bims-pideca Biomed News
on Class IA PI3K signalling in development and cancer
Issue of 2024‒09‒29
twenty-one papers selected by
Ralitsa Radostinova Madsen, MRC-PPU
  1. Vitessce: integrative visualization of multimodal and spatially resolved single-cell data.
  2. PIP5K-Ras bistability initiates plasma membrane symmetry breaking to regulate cell polarity and migration.
  3. Specific activation of the integrated stress response uncovers regulation of central carbon metabolism and lipid droplet biogenesis.
  4. mTOR activity paces human blastocyst stage developmental progression.
  5. Proteome-wide copy-number estimation from transcriptomics.
  6. Fluorescent tagging of endogenous IRS2 with an auxin-dependent degron to assess dynamic intracellular localization and function.
  7. Self-sustaining long-term 3D epithelioid cultures reveal drivers of clonal expansion in esophageal epithelium.
  8. Functional activation of the AKT-mTOR signalling axis in a real-world metastatic breast cancer cohort.
  9. Rapid generation of homozygous fluorescent knock-in human cells using CRISPR-Cas9 genome editing and validation by automated imaging and digital PCR screening.
  10. Engineering next generation vascularized organoids.
  11. Functional profiling of serine, threonine and tyrosine sites.
  12. Stress granule formation enables anchorage-independence survival in cancer cells.
  13. Thicker perineuronal nets disrupt insulin signalling.
  14. Senescence in the ageing skin: a new focus on mTORC1 and the lysosome.
  15. It took a long, long time: Ras and the race to cure cancer.
  16. MYC upstream region orchestrates resistance to PI3K inhibitors in cancer cells through FOXO3a-mediated autophagic adaptation.
  17. The inositol phosphate signalling network in physiology and disease.
  18. Designed endocytosis-inducing proteins degrade targets and amplify signals.
  19. PTEN controls alternative splicing of autism spectrum disorder-associated transcripts in primary neurons.
  20. Recovering single-cell expression profiles from spatial transcriptomics with scResolve.
  21. Insulin receptor substrate 1 is a novel member of EGFR signaling in pancreatic cells.
  1. Nat Methods. 2024 Sep 27.
    Vitessce: integrative visualization of multimodal and spatially resolved single-cell data.
    Mark S Keller, Ilan Gold, Chuck McCallum, Trevor Manz, Peter V Kharchenko, Nils Gehlenborg.
      Multiomics technologies with single-cell and spatial resolution make it possible to measure thousands of features across millions of cells. However, visual analysis of high-dimensional transcriptomic, proteomic, genome-mapped and imaging data types simultaneously remains a challenge. Here we describe Vitessce, an interactive web-based visualization framework for exploration of multimodal and spatially resolved single-cell data. We demonstrate integrative visualization of millions of data points, including cell-type annotations, gene expression quantities, spatially resolved transcripts and cell segmentations, across multiple coordinated views. The open-source software is available at http://vitessce.io .
    DOI:  https://doi.org/10.1038/s41592-024-02436-x
  2. bioRxiv. 2024 Sep 15. pii: 2024.09.15.613115. [Epub ahead of print]
    PIP5K-Ras bistability initiates plasma membrane symmetry breaking to regulate cell polarity and migration.
    Yu Deng, Tatsat Banerjee, Dhiman Sankar Pal, Parijat Banerjee, Huiwang Zhan, Jane Borleis, Pablo A Igleias, Peter N Devreotes.
      Symmetry breaking, polarity establishment, and spontaneous cell protrusion formation are fundamental but poorly explained cell behaviors. Here, we demonstrate that a biochemical network, where the mutually inhibitory localization of PIP5K and Ras activities plays a central role, governs these processes. First, in resting cells devoid of cytoskeletal activity, PIP5K is uniformly elevated on the plasma membrane, while Ras activity remains minimal. Symmetry is broken by spontaneous local displacements of PIP5K, coupled with simultaneous activations of Ras and downstream signaling events, including PI3K activation. Second, knockout of PIP5K dramatically increases both the incidence and size of Ras-PI3K activation patches, accompanied by branched F-actin assembly. This leads to enhanced cortical wave formation, increased protrusive activity, and a shift in migration mode. Third, high inducible overexpression of PIP5K virtually eliminates Ras-PI3K signaling, cytoskeletal activity, and cell migration, while acute recruitment of cytosolic PIP5K to the membrane induces contraction and blebs in cancer cells. These arrested phenotypes are reversed by reducing myosin II activity, indicating myosin's involvement in the PIP5K-Ras-centered regulatory network. Remarkably, low inducible overexpression of PIP5K unexpectedly facilitates polarity establishment, highlighting PIP5K as a highly sensitive master regulator of these processes. Simulations of a computational model combining an excitable system, cytoskeletal loops, and dynamic partitioning of PIP5K recreates the experimental observations. Taken together, our results reveal that a bistable, mutually exclusive localization of PIP5K and active Ras on the plasma membrane triggers the initial symmetry breaking. Coupled actomyosin reduction and increased actin polymerization lead to intermittently extended protrusions and, with feedback from the cytoskeleton, self-organizing, complementary gradients of PIP5K versus Ras steepen, raising the threshold of the networks at the rear and lowering it at the front to generate polarity for cell migration.
    DOI:  https://doi.org/10.1101/2024.09.15.613115
  3. Nat Commun. 2024 Sep 27. 15(1): 8301
    Specific activation of the integrated stress response uncovers regulation of central carbon metabolism and lipid droplet biogenesis.
    Katherine Labbé, Lauren LeBon, Bryan King, Ngoc Vu, Emily H Stoops, Nina Ly, Austin E Y T Lefebvre, Phillip Seitzer, Swathi Krishnan, Jin-Mi Heo, Bryson Bennett, Carmela Sidrauski.
      The integrated stress response (ISR) enables cells to cope with a variety of insults, but its specific contribution to downstream cellular outputs remains unclear. Using a synthetic tool, we selectively activate the ISR without co-activation of parallel pathways and define the resulting cellular state with multi-omics profiling. We identify time- and dose-dependent gene expression modules, with ATF4 driving only a small but sensitive subgroup that includes amino acid metabolic enzymes. This ATF4 response affects cellular bioenergetics, rerouting carbon utilization towards amino acid production and away from the tricarboxylic acid cycle and fatty acid synthesis. We also find an ATF4-independent reorganization of the lipidome that promotes DGAT-dependent triglyceride synthesis and accumulation of lipid droplets. While DGAT1 is the main driver of lipid droplet biogenesis, DGAT2 plays an essential role in buffering stress and maintaining cell survival. Together, we demonstrate the sufficiency of the ISR in promoting a previously unappreciated metabolic state.
    DOI:  https://doi.org/10.1038/s41467-024-52538-5
  4. Cell. 2024 Sep 18. pii: S0092-8674(24)00977-2. [Epub ahead of print]
    mTOR activity paces human blastocyst stage developmental progression.
    Dhanur P Iyer, Heidar Heidari Khoei, Vera A van der Weijden, Harunobu Kagawa, Saurabh J Pradhan, Maria Novatchkova, Afshan McCarthy, Teresa Rayon, Claire S Simon, Ilona Dunkel, Sissy E Wamaitha, Kay Elder, Phil Snell, Leila Christie, Edda G Schulz, Kathy K Niakan, Nicolas Rivron, Aydan Bulut-Karslioğlu.
      Many mammals can temporally uncouple conception from parturition by pacing down their development around the blastocyst stage. In mice, this dormant state is achieved by decreasing the activity of the growth-regulating mTOR signaling pathway. It is unknown whether this ability is conserved in mammals in general and in humans in particular. Here, we show that decreasing the activity of the mTOR signaling pathway induces human pluripotent stem cells (hPSCs) and blastoids to enter a dormant state with limited proliferation, developmental progression, and capacity to attach to endometrial cells. These in vitro assays show that, similar to other species, the ability to enter dormancy is active in human cells around the blastocyst stage and is reversible at both functional and molecular levels. The pacing of human blastocyst development has potential implications for reproductive therapies.
    Keywords:  blastoid; development; diapause; dormancy; human; mTOR; pluripotent stem cells
    DOI:  https://doi.org/10.1016/j.cell.2024.08.048
  5. Mol Syst Biol. 2024 Sep 27.
    Proteome-wide copy-number estimation from transcriptomics.
    Andrew J Sweatt, Cameron D Griffiths, Sarah M Groves, B Bishal Paudel, Lixin Wang, David F Kashatus, Kevin A Janes.
      Protein copy numbers constrain systems-level properties of regulatory networks, but proportional proteomic data remain scarce compared to RNA-seq. We related mRNA to protein statistically using best-available data from quantitative proteomics and transcriptomics for 4366 genes in 369 cell lines. The approach starts with a protein's median copy number and hierarchically appends mRNA-protein and mRNA-mRNA dependencies to define an optimal gene-specific model linking mRNAs to protein. For dozens of cell lines and primary samples, these protein inferences from mRNA outmatch stringent null models, a count-based protein-abundance repository, empirical mRNA-to-protein ratios, and a proteogenomic DREAM challenge winner. The optimal mRNA-to-protein relationships capture biological processes along with hundreds of known protein-protein complexes, suggesting mechanistic relationships. We use the method to identify a viral-receptor abundance threshold for coxsackievirus B3 susceptibility from 1489 systems-biology infection models parameterized by protein inference. When applied to 796 RNA-seq profiles of breast cancer, inferred copy-number estimates collectively re-classify 26-29% of luminal tumors. By adopting a gene-centered perspective of mRNA-protein covariation across different biological contexts, we achieve accuracies comparable to the technical reproducibility of contemporary proteomics.
    Keywords:  CCLE; CVB3; Pinferna; SWATH; TMT
    DOI:  https://doi.org/10.1038/s44320-024-00064-3
  6. J Biol Chem. 2024 Sep 19. pii: S0021-9258(24)02297-X. [Epub ahead of print] 107796
    Fluorescent tagging of endogenous IRS2 with an auxin-dependent degron to assess dynamic intracellular localization and function.
    Minjeong Jo, Ji-Sun Lee, Claire E Tocheny, Michael W Lero, Quyen Thu Bui, Jennifer S Morgan, Leslie M Shaw.
      Insulin Receptor Substrate 2 (IRS2) is a signaling adaptor protein for the insulin (IR) and Insulin-like Growth Factor-1 (IGF-1R) receptors. In breast cancer, IRS2 contributes to both initiation of primary tumor growth and establishment of secondary metastases through regulation of cancer stem cell (CSC) function and invasion. However, how IRS2 mediates its diverse functions is not well understood. We used CRISPR/Cas9-mediated gene editing to modify endogenous IRS2 to study the expression, localization, and function of this adaptor protein. A cassette containing an auxin inducible degradation (AID) sequence, 3X-FLAG tag and mNeon-green was introduced at the N-terminus of the IRS2 gene to provide rapid and reversible control of IRS2 protein degradation and analysis of endogenous IRS2 expression and localization. Live fluorescence imaging of these cells revealed that IRS2 shuttles between the cytoplasm and nucleus in response to growth regulatory signals in a PI3K-dependent manner. Inhibition of nuclear export or deletion of a putative nuclear export sequence in the C-terminal tail promotes nuclear retention of IRS2, implicating nuclear export in the mechanism by which IRS2 intracellular localization is regulated. Moreover, the acute induction of IRS2 degradation reduces tumor cell invasion, demonstrating the potential for therapeutic targeting of this adaptor protein. Our data highlight the value of our model of endogenously tagged IRS2 as a tool to study IRS2 localization and function.
    Keywords:  Auxin inducible degron; CRISPR/Cas9; Fluorescence imaging; IGF-1R; Insulin Receptor Substrate-2 (IRS2); nuclear translocation
    DOI:  https://doi.org/10.1016/j.jbc.2024.107796
  7. Nat Genet. 2024 Sep 23.
    Self-sustaining long-term 3D epithelioid cultures reveal drivers of clonal expansion in esophageal epithelium.
    Albert Herms, David Fernandez-Antoran, Maria P Alcolea, Argyro Kalogeropoulou, Ujjwal Banerjee, Gabriel Piedrafita, Emilie Abby, Jose Antonio Valverde-Lopez, Inês S Ferreira, Irene Caseda, Maria T Bejar, Stefan C Dentro, Sara Vidal-Notari, Swee Hoe Ong, Bartomeu Colom, Kasumi Murai, Charlotte King, Krishnaa Mahbubani, Kourosh Saeb-Parsy, Alan R Lowe, Moritz Gerstung, Philip H Jones.
      Aging epithelia are colonized by somatic mutations, which are subjected to selection influenced by intrinsic and extrinsic factors. The lack of suitable culture systems has slowed the study of this and other long-term biological processes. Here, we describe epithelioids, a facile, cost-effective method of culturing multiple mouse and human epithelia. Esophageal epithelioids self-maintain without passaging for at least 1 year, maintaining a three-dimensional structure with proliferative basal cells that differentiate into suprabasal cells, which eventually shed and retain genomic stability. Live imaging over 5 months showed that epithelioids replicate in vivo cell dynamics. Epithelioids support genetic manipulation and enable the study of mutant cell competition and selection in three-dimensional epithelia, and show how anti-cancer treatments modulate competition between transformed and wild-type cells. Finally, a targeted CRISPR-Cas9 screen shows that epithelioids recapitulate mutant gene selection in aging human esophagus and identifies additional drivers of clonal expansion, resolving the genetic networks underpinning competitive fitness.
    DOI:  https://doi.org/10.1038/s41588-024-01875-8
  8. Br J Cancer. 2024 Sep 25.
    Functional activation of the AKT-mTOR signalling axis in a real-world metastatic breast cancer cohort.
    Deepika Prasad, Elisa Baldelli, Edik M Blais, Justin Davis, Emna El Gazzah, Claudius Mueller, Alison Gomeiz, Aisha Ibrahim, Avani Vinayak Newrekar, Brian A Corgiat, Rick Dunetz, Emanuel F Petricoin Iii, Qi Wei, Mariaelena Pierobon.
      BACKGROUND: Mutations of the PIK3CA/AKT/mTOR axis are common events in metastatic breast cancers (MBCs). This study was designed to evaluate the extent to which genetic alterations of the PIK3CA/AKT/mTOR can predict protein activation of this signalling axis in MBCs.METHODS: Molecular profiles were generated by CLIA-certified laboratories from a real-world evidence cohort of 171 MBC patients. Genetic alterations of the PIK3CA pathway were measured using next-generation sequencing. Activation levels of AKT and downstream signalling molecules were quantified using two orthogonal proteomic methods. Protein activity was correlated with underlying genomic profiles and response to CDK4/6 inhibition in combination with endocrine treatment (ET).
    RESULTS: Oncogenic alterations of the PIK3CA/AKT/PTEN pathway were identified in 49.7% of cases. Genomic profiles emerged as poor predictors of protein activity (AUC:0.69), and AKT phosphorylation levels mimicked those of mutant lesions in 76.9% of wild-type tumours. High phosphorylation levels of the PI3K/AKT/mTOR downstream target p70S6 Kinase (T389) were associated with shorter PFS in patients treated with CDK4/6 inhibitors in combination with ET (HR:4.18 95%CI:1.19-14.63); this association was not seen when patients were classified by mutational status.
    CONCLUSIONS: Phosphoprotein-based measurements of drug targets and downstream substrates should be captured along with genomic information to identify MBCs driven by the PI3K/AKT/mTOR signalling.
    DOI:  https://doi.org/10.1038/s41416-024-02852-y
  9. Nat Protoc. 2024 Sep 20.
    Rapid generation of homozygous fluorescent knock-in human cells using CRISPR-Cas9 genome editing and validation by automated imaging and digital PCR screening.
    Andrea Callegari, Moritz Kueblbeck, Natalia Rosalía Morero, Beatriz Serrano-Solano, Jan Ellenberg.
      We previously described a protocol for genome engineering of mammalian cultured cells with clustered regularly interspaced short palindromic repeats and associated protein 9 (CRISPR-Cas9) to generate homozygous knock-ins of fluorescent tags into endogenous genes. Here we are updating this former protocol to reflect major improvements in the workflow regarding efficiency and throughput. In brief, we have improved our method by combining high-efficiency electroporation of optimized CRISPR-Cas9 reagents, screening of single cell-derived clones by automated bright-field and fluorescence imaging, rapidly assessing the number of tagged alleles and potential off-targets using digital polymerase chain reaction (PCR) and automated data analysis. Compared with the original protocol, our current procedure (1) substantially increases the efficiency of tag integration, (2) automates the identification of clones derived from single cells with correct subcellular localization of the tagged protein and (3) provides a quantitative and high throughput assay to measure the number of on- and off-target integrations with digital PCR. The increased efficiency of the new procedure reduces the number of clones that need to be analyzed in-depth by more than tenfold and yields to more than 26% of homozygous clones in polyploid cancer cell lines in a single genome engineering round. Overall, we were able to dramatically reduce the hands-on time from 30 d to 10 d during the overall ~10 week procedure, allowing a single person to process up to five genes in parallel, assuming that validated reagents-for example, PCR primers, digital PCR assays and western blot antibodies-are available.
    DOI:  https://doi.org/10.1038/s41596-024-01043-6
  10. Atherosclerosis. 2024 Jul 01. pii: S0021-9150(24)01097-9. [Epub ahead of print] 118529
    Engineering next generation vascularized organoids.
    Nicolas Werschler, Clement Quintard, Stephanie Nguyen, Josef Penninger.
      Organoids are self-organizing 3D cell culture models that are valuable for studying the mechanisms underlying both development and disease in multiple species, particularly, in humans. These 3D engineered tissues can mimic the structure and function of human organs in vitro. Methods to generate organoids have substantially improved to better resemble, in various ways, their in vivo counterpart. One of the major limitations in current organoid models is the lack of a functional vascular compartment. Here we discuss methodological approaches to generating perfusable blood vessel networks in organoid systems. Inclusion of perfused vascular compartments markedly enhances the physiological relevance of organoid systems and is a critical step in the establishment of next generation, higher-complexity in vitro systems for use in developmental, clinical, and drug-development settings.
    DOI:  https://doi.org/10.1016/j.atherosclerosis.2024.118529
  11. Nat Chem Biol. 2024 Sep 23.
    Functional profiling of serine, threonine and tyrosine sites.
    Yizhou Li, Tao Xu, Huazheng Ma, Di Yue, Qiezhong Lamao, Ying Liu, Zhuo Zhou, Wensheng Wei.
      Systematic perturbation of amino acids at endogenous loci provides diverse insights into protein function. Here, we performed a genome-wide screen to globally assess the cell fitness dependency of serine, threonine and tyrosine residues. Using an adenine base editor, we designed a whole-genome library comprising 817,089 single guide RNAs to perturb 584,337 S, T and Y sites. We identified 3,467 functional substitutions affecting cell fitness and 677 of them involving phosphorylation, including numerous phosphorylation-mediated gain-of-function substitutions that regulate phosphorylation levels of itself or downstream factors. Furthermore, our findings highlight that specific substitution types, notably serine to proline, are crucial for maintaining domain structure broadly. Lastly, we demonstrate that 309 enriched hits capable of initiating cell overproliferation might be potential cancer driver mutations. This study represents an extensive functional profiling of S, T and Y residues and provides insights into the distinctive roles of these amino acids in biological mechanisms and tumor progression.
    DOI:  https://doi.org/10.1038/s41589-024-01731-0
  12. bioRxiv. 2024 Sep 15. pii: 2024.09.14.613064. [Epub ahead of print]
    Stress granule formation enables anchorage-independence survival in cancer cells.
    Seungwon Yang, Anaïs Aulas, Paul J Anderson, Pavel Ivanov.
      Stress granules (SGs) are dynamic cytoplasmic structures assembled in response to various stress stimuli that enhance cell survival under adverse environmental conditions. Here we show that SGs contribute to breast cancer progression by enhancing the survival of cells subjected to anoikis stress. SG assembly is triggered by inhibition of Focal Adhesion Kinase (FAK) or loss of adhesion signals. Combined proteomic analysis and functional studies reveal that SG formation enhances cancer cell proliferation, resistance to metabolic stress, anoikis resistance, and migration. Importantly, inhibiting SG formation promotes the sensitivity of cancer cells to FAK inhibitors being developed as cancer therapeutics. Furthermore, we identify the Rho-ROCK- PERK-eIF2α axis as a critical signaling pathway activated by loss of adhesion signals and inhibition of the FAK-mTOR-eIF4F complex in breast cancer cells. By triggering SG assembly and AKT activation in response to anoikis stress, PERK functions as an oncoprotein in breast cancer cells. Overall, our study highlights the significance of SG formation in breast cancer progression and suggests that therapeutic inhibition of SG assembly may reverse anoikis resistance in treatment-resistant cancers such as triple-negative breast cancer (TNBC).Highlights: Either anoikis stress or loss of adhesion induce stress granule (SG) formationThe Rho-ROCK-PERK-eIF2α axis is a crucial signaling pathway triggered by the absence of adhesion signals, leading to the promotion of SG formation along with the inhibition of the FAK- AKT/mTOR-eIF4F complex under anoikis stress.PERK functions as an oncogene in breast cancer cells, initiating SG formation and activating AKT under anoikis stress.Inhibiting SG formation significantly enhances the sensitivity to Focal Adhesion Kinase (FAK) inhibitors, suggesting a potential for combined therapy to improve cancer treatment efficacy.
    DOI:  https://doi.org/10.1101/2024.09.14.613064
  13. Nat Rev Neurosci. 2024 Sep 26.
    Thicker perineuronal nets disrupt insulin signalling.
    Sian Lewis.
      
    DOI:  https://doi.org/10.1038/s41583-024-00872-4
  14. FEBS J. 2024 Sep 26.
    Senescence in the ageing skin: a new focus on mTORC1 and the lysosome.
    Phineas Smith, Bernadette Carroll.
      Ageing is defined as the progressive loss of tissue function and regenerative capacity and is caused by both intrinsic factors i.e. the natural accumulation of damage, and extrinsic factors i.e. damage from environmental stressors. Cellular senescence, in brief, is an irreversible exit from the cell cycle that occurs primarily in response to excessive cellular damage, such as from ultraviolet (UV) exposure and oxidative stress, and it has been comprehensively demonstrated to contribute to tissue and organismal ageing. In this review, we will focus on the skin, an organ which acts as an essential protective barrier against injury, insults, and infection. We will explore the evidence for the existence and contribution of cellular senescence to skin ageing. We discuss the known molecular mechanisms driving senescence in the skin, with a focus on the dysregulation of the master growth regulator, mechanistic Target of Rapamycin Complex 1 (mTORC1). We explore the interplay of dysregulated mTORC1 with lysosomes and how they contribute to senescence phenotypes.
    Keywords:  ageing; lysosome; mTORC1; senescence; skin
    DOI:  https://doi.org/10.1111/febs.17281
  15. Cell. 2024 Sep 19. pii: S0092-8674(24)01028-6. [Epub ahead of print]
    It took a long, long time: Ras and the race to cure cancer.
    Robert A Weinberg.
      
    DOI:  https://doi.org/10.1016/j.cell.2024.09.010
  16. Oncogene. 2024 09 22.
    MYC upstream region orchestrates resistance to PI3K inhibitors in cancer cells through FOXO3a-mediated autophagic adaptation.
    Rosa Bordone, Devon Michael Ivy, Rodrigo D'Amico, Martina Barba, Miriam Gaggianesi, Fiorella Di Pastena, Bianca Cesaro, Francesca Bufalieri, Alessio Balzerano, Enrico De Smaele, Giuseppe Giannini, Lucia Di Marcotullio, Alessandro Fatica, Giorgio Stassi, Laura Di Magno, Sonia Coni, Gianluca Canettieri.
      The MYC oncogene is frequently overexpressed in tumors and inhibition of its translation is considered an attractive therapeutic opportunity. Despite numerous reports proposing an internal ribosome entry site (IRES) within the MYC Upstream Region (MYC UR) to sustain MYC translation during cellular stress or chemotherapy, conflicting evidence remains regarding the validity of such a mechanism. Through comprehensive investigations in MYC-driven Colorectal Cancer (CRC) and Burkitt Lymphoma (BL) cells, we demonstrate that MYC UR does not facilitate cap-independent translation, but instead orchestrates resistance to PI3K inhibitors. Genomic deletion of MYC UR neither impacts MYC protein levels nor viability in CRC cells, either untreated or exposed to cellular stress. However, in response to PI3K inhibitors, MYC UR drives a FOXO3a-dependent transcriptional upregulation of MYC, conferring drug resistance. This resistance is mediated by enhanced autophagic flux, governed by MYC, and blockade of autophagy sensitizes CRC cells to PI3K inhibition in vitro and in vivo. Remarkably, BL cells lacking the translocation of MYC UR exhibit sensitivity to PI3K inhibitors, whereas MYC UR-translocated cells respond to these drugs only when autophagy is inhibited. These findings challenge previous notions regarding IRES-mediated translation and highlight a promising strategy to overcome resistance to PI3K inhibitors in MYC-driven malignancies, offering potential clinical implications for CRC and BL treatment.
    DOI:  https://doi.org/10.1038/s41388-024-03170-6
  17. Trends Biochem Sci. 2024 Sep 23. pii: S0968-0004(24)00192-0. [Epub ahead of print]
    The inositol phosphate signalling network in physiology and disease.
    Seyun Kim, Rashna Bhandari, Charles A Brearley, Adolfo Saiardi.
      Combinatorial substitution of phosphate groups on the inositol ring gives rise to a plethora of inositol phosphates (InsPs) and inositol pyrophosphates (PP-InsPs). These small molecules constitute an elaborate metabolic and signalling network that influences nearly every cellular function. This review delves into the knowledge accumulated over the past decades regarding the biochemical principles and significance of InsP metabolism. We focus on the biological actions of InsPs in mammals, with an emphasis on recent findings regarding specific target proteins. We further discuss the roles of InsP metabolism in contributing to physiological homeostasis and pathological conditions. A deeper understanding of InsPs and their metabolic pathways holds the potential to address unresolved questions and propel advances towards therapeutic applications.
    Keywords:  kinase; metabolism; phosphatase; physiology; signal transduction
    DOI:  https://doi.org/10.1016/j.tibs.2024.08.005
  18. Nature. 2024 Sep 25.
    Designed endocytosis-inducing proteins degrade targets and amplify signals.
    Buwei Huang, Mohamad Abedi, Green Ahn, Brian Coventry, Isaac Sappington, Cong Tang, Rong Wang, Thomas Schlichthaerle, Jason Z Zhang, Yujia Wang, Inna Goreshnik, Ching Wen Chiu, Adam Chazin-Gray, Sidney Chan, Stacey Gerben, Analisa Murray, Shunzhi Wang, Jason O'Neill, Li Yi, Ronald Yeh, Ayesha Misquith, Anitra Wolf, Luke M Tomasovic, Dan I Piraner, Maria J Duran Gonzalez, Nathaniel R Bennett, Preetham Venkatesh, Maggie Ahlrichs, Craig Dobbins, Wei Yang, Xinru Wang, Danny D Sahtoe, Dionne Vafeados, Rubul Mout, Shirin Shivaei, Longxing Cao, Lauren Carter, Lance Stewart, Jamie B Spangler, Kole T Roybal, Per Jr Greisen, Xiaochun Li, Gonçalo J L Bernardes, Carolyn R Bertozzi, David Baker.
      Endocytosis and lysosomal trafficking of cell surface receptors can be triggered by endogenous ligands. Therapeutic approaches such as lysosome-targeting chimaeras1,2 (LYTACs) and cytokine receptor-targeting chimeras3 (KineTACs) have used this to target specific proteins for degradation by fusing modified native ligands to target binding proteins. Although powerful, these approaches can be limited by competition with native ligands and requirements for chemical modification that limit genetic encodability and can complicate manufacturing, and, more generally, there may be no native ligands that stimulate endocytosis through a given receptor. Here we describe computational design approaches for endocytosis-triggering binding proteins (EndoTags) that overcome these challenges. We present EndoTags for insulin-like growth factor 2 receptor (IGF2R) and asialoglycoprotein receptor (ASGPR), sortilin and transferrin receptors, and show that fusing these tags to soluble or transmembrane target protein binders leads to lysosomal trafficking and target degradation. As these receptors have different tissue distributions, the different EndoTags could enable targeting of degradation to different tissues. EndoTag fusion to a PD-L1 antibody considerably increases efficacy in a mouse tumour model compared to antibody alone. The modularity and genetic encodability of EndoTags enables AND gate control for higher-specificity targeted degradation, and the localized secretion of degraders from engineered cells. By promoting endocytosis, EndoTag fusion increases signalling through an engineered ligand-receptor system by nearly 100-fold. EndoTags have considerable therapeutic potential as targeted degradation inducers, signalling activators for endocytosis-dependent pathways, and cellular uptake inducers for targeted antibody-drug and antibody-RNA conjugates.
    DOI:  https://doi.org/10.1038/s41586-024-07948-2
  19. Brain. 2024 Sep 26. pii: awae306. [Epub ahead of print]
    PTEN controls alternative splicing of autism spectrum disorder-associated transcripts in primary neurons.
    Sebastian Rademacher, Marco Preußner, Marie C Rehm, Joachim Fuchs, Florian Heyd, Britta J Eickholt.
      Phosphatase and tensin homologue (PTEN) is the main antagonist of the phosphatidylinositol-3-kinase (PI3K)/AKT/mTOR signalling pathway and mutated in 10-20% of individuals with autism spectrum disorder (ASD) exhibiting macrocephaly. Hyperactive mTOR signalling is responsible for some aspects during PTEN-ASD progression, e.g. neuronal hypertrophy and -excitability, but PI3K/mTOR-independent processes have additionally been described. There is emerging evidence that PTEN regulates gene transcription, spliceosome formation and pre-mRNA splicing independently of PI3K/mTOR. Altered splicing is a hallmark of brains from individuals with idiopathic and PTEN-ASD, however, molecular mechanisms are yet to be identified. We performed RNA-Seq followed by analysis of altered transcript splicing in Pten-deficient primary cortical mouse neurons, which we compared with published data from PTEN-deficient human neuronal stem cells. This analysis identified that transcripts were globally mis-spliced in a developmentally regulated fashion and cluster in synaptic and gene expression regulatory processes. Strikingly, splicing defects following Pten-deficiency represent a significant number of other known ASD-susceptibility genes. Furthermore, we show that exons with strong 3' splice sites are more frequently mis-spliced under Pten-deficient conditions. Our study indicates that PTEN-ASD is a multifactorial condition involving the dysregulation of other known ASD-susceptibility genes.
    DOI:  https://doi.org/10.1093/brain/awae306
  20. Cell Rep Methods. 2024 Sep 17. pii: S2667-2375(24)00242-X. [Epub ahead of print] 100864
    Recovering single-cell expression profiles from spatial transcriptomics with scResolve.
    Hao Chen, Young Je Lee, Jose A Ovando-Ricardez, Lorena Rosas, Mauricio Rojas, Ana L Mora, Ziv Bar-Joseph, Jose Lugo-Martinez.
      Many popular spatial transcriptomics techniques lack single-cell resolution. Instead, these methods measure the collective gene expression for each location from a mixture of cells, potentially containing multiple cell types. Here, we developed scResolve, a method for recovering single-cell expression profiles from spatial transcriptomics measurements at multi-cellular resolution. scResolve accurately restores expression profiles of individual cells at their locations, which is unattainable with cell type deconvolution. Applications of scResolve on human breast cancer data and human lung disease data demonstrate that scResolve enables cell-type-specific differential gene expression analysis between different tissue contexts and accurate identification of rare cell populations. The spatially resolved cellular-level expression profiles obtained through scResolve facilitate more flexible and precise spatial analysis that complements raw multi-cellular level analysis.
    Keywords:  CP: Systems biology; cell segmentation; cellular senescence; spatial differential expression; spatial transcriptomics; super-resolution
    DOI:  https://doi.org/10.1016/j.crmeth.2024.100864
  21. Eur J Cell Biol. 2024 Sep 18. pii: S0171-9335(24)00074-8. [Epub ahead of print]103(4): 151457
    Insulin receptor substrate 1 is a novel member of EGFR signaling in pancreatic cells.
    Tamás Takács, Loretta László, Álmos Tilajka, Julianna Novák, László Buday, Virag Vas.
      Pancreatic ductal adenocarcinoma is an extremely incurable cancer type characterized by cells with highly proliferative capacity and resistance against the current therapeutic options. Our study reveals that IRS1 acts as a bridging molecule between EGFR and IGFR/InsR signalization providing a potential mechanism for the interplay between signaling pathways and bypassing EGFR-targeted or IGFR/InsR-targeted therapies. The analysis of IRS1 phosphorylation status in four pancreatic cell lines identified the impact of EGFR signaling on IRS1 activation in comparison with InsR/IGFR signaling. Significantly reduced viability was observed in IRS1-silenced cells even upon EGF stimulation showing the critical role of IRS1 in the EGFR signaling network in both malignant and normal pancreatic cells. This study also demonstrated that EGFR binds directly to IRS1 and at least on two tyrosine sites, Y612 and Y896, IRS1 becomes phosphorylated in response to EGF stimulation. Mechanistically, the EGFR-mediated phosphorylation of IRS1 can further activate the MAPK signaling pathway with the recruitment of GRB2 protein. Collectively, in this study, IRS1 was identified as a crucial regulator in the EGFR signaling suggesting IRS1 as a potential target for more durable responses to targeted PDAC therapy.
    Keywords:  EGF; EGF receptor; GRB2; IRS1; Pancreatic adenocarcinoma; Signaling crosstalk; Tyrosine kinase
    DOI:  https://doi.org/10.1016/j.ejcb.2024.151457
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