FILIP1, a mediator for filamin degradation. Altogether, we present a model in which dephosphorylation of FLNc d20 by the dominant action of PP1c prevails over AKT activity to promote the binding of the filamin degradation-inducing factor FILIP1 during acute mechanical stress.
    DOI:  https://doi.org/10.1038/s41598-024-78953-8
  19. Nat Commun. 2024 Nov 07. 15(1): 9635
    The massed-spaced learning effect in non-neural human cells.
    N V Kukushkin, R E Carney, T Tabassum, T J Carew.
      The massed-spaced effect is a hallmark feature of memory formation. We now demonstrate this effect in two separate non-neural, immortalized cell lines stably expressing a short-lived luciferase reporter controlled by a CREB-dependent promoter. We emulate training using repeated pulses of forskolin and/or phorbol ester, and, as a proxy for memory, measure luciferase expression at various points after training. Four spaced pulses of either agonist elicit stronger and more sustained luciferase expression than a single "massed" pulse. Spaced pulses also result in stronger and more sustained activation of molecular factors critical for memory formation, ERK and CREB, and inhibition of ERK or CREB blocks the massed-spaced effect. Our findings show that canonical features of memory do not necessarily depend on neural circuitry, but can be embedded in the dynamics of signaling cascades conserved across different cell types.
    DOI:  https://doi.org/10.1038/s41467-024-53922-x
  20. Elife. 2024 Nov 14. pii: RP92025. [Epub ahead of print]13
    Proteomic and functional comparison between human induced and embryonic stem cells.
    Alejandro J Brenes, Eva Griesser, Linda V Sinclair, Lindsay Davidson, Alan R Prescott, Francois Singh, Elizabeth K J Hogg, Carmen Espejo-Serrano, Hao Jiang, Harunori Yoshikawa, Melpomeni Platani, Jason R Swedlow, Greg M Findlay, Doreen A Cantrell, Angus I Lamond.
      Human induced pluripotent stem cells (hiPSCs) have great potential to be used as alternatives to embryonic stem cells (hESCs) in regenerative medicine and disease modelling. In this study, we characterise the proteomes of multiple hiPSC and hESC lines derived from independent donors and find that while they express a near-identical set of proteins, they show consistent quantitative differences in the abundance of a subset of proteins. hiPSCs have increased total protein content, while maintaining a comparable cell cycle profile to hESCs, with increased abundance of cytoplasmic and mitochondrial proteins required to sustain high growth rates, including nutrient transporters and metabolic proteins. Prominent changes detected in proteins involved in mitochondrial metabolism correlated with enhanced mitochondrial potential, shown using high-resolution respirometry. hiPSCs also produced higher levels of secreted proteins, including growth factors and proteins involved in the inhibition of the immune system. The data indicate that reprogramming of fibroblasts to hiPSCs produces important differences in cytoplasmic and mitochondrial proteins compared to hESCs, with consequences affecting growth and metabolism. This study improves our understanding of the molecular differences between hiPSCs and hESCs, with implications for potential risks and benefits for their use in future disease modelling and therapeutic applications.
    Keywords:  biochemistry; chemical biology; hESC; human; iPSC; mass spectrometry; protein content; proteomics; regenerative medicine; stem cell; stem cells
    DOI:  https://doi.org/10.7554/eLife.92025
  21. Radiol Case Rep. 2025 Jan;20(1): 427-431
    Vascular lesions: Hemangioma or venous malformation?
    Brandon Thinh Duc Dang, Alan Victor Krauthamer.
      We present a case of a 62-year-old female who was incidentally found to have a venous malformation. Venous malformations are part of a larger category of slow flow vascular malformations and are associated with various familial syndromes and localized intravascular coagulation. Venous malformations were often misdiagnosed as hemangiomas; however, the treatment modalities of vascular malformations and hemangiomas vary significantly. Here we elucidate the imaging findings of venous malformations from various vascular tumors and other malformations.
    Keywords:  Hemangioma; MRI; Vascular tumors; Venous malformation
    DOI:  https://doi.org/10.1016/j.radcr.2024.10.069
  22. Nat Commun. 2024 Nov 15. 15(1): 9932
    scPair: Boosting single cell multimodal analysis by leveraging implicit feature selection and single cell atlases.
    Hongru Hu, Gerald Quon.
      Multimodal single-cell assays profile multiple sets of features in the same cells and are widely used for identifying and mapping cell states between chromatin and mRNA and linking regulatory elements to target genes. However, the high dimensionality of input features and shallow sequencing depth compared to unimodal assays pose challenges in data analysis. Here we present scPair, a multimodal single-cell data framework that overcomes these challenges by employing an implicit feature selection approach. scPair uses dual encoder-decoder structures trained on paired data to align cell states across modalities and predict features from one modality to another. We demonstrate that scPair outperforms existing methods in accuracy and execution time, and facilitates downstream tasks such as trajectory inference. We further show scPair can augment smaller multimodal datasets with larger unimodal atlases to increase statistical power to identify groups of transcription factors active during different stages of neural differentiation.
    DOI:  https://doi.org/10.1038/s41467-024-53971-2
  23. Nucleic Acids Res. 2024 Nov 13. pii: gkae1108. [Epub ahead of print]
    Rediscovering publicly available single-cell data with the DISCO platform.
    Mengwei Li, Kok Siong Ang, Brian Teo, Uddamvathanak Rom, Minh N Nguyen, Sebastian Maurer-Stroh, Jinmiao Chen.
      Single-cell RNA sequencing (scRNA-seq) has emerged as the key technique for studying transcriptomics at the single-cell level. In our previous work, we presented the DISCO database (https://www.immunesinglecell.org/) that integrates publicly available human scRNA-seq data. We now introduce an enhanced version of DISCO, which has expanded fourfold to include >100 million cells from >17 thousand samples. It provides uniformly realigned read count tables, curated metadata, integrated tissue and phenotype specific atlases, and harmonized cell type annotations. It also hosts a single-cell enhanced knowledgebase of cell type ontology and gene signatures relating to cell types and phenotypes. Lastly, it offers a suite of tools for data retrieval, integration, annotation, and mapping, allowing users to construct customized atlases and perform integrated analysis with their own data. These tools are also available in a standalone R package for offline analysis.
    DOI:  https://doi.org/10.1093/nar/gkae1108
  24. Am J Respir Cell Mol Biol. 2024 Nov 08.
    Enhanced Gαq Signaling in TSC2-deficient Cells Is Required for Their Neoplastic Behavior.
    Aurelie Trefier, Nihad Tousson-Abouelazm, Lama Yamani, Sajida Ibrahim, Kwang-Bo Joung, Adam Pietrobon, Julien Yockell-Lelievre, Terence E Hébert, Reese J Ladak, Tomoko Takano, Mark Nellist, Yoon Namkung, David Chatenet, William L Stanford, Stephane A Laporte, Arnold S Kristof.
      Inherited or sporadic loss of the TSC2 gene can lead to pulmonary lymphangioleiomyomatosis (LAM), a rare cystic lung disease caused by protease-secreting interstitial tumor nodules. The nodules arise by metastasis of cells that exhibit features of neural crest and smooth muscle lineage ('LAM cells'). Their aberrant growth is attributed to increased activity of 'mechanistic target of rapamycin complex 1' (mTORC1), an anabolic protein kinase that is normally suppressed by the TSC1-TSC2 protein complex. The mTORC1 inhibitor rapamycin slows the progression of LAM, but fails to eradicate disease, indicating a role for mTORC1-independent mechanisms in LAM pathogenesis. Our previous studies revealed G-protein coupled urotensin-II receptor (UT) signaling as a candidate mechanism, but how it promotes oncogenic signaling in TSC2-deficient cells remained unknown. Using a human pluripotent stem cell-derived in vitro model of LAM, we now show hyperactivation of UT, which was required for their enhanced migration and pro-neoplastic signaling in a rapamycin-insensitive mechanism that required heterotrimeric Gαq/11 (Gαq). Bioluminescence resonance energy transfer assays in HEK 293T cells lacking TSC2 demonstrated selective and enhanced activation of Gαq and its RhoA-associated effectors compared to wild-type control cells. By immunoprecipitation, recombinant UT was physically associated with Gαq and TSC2. The augmented Gαq signaling in TSC2-deleted cells was independent of mTOR activity, and associated with increased endosomal targeting of p63RhoGEF, a known RhoA-activating effector of Gαq. These studies identify potential mTORC1-independent pro-neoplastic mechanisms that can be targeted for prevention or eradication of pulmonary and extrapulmonary LAM tumors.
    Keywords:  Gαq; Lymphangioleiomyomatosis; Mechanistic target of rapamycin; Tuberous sclerosis complex; Urotensin-II receptor
    DOI:  https://doi.org/10.1165/rcmb.2024-0111OC
  25. EMBO Rep. 2024 Nov 12.
    Cultivating the next generation of leaders : How postdocs, principal investigators and institutes can nurture and select for leadership competencies.
    James W Bryson, Ülkü Uzun, Victor O Oria, Jamie Y Auxillos, Iman Safari, Alexia M Lopresti, Agnieszka Krzyzanowska, Katrine Sonne-Hansen.
      
    DOI:  https://doi.org/10.1038/s44319-024-00308-1
  26. Nat Methods. 2024 Nov 14.
    A comprehensive human embryo reference tool using single-cell RNA-sequencing data.
    Cheng Zhao, Alvaro Plaza Reyes, John Paul Schell, Jere Weltner, Nicolás M Ortega, Yi Zheng, Åsa K Björklund, Laura Baqué-Vidal, Joonas Sokka, Ras Torokovic, Brian Cox, Janet Rossant, Jianping Fu, Sophie Petropoulos, Fredrik Lanner.
      Stem cell-based embryo models offer unprecedented experimental tools for studying early human development. The usefulness of embryo models hinges on their molecular, cellular and structural fidelities to their in vivo counterparts. To authenticate human embryo models, single-cell RNA sequencing has been utilized for unbiased transcriptional profiling. However, an organized and integrated human single-cell RNA-sequencing dataset, serving as a universal reference for benchmarking human embryo models, remains unavailable. Here we developed such a reference through the integration of six published human datasets covering development from the zygote to the gastrula. Lineage annotations are contrasted and validated with available human and nonhuman primate datasets. Using stabilized Uniform Manifold Approximation and Projection, we constructed an early embryogenesis prediction tool, where query datasets can be projected on the reference and annotated with predicted cell identities. Using this reference tool, we examined published human embryo models, highlighting the risk of misannotation when relevant references are not utilized for benchmarking and authentication.
    DOI:  https://doi.org/10.1038/s41592-024-02493-2
  27. Heliyon. 2024 Nov 15. 10(21): e39752
    TPC1 regulates melanoma tumourigenesis via mTORC1 and TFEB.
    Xuhui Jin, Ali A Hanbashi, Faroq Kamli, Xiaoqi Pan, Colin R Goding, John Parrington.
      The major cause of death in cancer patients is a combination of metastatic dissemination combined with therapy resistance. Over recent years, intratumour phenotypic heterogeneity arising from the bi-directional interplay between plastic cancer cells and the microenvironment has been identified as key to disease progression. Most notably metastatic outgrowth and resistance to targeted therapies are frequently associated with activity of mTORC1, a key metabolic hub that promotes protein synthesis and proliferation in the presence of nutrients. Yet while the regulation of mTORC1 by amino acids and glucose availability is well characterized, whether other mechanisms are important in controlling mTORC1 and its downstream signalling is less well understood. Here we show, using the murine B16-F0 melanoma cell line as a model, that mTORC1 activity is decreased following the knockout (KO) of TPC1, a cation channel localised to early and recycling endosomes. Consequently, TPC1 KO melanoma cells exhibit reduced proliferation and invasiveness, as well as increased pigmentation associated with nuclear localisation of the MITF-related transcription factor TFEB. Our results demonstrate that the knockout of TPC1 has induced significant tumour-suppressive effects in melanoma, during which the altered activity of mTORC1 and TFEB play the key roles. The results help us further understand the link between mTORC1 and endolysosomal ion channels, and reveal that TPC1 controls melanoma progression and represents a potential therapeutic target.
    Keywords:  MITF; Melanoma; TFEB; TPC1; mTORC1
    DOI:  https://doi.org/10.1016/j.heliyon.2024.e39752
  28. Nat Commun. 2024 Nov 09. 15(1): 9699
    Reconstructing the regulatory programs underlying the phenotypic plasticity of neural cancers.
    Ida Larsson, Felix Held, Gergana Popova, Alper Koc, Soumi Kundu, Rebecka Jörnsten, Sven Nelander.
      Nervous system cancers exhibit diverse transcriptional cell states influenced by normal development, injury response, and growth. However, the understanding of these states' regulation and pharmacological relevance remains limited. Here we present "single-cell regulatory-driven clustering" (scregclust), a method that reconstructs cellular regulatory programs from extensive collections of single-cell RNA sequencing (scRNA-seq) data from both tumors and developing tissues. The algorithm efficiently divides target genes into modules, predicting key transcription factors and kinases with minimal computational time. Applying this method to adult and childhood brain cancers, we identify critical regulators and suggest interventions that could improve temozolomide treatment in glioblastoma. Additionally, our integrative analysis reveals a meta-module regulated by SPI1 and IRF8 linked to an immune-mediated mesenchymal-like state. Finally, scregclust's flexibility is demonstrated across 15 tumor types, uncovering both pan-cancer and specific regulators. The algorithm is provided as an easy-to-use R package that facilitates the exploration of regulatory programs underlying cell plasticity.
    DOI:  https://doi.org/10.1038/s41467-024-53954-3
  29. Oncogene. 2024 Nov 13.
    Unveiling RACK1: a key regulator of the PI3K/AKT pathway in prostate cancer development.
    Cancan Lyu, Prasanna Kuma Vaddi, Said Elshafae, Anirudh Pradeep, Deqin Ma, Songhai Chen.
      The dysregulated PI3K/AKT pathway is pivotal in the onset and progression of various cancers, including prostate cancer. However, targeting this pathway directly poses challenges due to compensatory upregulation of alternative oncogenic pathways. This study focuses on the novel regulatory activity of the Receptor for Activated Protein Kinase (RACK1), a scaffolding/adaptor protein, in governing the PI3K/AKT pathway within prostate cancer. Through a genetic mouse model, our research unveils RACK1's pivotal role in orchestrating AKT activation and the genesis of prostate cancer. RACK1 deficiency hampers AKT activation, effectively impeding prostate tumor formation induced by PTEN and p53 deficiency. Mechanistically, RACK1 facilitates AKT membrane translocation and fosters its interaction with mTORC2, thereby promoting AKT activation and subsequent tumor cell proliferation and tumor formation. Notably, inhibiting AKT activation via RACK1 deficiency does not trigger feedback upregulation of HER3 and androgen receptor (AR) expression and activation, distinguishing it from direct PI3K or AKT targeting. These findings position RACK1 as a critical regulator of the PI3K/AKT pathway and a promising target for curtailing prostate cancer development arising from pathway aberrations.
    DOI:  https://doi.org/10.1038/s41388-024-03224-9
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