bims-cagime 2024-02-25 papers

bims-cagime

Biomed News

on Cancer, aging and metabolism

Issue of 2024–02–25
forty-one papers selected by
Kıvanç Görgülü, Technical University of Munich

UDP-glucose dehydrogenase supports autophagy-deficient PDAC growth via increasing hyaluronic acid biosynthesis.
Clearance of dead cells by efferocytosis promotes liver metastasis in pancreatic cancer.
Exosomes define a local and systemic communication network in healthy pancreas and pancreatic ductal adenocarcinoma.
Inhibition of ULK1/2 and KRAS G12C controls tumor growth in preclinical models of lung cancer.
Selective degradation of ribosomes during oncogene-induced senescence: molecular insights and biological perspectives.
SREBP-dependent regulation of lipid homeostasis is required for progression and growth of pancreatic ductal adenocarcinoma.
Genome-wide screens identify SEL1L as an intracellular rheostat controlling collagen turnover.
Mesothelin CAR T-cells secreting anti-FAP/anti-CD3 molecules efficiently target pancreatic adenocarcinoma and its stroma.
Plasma membrane damage causes cellular senescence.
Substrate stiffness modulates the emergence and magnitude of senescence phenotypes.
Hypoxia at 3D organoid establishment selects essential subclones within heterogenous pancreatic cancer.
Epithelial and Mesenchymal-like Pancreatic Cancer Cells Exhibit Different Stem Cell Phenotypes Associated with Different Metastatic Propensities.
Clinical Evaluation of the Pancreatic Cancer Microenvironment: Opportunities and Challenges.
Multidimensional screening of pancreatic cancer spheroids reveals vulnerabilities in mitotic and cell-matrix adhesion signaling that associate with metastatic progression and decreased patient survival.
The enzymes of serine synthesis pathway in cancer metastasis.
Genetically encoded fluorescent sensor to monitor intracellular arginine methylation.
Drosophila TMEM63 and mouse TMEM63A are lysosomal mechanosensory ion channels.
Identification of MIMAS, a multifunctional mega-assembly integrating metabolic and respiratory biogenesis factors of mitochondria.
IgG is an aging factor that drives adipose tissue fibrosis and metabolic decline.
Fateful fat: Intra-pancreatic lipids cause pancreatic cancer.
Mechanism and cellular function of direct membrane binding by the ESCRT and ERES-associated Ca2+-sensor ALG-2.
Annexins-a family of proteins with distinctive tastes for cell signaling and membrane dynamics.
Cellular senescence: Neither irreversible nor reversible.
Lysosomal channels sensing forces.
Plasma membrane damage limits replicative lifespan in yeast and induces premature senescence in human fibroblasts.
PC4: A new regulator of cyclin D1 transcript levels.
Plasticity-induced repression of Irf6 underlies acquired resistance to cancer immunotherapy in pancreatic ductal adenocarcinoma.
Chromosome evolution screens recapitulate tissue-specific tumor aneuploidy patterns.
Recording physiological history of cells with chemical labeling.
Reporter cell lines to screen for inhibitors or regulators of the KRAS-RAF-MEK1/2-ERK1/2 pathway.
Phase-shifting optothermal microscopy enables live-cell mid-infrared hyperspectral imaging of large cell populations at high confluency.
ColabSeg: An interactive tool for editing, processing, and visualizing membrane segmentations from cryo-ET data.
mTORC1 activity oscillates throughout the cell cycle promoting mitotic entry and differentially influencing autophagy induction.
ITIH5 as a multifaceted player in pancreatic cancer suppression, impairing tyrosine kinase signaling, cell adhesion and migration.
Asymmetric oligomerization state and sequence patterning can tune multiphase condensate miscibility.
Magnetic-optical dual-modality imaging monitoring chemotherapy efficacy of pancreatic ductal adenocarcinoma with a low-dose fibronectin-targeting Gd-based contrast agent.
NINJ1 induces plasma membrane rupture and release of damage-associated molecular pattern molecules during ferroptosis.
Longitudinal Fragility Phenotyping Predicts Lifespan and Age-Associated Morbidity in C57BL/6 and Diversity Outbred Mice.
Fusion genes in pancreatic tumors.
Sleep and risk of pancreatic cancer in the UK Biobank.
PARP1-targeted fluorescence molecular endoscopy as novel tool for early detection of esophageal dysplasia and adenocarcinoma.

Cell Rep. 2024 Feb 16. pii: S2211-1247(24)00136-0. [Epub ahead of print]43(2): 113808

UDP-glucose dehydrogenase supports autophagy-deficient PDAC growth via increasing hyaluronic acid biosynthesis.

Minghe Fan, Sihan Huo, Yuyao Guo, Ruoxuan Wang, Wenqin Hao, Ziyang Zhang, Lina Wang, Ying Zhao.

  Autophagy is an essential degradation and recycling process that maintains cellular homeostasis during stress or nutrient deprivation. However, certain types of tumors such as pancreatic cancers can circumvent autophagy inhibition to sustain growth. The mechanism that autophagy-deficient pancreatic ductal adenocarcinoma (PDAC) uses to grow under nutrient deprivation is poorly understood. Our data show that nutrient deprivation in PDAC results in UDP-glucose dehydrogenase (UGDH) degradation, which is dependent on autophagic cargo receptor sequestosome 1 (p62). Moreover, we demonstrate that accumulated UGDH is indispensable for autophagy-deficient PDAC cells proliferation by promoting hyaluronic acid (HA) synthesis upon energy deprivation. Using an orthotopic mouse model of PDAC, we find that inhibition of HA synthesis by targeting UGDH in PDAC reduces tumor weight. Thus, the combined inhibition of HA and autophagy might be an attractive strategy for PDAC treatment.

Keywords:  CP: Cancer; CP: Metabolism; HA; PDAC; UDP-glucose dehydrogenase; UGDH; autophagy; hyaluronic acid; p62; pancreatic ductal adenocarcinoma; sequestosome 1

DOI:  https://doi.org/10.1016/j.celrep.2024.113808
Nat Cancer. 2024 Feb 20.

Clearance of dead cells by efferocytosis promotes liver metastasis in pancreatic cancer.

DOI: https://doi.org/10.1038/s43018-024-00732-1
Nat Commun. 2024 Feb 21. 15(1): 1496

Exosomes define a local and systemic communication network in healthy pancreas and pancreatic ductal adenocarcinoma.

Bárbara Adem, Nuno Bastos, Carolina F Ruivo, Sara Sousa-Alves, Carolina Dias, Patrícia F Vieira, Inês A Batista, Bruno Cavadas, Dieter Saur, José C Machado, Dawen Cai, Sonia A Melo.

Pancreatic ductal adenocarcinoma (PDAC), a lethal disease, requires a grasp of its biology for effective therapies. Exosomes, implicated in cancer, are poorly understood in living systems. Here we use the genetically engineered mouse model (ExoBow) to map the spatiotemporal distribution of exosomes from healthy and PDAC pancreas in vivo to determine their biological significance. We show that, within the PDAC microenvironment, cancer cells establish preferential communication routes through exosomes with cancer associated fibroblasts and endothelial cells. The latter being a conserved event in the healthy pancreas. Inhibiting exosomes secretion in both scenarios enhances angiogenesis, underscoring their contribution to vascularization and to cancer. Inter-organ communication is significantly increased in PDAC with specific organs as most frequent targets of exosomes communication occurring in health with the thymus, bone-marrow, brain, and intestines, and in PDAC with the kidneys, lungs and thymus. In sum, we find that exosomes mediate an organized intra- and inter- pancreas communication network with modulatory effects in vivo.

DOI: https://doi.org/10.1038/s41467-024-45753-7
bioRxiv. 2024 Feb 08. pii: 2024.02.06.579200. [Epub ahead of print]

Inhibition of ULK1/2 and KRAS G12C controls tumor growth in preclinical models of lung cancer.

Phaedra C Ghazi, Kayla T O'Toole, Sanjana Srinivas Boggaram, Michael T Scherzer, Mark R Silvis, Yun Zhang, Madhumita Bogdan, Bryan D Smith, Guillermina Lozano, Daniel L Flynn, Eric L Snyder, Conan G Kinsey, Martin McMahon.

Mutational activation of KRAS occurs commonly in lung carcinogenesis and, with the recent FDA approval of covalent inhibitors of KRAS G12C such as sotorasib or adagrasib, KRAS oncoproteins are important pharmacological targets in non-small cell lung cancer (NSCLC). However, not all KRAS G12C -driven NSCLCs respond to these inhibitors, and the emergence of drug resistance in those patients that do respond can be rapid and pleiotropic. Hence, based on a backbone of covalent inhibition of KRAS G12C , efforts are underway to develop effective combination therapies. Here we report that inhibition of KRAS G12C signaling increases autophagy in KRAS G12C expressing lung cancer cells. Moreover, the combination of DCC-3116, a selective ULK1/2 inhibitor, plus sotorasib displays cooperative/synergistic suppression of human KRAS G12C -driven lung cancer cell proliferation in vitro and superior tumor control in vivo . Additionally, in genetically engineered mouse models of KRAS G12C -driven NSCLC, inhibition of either KRAS G12C or ULK1/2 decreases tumor burden and increases mouse survival. Consequently, these data suggest that ULK1/2-mediated autophagy is a pharmacologically actionable cytoprotective stress response to inhibition of KRAS G12C in lung cancer.

DOI: https://doi.org/10.1101/2024.02.06.579200
Autophagy. 2024 Feb 21. 1-3

Selective degradation of ribosomes during oncogene-induced senescence: molecular insights and biological perspectives.

Aida Rodríguez López, Lisa B Frankel.

  Ribosomes are conserved macromolecular machines that are responsible for protein synthesis in all cells. While our knowledge of ribosome biogenesis and function has increased significantly in recent years, little is known about how ribosomes are degraded under specific cellular conditions. We recently uncovered that ribosomes are efficiently turned over by selective macroautophagy/autophagy during oncogene-induced senescence (OIS). By profiling the ribosome interactome in human fibroblasts undergoing OIS, we discovered a key role for the de-ubiquitinating enzyme USP10 in guiding this process. Release of USP10 from ribosomes during senescence leads to their enhanced ubiquitination and selective sequestering by autophagy through the SQSTM1/p62 receptor protein. This process is important for sustaining senescence-associated metabolome and secretome alterations.

Keywords:  Oncogene-induced senescence; USP10; ribosomes; selective autophagy; translation; ubiquitination

DOI:  https://doi.org/10.1080/15548627.2024.2319022
bioRxiv. 2024 Feb 08. pii: 2024.02.04.578802. [Epub ahead of print]

SREBP-dependent regulation of lipid homeostasis is required for progression and growth of pancreatic ductal adenocarcinoma.

Chiaki T Ishida, Stephanie L Myers, Wei Shao, Meredith R McGuire, Chune Liu, Casie S Kubota, Theodore E Ewachiw, Debaditya Mukhopadhyay, Suqi Ke, Hao Wang, Zeshaan A Rasheed, Robert A Anders, Peter J Espenshade.

Metabolic reprogramming is a necessary component of oncogenesis and cancer progression that solid tumors undergo when their growth outstrips local nutrient supply. The supply of lipids such as cholesterol and fatty acids is required for continued tumor cell proliferation, and oncogenic mutations stimulate de novo lipogenesis to support tumor growth. Sterol regulatory element-binding protein (SREBP) transcription factors control cellular lipid homeostasis by activating genes required for lipid synthesis and uptake. SREBPs have been implicated in the progression of multiple cancers, including brain, breast, colon, liver, and prostate. However, the role the SREBP pathway and its central regulator SREBP cleavage activating protein (SCAP) in pancreatic ductal adenocarcinoma (PDAC) has not been studied in detail. Here, we demonstrated that pancreas-specific knockout of Scap has no effect on mouse pancreas development or function, allowing for examination of the role for Scap in the murine KPC model of PDAC. Notably, heterozygous loss of Scap prolonged survival in KPC mice, and homozygous loss of Scap impaired PDAC tumor progression. Using subcutaneous and orthotopic xenograft models, we showed that S CAP is required for human PDAC tumor growth. Mechanistically, chemical or genetic inhibition of the SREBP pathway prevented PDAC cell growth under low serum conditions due to a lack of lipid supply. Highlighting the clinical importance of this pathway, the SREBP pathway is broadly required for cancer cell growth, SREBP target genes are upregulated in human PDAC tumors, and increased expression of SREBP targets genes is associated with poor survival in PDAC patients. Collectively, these results demonstrate that SCAP and the SREBP pathway activity are essential for PDAC cell and tumor growth in vitro and in vivo , identifying SCAP as a potential therapeutic target for PDAC.
SIGNIFICANCE: Our findings demonstrate that SREBP pathway activation is a critical part of the metabolic reprogramming that occurs in PDAC development and progression. Therefore, targeting the SREBP pathway has significant therapeutic potential.

DOI: https://doi.org/10.1101/2024.02.04.578802
Nat Commun. 2024 Feb 20. 15(1): 1531

Genome-wide screens identify SEL1L as an intracellular rheostat controlling collagen turnover.

Michael J Podolsky, Benjamin Kheyfets, Monika Pandey, Afaq H Beigh, Christopher D Yang, Carlos O Lizama, Ritwik Datta, Liangguang L Lin, Zhihong Wang, Paul J Wolters, Michael T McManus, Ling Qi, Kamran Atabai.

Accumulating evidence has implicated impaired extracellular matrix (ECM) clearance as a key factor in fibrotic disease. Despite decades of research elucidating the effectors of ECM clearance, relatively little is understood regarding the upstream regulation of this process. Collagen is the most abundant constituent of normal and fibrotic ECM in mammalian tissues. Its catabolism occurs through extracellular proteolysis and cell-mediated uptake of collagen fragments for intracellular degradation. Given the paucity of information regarding the regulation of this latter process, here we execute unbiased genome-wide screens to understand the molecular underpinnings of cell-mediated collagen clearance. Using this approach, we discover a mechanism through which collagen biosynthesis is sensed by cells internally and directly regulates clearance of extracellular collagen. The sensing mechanism appears to be dependent on endoplasmic reticulum-resident protein SEL1L and occurs via a noncanonical function of this protein. This pathway functions as a homeostatic negative feedback loop that limits collagen accumulation in tissues. In human fibrotic lung disease, the induction of this collagen clearance pathway by collagen synthesis is impaired, thereby contributing to the pathological accumulation of collagen in lung tissue. Thus, we describe cell-autonomous, rheostatic collagen clearance as an important pathway of tissue homeostasis.

DOI: https://doi.org/10.1038/s41467-024-45817-8
Clin Cancer Res. 2024 Feb 23.

Mesothelin CAR T-cells secreting anti-FAP/anti-CD3 molecules efficiently target pancreatic adenocarcinoma and its stroma.

Marc Wehrli, Samantha Guinn, Filippo Birocchi, Adam Kuo, Yi Sun, Rebecca C Larson, Antonio J Almazan, Irene Scarfò, Amanda A Bouffard, Stefanie R Bailey, Praju Vikas Anekal, Paula Montero Lopis, Linda T Nieman, Yuhui Song, Katherine H Xu, Trisha R Berger, Michael C Kann, Mark B Leick, Harrison Silva, Diego Salas-Benito, Tamina Kienka, Korneel Grauwet, Todd D Armstrong, Rui Zhang, Qingfeng Zhu, Juan Fu, Andrea Schmidts, Felix Korell, Max Jan, Bryan D Choi, Andrew S Liss, Genevieve M Boland, David T Ting, Richard A Burkhart, Russell W Jenkins, Lei Zheng, Elizabeth M Jaffee, Jacquelyn W Zimmerman, Marcela V Maus.

PURPOSE: Targeting solid tumors with CAR T-cells remains challenging due to heterogenous target antigen expression, antigen escape, and the immunosuppressive tumor microenvironment (TME). Pancreatic cancer is characterized by a thick stroma generated by cancer-associated fibroblasts (CAFs), which may contribute to the limited efficacy of mesothelin-directed CAR T-cells in early-phase clinical trials. To provide a more favorable TME for CAR T-cells to target pancreatic ductal adenocarcinoma (PDAC), we generated T-cells with an anti-mesothelin CAR and a secreted T-cell-engaging molecule (TEAM) that targets CAFs through fibroblast activation protein (FAP) and engages T-cells through CD3 (termed mesoFAP CAR-TEAM cells).
EXPERIMENTAL DESIGN: Using a suite of in vitro, in vivo, and ex vivo patient-derived models containing cancer cells and CAFs, we examined the ability of mesoFAP CAR-TEAM cells to target PDAC cells and CAFs within the TME. We developed and used patient-derived ex vivo models including patient-derived organoids with patient-matched CAFs and patient-derived organotypic tumor spheroids (PDOTS).
RESULTS: We demonstrated specific and significant binding of the TEAM to its respective antigens (CD3 and FAP) when released from mesothelin-targeting CAR T cells, leading to T cell activation and cytotoxicity of the target cell. MesoFAP CAR-TEAM cells were superior in eliminating PDAC and CAFs compared to T cells engineered to target either antigen alone in our ex-vivo patient-derived models and in mouse models of PDAC with primary or metastatic liver tumors.
CONCLUSIONS: CAR-TEAM cells enable modification of tumor stroma, leading to increased elimination of PDAC tumors. This approach represents a promising treatment option for pancreatic cancer.

DOI: https://doi.org/10.1158/1078-0432.CCR-23-3841
Nat Aging. 2024 Feb 22.

Plasma membrane damage causes cellular senescence.

Stine Lauritzen Sønder, Jesper Nylandsted.

DOI: https://doi.org/10.1038/s43587-024-00584-5
bioRxiv. 2024 Feb 07. pii: 2024.02.06.579151. [Epub ahead of print]

Substrate stiffness modulates the emergence and magnitude of senescence phenotypes.

Bartholomew Starich, Fan Yang, Derin Tanrioven, Heng-Chung Kung, Joanne Baek, Praful R Nair, Pratik Kamat, Nico Macaluso, Joon Eoh, Kyu Sang Han, Luo Gu, Sean Sun, Pei-Hsun Wu, Denis Wirtz, Jude M Phillip.

Cellular senescence is a major driver of aging and disease. Here we show that substrate stiffness modulates the emergence and magnitude of senescence phenotypes post induction. Using a primary dermal fibroblast model of senescence, we show that decreased substrate stiffness accelerates cell-cycle arrest during senescence development and regulate expression of conventional protein-based biomarkers of senescence. We found that the expression of these senescence biomarkers, namely p21 WAF1/CIP1 ( CDKN1a ) and p16 INK4a ( CDKN2a ) are mechanosensitive and are in-part regulated by myosin contractility through focal adhesion kinase (FAK)-ROCK signaling. Interestingly, at the protein level senescence-induced dermal fibroblasts on soft substrates (0.5 kPa) do not express p21 WAF1/CIP1 and p16 INK4a at comparable levels to induced cells on stiff substrates (4GPa). However, cells do express CDKN1a, CDKN2a, and IL6 at the RNA level across both stiff and soft substrates. When cells were transferred from soft to stiff substrates, senescent cells recover an elevated expression expressing p21 WAF1/CIP1 and p16 INK4a at levels comparable to senescence cells on stiff substrates, pointing to a mechanosensitive regulation of the senescence phenotypes. Together, our results indicate that the induction of senescence programs depends critically on the mechanical environments of cells and that senescent cells actively respond and adapt to changing mechanical cues.

DOI: https://doi.org/10.1101/2024.02.06.579151
Front Cell Dev Biol. 2024 ;12 1327772

Hypoxia at 3D organoid establishment selects essential subclones within heterogenous pancreatic cancer.

Koichiro Kumano, Hiromitsu Nakahashi, Pakavarin Louphrasitthiphol, Yukihito Kuroda, Yoshihiro Miyazaki, Osamu Shimomura, Shinji Hashimoto, Yoshimasa Akashi, Bryan J Mathis, Jaejeong Kim, Yohei Owada, Colin R Goding, Tatsuya Oda.

  Pancreatic ductal adenocarcinoma (PDAC) is especially hypoxic and composed of heterogeneous cell populations containing hypoxia-adapted cells. Hypoxia as a microenvironment of PDAC is known to cause epithelial-mesenchymal transition (EMT) and resistance to therapy. Therefore, cells adapted to hypoxia possess malignant traits that should be targeted for therapy. However, current 3D organoid culture systems are usually cultured under normoxia, losing hypoxia-adapted cells due to selectivity bias at the time of organoid establishment. To overcome any potential selection bias, we focused on oxygen concentration during the establishment of 3D organoids. We subjected identical PDAC surgical samples to normoxia (O2 20%) or hypoxia (O2 1%), yielding glandular and solid organoid morphology, respectively. Pancreatic cancer organoids established under hypoxia displayed higher expression of EMT-related proteins, a Moffitt basal-like subtype transcriptome, and higher 5-FU resistance in contrast to organoids established under normoxia. We suggest that hypoxia during organoid establishment efficiently selects for hypoxia-adapted cells possibly responsible for PDAC malignant traits, facilitating a fundamental source for elucidating and developing new treatment strategies against PDAC.

Keywords:  3D organoid; hypoxia; pancreatic cancer; selection bias; tumor heterogeneity

DOI:  https://doi.org/10.3389/fcell.2024.1327772
Cancers (Basel). 2024 Feb 06. pii: 686. [Epub ahead of print]16(4):

Epithelial and Mesenchymal-like Pancreatic Cancer Cells Exhibit Different Stem Cell Phenotypes Associated with Different Metastatic Propensities.

Lisa-Marie Philipp, Umut-Ulas Yesilyurt, Arne Surrow, Axel Künstner, Anne-Sophie Mehdorn, Charlotte Hauser, Jan-Paul Gundlach, Olga Will, Patrick Hoffmann, Lea Stahmer, Sören Franzenburg, Hendrike Knaack, Udo Schumacher, Hauke Busch, Susanne Sebens.

  Pancreatic ductal adenocarcinoma (PDAC) is mostly diagnosed at advanced or even metastasized stages, limiting the prognoses of patients. Metastasis requires high tumor cell plasticity, implying phenotypic switching in response to changing environments. Here, epithelial-mesenchymal transition (EMT), being associated with an increase in cancer stem cell (CSC) properties, and its reversion are important. Since it is poorly understood whether different CSC phenotypes exist along the EMT axis and how these impact malignancy-associated properties, we aimed to characterize CSC populations of epithelial and mesenchymal-like PDAC cells. Single-cell cloning revealed CSC (Holoclone) and non-CSC (Paraclone) clones from the PDAC cell lines Panc1 and Panc89. The Panc1 Holoclone cells showed a mesenchymal-like phenotype, dominated by a high expression of the stemness marker Nestin, while the Panc89 Holoclone cells exhibited a SOX2-dominated epithelial phenotype. The Panc89 Holoclone cells showed enhanced cell growth and a self-renewal capacity but slow cluster-like invasion. Contrarily, the Panc1 Holoclone cells showed slower cell growth and self-renewal ability but were highly invasive. Moreover, cell variants differentially responded to chemotherapy. In vivo, the Panc1 and Panc89 cell variants significantly differed regarding the number and size of metastases, as well as organ manifestation, leading to different survival outcomes. Overall, these data support the existence of different CSC phenotypes along the EMT axis in PDAC, manifesting different metastatic propensities.

Keywords:  EMT; PDAC; adhesion; cancer stem cells; epithelial–mesenchymal-transition; heterogeneity; invasion; metastasis; migration; pancreatic adenocarcinoma; plasticity

DOI:  https://doi.org/10.3390/cancers16040686
Cancers (Basel). 2024 Feb 15. pii: 794. [Epub ahead of print]16(4):

Clinical Evaluation of the Pancreatic Cancer Microenvironment: Opportunities and Challenges.

Julianne M Szczepanski, Mark A Rudolf, Jiaqi Shi.

  Advances in our understanding of pancreatic ductal adenocarcinoma (PDAC) and its tumor microenvironment (TME) have the potential to transform treatment for the hundreds of thousands of patients who are diagnosed each year. Whereas the clinical assessment of cancer cell genetics has grown increasingly sophisticated and personalized, current protocols to evaluate the TME have lagged, despite evidence that the TME can be heterogeneous within and between patients. Here, we outline current protocols for PDAC diagnosis and management, review novel biomarkers, and highlight potential opportunities and challenges when evaluating the PDAC TME as we prepare to translate emerging TME-directed therapies to the clinic.

Keywords:  biomarker; management; pancreatic ductal adenocarcinoma; pathology; tumor microenvironment

DOI:  https://doi.org/10.3390/cancers16040794
Biochem Biophys Res Commun. 2024 Feb 06. pii: S0006-291X(24)00110-4. [Epub ahead of print]703 149575

Multidimensional screening of pancreatic cancer spheroids reveals vulnerabilities in mitotic and cell-matrix adhesion signaling that associate with metastatic progression and decreased patient survival.

Albert-Fred Aquino, Farhana Runa, Jannatul F Shoma, Audrey Todd, Matthew Wallace, Natan Roberto de Barros, Jonathan A Kelber.

  Pancreatic ductal adenocarcinoma (PDAC) is a highly aggressive malignancy, with a median survival of less than 12 months and a 5-year survival of less than 10 %. Here, we have established an image-based screening pipeline for quantifying single PDAC spheroid dynamics in genetically and phenotypically diverse PDAC cell models. Wild-type KRas PDAC cells formed tight/compact spheroids - compaction of these structures was completely blocked by cytoplasmic dynein and focal adhesion kinase (FAK) inhibitors. In contrast, PDAC cells containing mutant KRas formed loosely aggregated spheroids that grew significantly slower following inhibition of polo-like kinase 1 (PLK1) or focal adhesion kinase (FAK). Independent of genetic background, multicellular PDAC-mesenchymal stromal cell (MSC) spheroids self-organized into structures with an MSC-dominant core. The inclusion of MSCs into wild-type KRas PDAC spheroids modestly affected their compaction; however, MSCs significantly increased the compaction and growth of mutant KRas PDAC spheroids. Notably, exogenous collagen 1 potentiated PANC1 spheroid compaction while ITGA1 knockdown in PANC1 cells blocked MSC-induced PANC1 spheroid compaction. In agreement with a role for collagen-based integrin adhesion complexes in stromal cell-induced PDAC phenotypes, we also discovered that MSC-induced PANC1 spheroid growth was completely blocked by the ITGB1 immunoneutralizing antibody mAb13. Finally, multiplexed single-cell immunohistochemical analysis of a 25 patient PDAC tissue microarray revealed a relationship between decreased variance in Spearman r correlation for ITGA1 and PLK1 expression within the tumor cell compartment of PDAC in patients with advanced disease stage, and elevated expression of both ITGA1 and PLK1 in PDAC was found to be associated with decreased patient survival. Taken together, this work uncovers new therapeutic vulnerabilities in PDAC that are relevant to the progression of this stromal cell-rich malignancy and which may reveal strategies for improving patient outcomes.

Keywords:  Cell adhesion; Integrins; KRas; Mitosis; Pancreatic cancer; Spheroids

DOI:  https://doi.org/10.1016/j.bbrc.2024.149575
Biochim Biophys Acta Mol Cell Res. 2024 Feb 19. pii: S0167-4889(24)00040-5. [Epub ahead of print]1871(4): 119697

The enzymes of serine synthesis pathway in cancer metastasis.

Lei Li, Yuting Qin, Yuping Chen.

  Metastasis, the major cause of cancer mortality, requires cancer cells to reprogram their metabolism to adapt to and thrive in different environments, thereby leaving metastatic cells metabolic characteristics different from their parental cells. Mounting research has revealed that the de novo serine synthesis pathway (SSP), a glycolytic branching pathway that consumes glucose carbons for serine makeup and α-ketoglutarate generation and thus supports the proliferation, survival, and motility of cancer cells, is one such reprogrammed metabolic pathway. During different metastatic cascades, the SSP enzyme proteins or their enzymatic activity are both dynamically altered; manipulating their expression or catalytic activity could effectively prevent the progression of cancer metastasis; and the SSP enzymatic proteins could even conduce to metastasis via their nonenzymatic functions. In this article we overview the SSP dynamics during cancer metastasis and put the focuses on the regulatory role of the SSP in metastasis and the underlying mechanisms that mainly involve cellular anabolism/catabolism, redox balance, and epigenetics, aiming to provide a theoretical basis for the development of therapeutic strategies for targeting metastatic lesions.

Keywords:  Metabolic reprogramming; Metastasis; PHGDH; Serine biosynthesis; Serine metabolism

DOI:  https://doi.org/10.1016/j.bbamcr.2024.119697
J Photochem Photobiol B. 2024 Feb 15. pii: S1011-1344(24)00027-7. [Epub ahead of print]252 112867

Genetically encoded fluorescent sensor to monitor intracellular arginine methylation.

Fangrong Zhang, Helmut Bischof, Sandra Burgstaller, Benjamin M R Bourgeois, Roland Malli, Tobias Madl.

  Arginine methylation (ArgMet), as a post-translational modification, plays crucial roles in RNA processing, transcriptional regulation, signal transduction, DNA repair, apoptosis and liquid-liquid phase separation (LLPS). Since arginine methylation is associated with cancer pathogenesis and progression, protein arginine methyltransferases have gained interest as targets for anti-cancer therapy. Despite considerable process made to elucidate (patho)physiological mechanisms regulated by arginine methylation, there remains a lack of tools to visualize arginine methylation with high spatiotemporal resolution in live cells. To address this unmet need, we generated an ArgMet-sensitive genetically encoded, Förster resonance energy transfer-(FRET) based biosensor, called GEMS, capable of quantitative real-time monitoring of ArgMet dynamics. We optimized these biosensors by using different ArgMet-binding domains, arginine-glycine-rich regions and adjusting the linkers within the biosensors to improve their performance. Using a set of mammalian cell lines and modulators, we demonstrated the applicability of GEMS for monitoring changes in arginine methylation with single-cell and temporal resolution. The GEMS can facilitate the in vitro screening to find potential protein arginine methyltransferase inhibitors and will contribute to a better understanding of the regulation of ArgMet related to differentiation, development and disease.

Keywords:  Arginine methylation; Förster resonance energy transfer; Live-cell imaging; PRMT inhibitors; Wnt signaling

DOI:  https://doi.org/10.1016/j.jphotobiol.2024.112867
Nat Cell Biol. 2024 Feb 22.

Drosophila TMEM63 and mouse TMEM63A are lysosomal mechanosensory ion channels.

Kai Li, Yanmeng Guo, Yayu Wang, Ruijun Zhu, Wei Chen, Tong Cheng, Xiaofan Zhang, Yinjun Jia, Ting Liu, Wei Zhang, Lily Yeh Jan, Yuh Nung Jan.

Cells sense physical forces and convert them into electrical or chemical signals, a process known as mechanotransduction. Whereas extensive studies focus on mechanotransduction at the plasma membrane, little is known about whether and how intracellular organelles sense mechanical force and the physiological functions of organellar mechanosensing. Here we identify the Drosophila TMEM63 (DmTMEM63) ion channel as an intrinsic mechanosensor of the lysosome, a major degradative organelle. Endogenous DmTMEM63 proteins localize to lysosomes, mediate lysosomal mechanosensitivity and modulate lysosomal morphology and function. Tmem63 mutant flies exhibit impaired lysosomal degradation, synaptic loss, progressive motor deficits and early death, with some of these mutant phenotypes recapitulating symptoms of TMEM63-associated human diseases. Importantly, mouse TMEM63A mediates lysosomal mechanosensitivity in Neuro-2a cells, indicative of functional conservation in mammals. Our findings reveal DmTMEM63 channel function in lysosomes and its physiological roles in vivo and provide a molecular basis to explore the mechanosensitive process in subcellular organelles.

DOI: https://doi.org/10.1038/s41556-024-01353-7
Cell Rep. 2024 Feb 21. pii: S2211-1247(24)00100-1. [Epub ahead of print]43(3): 113772

Identification of MIMAS, a multifunctional mega-assembly integrating metabolic and respiratory biogenesis factors of mitochondria.

Patrick Horten, Kuo Song, Joshua Garlich, Robert Hardt, Lilia Colina-Tenorio, Susanne E Horvath, Uwe Schulte, Bernd Fakler, Martin van der Laan, Thomas Becker, Rosemary A Stuart, Nikolaus Pfanner, Heike Rampelt.

  The mitochondrial inner membrane plays central roles in bioenergetics and metabolism and contains several established membrane protein complexes. Here, we report the identification of a mega-complex of the inner membrane, termed mitochondrial multifunctional assembly (MIMAS). Its large size of 3 MDa explains why MIMAS has escaped detection in the analysis of mitochondria so far. MIMAS combines proteins of diverse functions from respiratory chain assembly to metabolite transport, dehydrogenases, and lipid biosynthesis but not the large established supercomplexes of the respiratory chain, ATP synthase, or prohibitin scaffold. MIMAS integrity depends on the non-bilayer phospholipid phosphatidylethanolamine, in contrast to respiratory supercomplexes whose stability depends on cardiolipin. Our findings suggest that MIMAS forms a protein-lipid mega-assembly in the mitochondrial inner membrane that integrates respiratory biogenesis and metabolic processes in a multifunctional platform.

Keywords:  CP: Metabolism; CP: Molecular biology; membrane protein complex; metabolism; metabolite carriers; mitochondria; phosphatidylethanolamine; phospholipids; protein assembly; respiratory chain

DOI:  https://doi.org/10.1016/j.celrep.2024.113772
Cell Metab. 2024 Feb 12. pii: S1550-4131(24)00015-9. [Epub ahead of print]

IgG is an aging factor that drives adipose tissue fibrosis and metabolic decline.

Lexiang Yu, Qianfen Wan, Qiongming Liu, Yong Fan, Qiuzhong Zhou, Alicja A Skowronski, Summer Wang, Zhengping Shao, Chen-Yu Liao, Lei Ding, Brian K Kennedy, Shan Zha, Jianwen Que, Charles A LeDuc, Lei Sun, Liheng Wang, Li Qiang.

  Aging is underpinned by pronounced metabolic decline; however, the drivers remain obscure. Here, we report that IgG accumulates during aging, particularly in white adipose tissue (WAT), to impair adipose tissue function and metabolic health. Caloric restriction (CR) decreases IgG accumulation in WAT, whereas replenishing IgG counteracts CR's metabolic benefits. IgG activates macrophages via Ras signaling and consequently induces fibrosis in WAT through the TGF-β/SMAD pathway. Consistently, B cell null mice are protected from aging-associated WAT fibrosis, inflammation, and insulin resistance, unless exposed to IgG. Conditional ablation of the IgG recycling receptor, neonatal Fc receptor (FcRn), in macrophages prevents IgG accumulation in aging, resulting in prolonged healthspan and lifespan. Further, targeting FcRn by antisense oligonucleotide restores WAT integrity and metabolic health in aged mice. These findings pinpoint IgG as a hidden culprit in aging and enlighten a novel strategy to rejuvenate metabolic health.

Keywords:  IgG; adipose tissue; aging; fibrosis; metabolic dysfunction

DOI:  https://doi.org/10.1016/j.cmet.2024.01.015
Cell Rep Med. 2024 Feb 20. pii: S2666-3791(24)00051-X. [Epub ahead of print]5(2): 101428

Fateful fat: Intra-pancreatic lipids cause pancreatic cancer.

Maxim S Petrov.

In a Mendelian randomization and prospective cohort study,1 intra-pancreatic fat increases the risk of pancreatic cancer. This provides persuasive human evidence of causal relation between lipids and cancer in the pancreas, which confirms a prediction of the PANDORA hypothesis.

DOI: https://doi.org/10.1016/j.xcrm.2024.101428
Proc Natl Acad Sci U S A. 2024 Feb 27. 121(9): e2318046121

Mechanism and cellular function of direct membrane binding by the ESCRT and ERES-associated Ca2+-sensor ALG-2.

Sankalp Shukla, Wei Chen, Shanlin Rao, Serim Yang, Chenxi Ou, Kevin P Larsen, Gerhard Hummer, Phyllis I Hanson, James H Hurley.

  Apoptosis linked Gene-2 (ALG-2) is a multifunctional intracellular Ca2+ sensor and the archetypal member of the penta-EF hand protein family. ALG-2 functions in the repair of damage to both the plasma and lysosome membranes and in COPII-dependent budding at endoplasmic reticulum exit sites (ERES). In the presence of Ca2+, ALG-2 binds to ESCRT-I and ALIX in membrane repair and to SEC31A at ERES. ALG-2 also binds directly to acidic membranes in the presence of Ca2+ by a combination of electrostatic and hydrophobic interactions. By combining giant unilamellar vesicle-based experiments and molecular dynamics simulations, we show that charge-reversed mutants of ALG-2 at these locations disrupt membrane recruitment. ALG-2 membrane binding mutants have reduced or abrogated ERES localization in response to Thapsigargin-induced Ca2+ release but still localize to lysosomes following lysosomal Ca2+ release. In vitro reconstitution shows that the ALG-2 membrane-binding defect can be rescued by binding to ESCRT-I. These data thus reveal the nature of direct Ca2+-dependent membrane binding and its interplay with Ca2+-dependent protein binding in the cellular functions of ALG-2.

Keywords:  ESCRT; calcium-binding protein; lysosome; membrane repair; reconstitution

DOI:  https://doi.org/10.1073/pnas.2318046121
Nat Commun. 2024 Feb 21. 15(1): 1574

Annexins-a family of proteins with distinctive tastes for cell signaling and membrane dynamics.

Volker Gerke, Felicity N E Gavins, Michael Geisow, Thomas Grewal, Jyoti K Jaiswal, Jesper Nylandsted, Ursula Rescher.

Annexins are cytosolic proteins with conserved three-dimensional structures that bind acidic phospholipids in cellular membranes at elevated Ca2+ levels. Through this they act as Ca2+-regulated membrane binding modules that organize membrane lipids, facilitating cellular membrane transport but also displaying extracellular activities. Recent discoveries highlight annexins as sensors and regulators of cellular and organismal stress, controlling inflammatory reactions in mammals, environmental stress in plants, and cellular responses to plasma membrane rupture. Here, we describe the role of annexins as Ca2+-regulated membrane binding modules that sense and respond to cellular stress and share our view on future research directions in the field.

DOI: https://doi.org/10.1038/s41467-024-45954-0
J Exp Med. 2024 Apr 01. pii: e20232136. [Epub ahead of print]221(4):

Cellular senescence: Neither irreversible nor reversible.

Maurice Reimann, Soyoung Lee, Clemens A Schmitt.

Cellular senescence is a critical stress response program implicated in embryonic development, wound healing, aging, and immunity, and it backs up apoptosis as an ultimate cell-cycle exit mechanism. In analogy to replicative exhaustion of telomere-eroded cells, premature types of senescence-referring to oncogene-, therapy-, or virus-induced senescence-are widely considered irreversible growth arrest states as well. We discuss here that entry into full-featured senescence is not necessarily a permanent endpoint, but dependent on essential maintenance components, potentially transient. Unlike a binary state switch, we view senescence with its extensive epigenomic reorganization, profound cytomorphological remodeling, and distinctive metabolic rewiring rather as a journey toward a full-featured arrest condition of variable strength and depth. Senescence-underlying maintenance-essential molecular mechanisms may allow cell-cycle reentry if not continuously provided. Importantly, senescent cells that resumed proliferation fundamentally differ from those that never entered senescence, and hence would not reflect a reversion but a dynamic progression to a post-senescent state that comes with distinct functional and clinically relevant ramifications.

DOI: https://doi.org/10.1084/jem.20232136
Nat Cell Biol. 2024 Feb 22.

Lysosomal channels sensing forces.

Erika Riederer, Dejian Ren.

DOI: https://doi.org/10.1038/s41556-024-01347-5
Nat Aging. 2024 Feb 22.

Plasma membrane damage limits replicative lifespan in yeast and induces premature senescence in human fibroblasts.

Kojiro Suda, Yohsuke Moriyama, Nurhanani Razali, Yatzu Chiu, Yumiko Masukagami, Koutarou Nishimura, Hunter Barbee, Hiroshi Takase, Shinju Sugiyama, Yuta Yamazaki, Yoshikatsu Sato, Tetsuya Higashiyama, Yoshikazu Johmura, Makoto Nakanishi, Keiko Kono.

Plasma membrane damage (PMD) occurs in all cell types due to environmental perturbation and cell-autonomous activities. However, cellular outcomes of PMD remain largely unknown except for recovery or death. In this study, using budding yeast and normal human fibroblasts, we found that cellular senescence-stable cell cycle arrest contributing to organismal aging-is the long-term outcome of PMD. Our genetic screening using budding yeast unexpectedly identified a close genetic association between PMD response and replicative lifespan regulations. Furthermore, PMD limits replicative lifespan in budding yeast; upregulation of membrane repair factors ESCRT-III (SNF7) and AAA-ATPase (VPS4) extends it. In normal human fibroblasts, PMD induces premature senescence via the Ca2+-p53 axis but not the major senescence pathway, DNA damage response pathway. Transient upregulation of ESCRT-III (CHMP4B) suppressed PMD-dependent senescence. Together with mRNA sequencing results, our study highlights an underappreciated but ubiquitous senescent cell subtype: PMD-dependent senescent cells.

DOI: https://doi.org/10.1038/s43587-024-00575-6
J Cell Biol. 2024 Mar 04. pii: e202401056. [Epub ahead of print]223(3):

PC4: A new regulator of cyclin D1 transcript levels.

Anne Fassl, Piotr Sicinski.

The expression of cyclin proteins is tightly regulated during the cell cycle, to allow precise activation of cyclin-dependent kinases. In this issue, Pan et al. (https://doi.org/10.1083/jcb.202308066) identify an RNA-binding protein, PC4, as a regulator of cyclin D1 mRNA stability in hepatocellular carcinoma cells. This study provides a new mechanism regulating the levels of a key cell cycle protein, cyclin D1, in human cells.

DOI: https://doi.org/10.1083/jcb.202401056
Nat Commun. 2024 Feb 20. 15(1): 1532

Plasticity-induced repression of Irf6 underlies acquired resistance to cancer immunotherapy in pancreatic ductal adenocarcinoma.

Il-Kyu Kim, Mark S Diamond, Salina Yuan, Samantha B Kemp, Benjamin M Kahn, Qinglan Li, Jeffrey H Lin, Jinyang Li, Robert J Norgard, Stacy K Thomas, Maria Merolle, Takeshi Katsuda, John W Tobias, Timour Baslan, Katerina Politi, Robert H Vonderheide, Ben Z Stanger.

Acquired resistance to immunotherapy remains a critical yet incompletely understood biological mechanism. Here, using a mouse model of pancreatic ductal adenocarcinoma (PDAC) to study tumor relapse following immunotherapy-induced responses, we find that resistance is reproducibly associated with an epithelial-to-mesenchymal transition (EMT), with EMT-transcription factors ZEB1 and SNAIL functioning as master genetic and epigenetic regulators of this effect. Acquired resistance in this model is not due to immunosuppression in the tumor immune microenvironment, disruptions in the antigen presentation machinery, or altered expression of immune checkpoints. Rather, resistance is due to a tumor cell-intrinsic defect in T-cell killing. Molecularly, EMT leads to the epigenetic and transcriptional silencing of interferon regulatory factor 6 (Irf6), rendering tumor cells less sensitive to the pro-apoptotic effects of TNF-α. These findings indicate that acquired resistance to immunotherapy may be mediated by programs distinct from those governing primary resistance, including plasticity programs that render tumor cells impervious to T-cell killing.

DOI: https://doi.org/10.1038/s41467-024-46048-7
Nat Genet. 2024 Feb 22.

Chromosome evolution screens recapitulate tissue-specific tumor aneuploidy patterns.

Emma V Watson, Jake June-Koo Lee, Doga C Gulhan, Giorgio E M Melloni, Sergey V Venev, Rayna Y Magesh, Abdulrazak Frederick, Kunitoshi Chiba, Eric C Wooten, Kamila Naxerova, Job Dekker, Peter J Park, Stephen J Elledge.

Whole chromosome and arm-level copy number alterations occur at high frequencies in tumors, but their selective advantages, if any, are poorly understood. Here, utilizing unbiased whole chromosome genetic screens combined with in vitro evolution to generate arm- and subarm-level events, we iteratively selected the fittest karyotypes from aneuploidized human renal and mammary epithelial cells. Proliferation-based karyotype selection in these epithelial lines modeled tissue-specific tumor aneuploidy patterns in patient cohorts in the absence of driver mutations. Hi-C-based translocation mapping revealed that arm-level events usually emerged in multiples of two via centromeric translocations and occurred more frequently in tetraploids than diploids, contributing to the increased diversity in evolving tetraploid populations. Isogenic clonal lineages enabled elucidation of pro-tumorigenic mechanisms associated with common copy number alterations, revealing Notch signaling potentiation as a driver of 1q gain in breast cancer. We propose that intrinsic, tissue-specific proliferative effects underlie tumor copy number patterns in cancer.

DOI: https://doi.org/10.1038/s41588-024-01665-2
Science. 2024 Feb 23. 383(6685): 890-897

Recording physiological history of cells with chemical labeling.

Magnus-Carsten Huppertz, Jonas Wilhelm, Vincent Grenier, Martin W Schneider, Tjalda Falt, Nicola Porzberg, David Hausmann, Dirk C Hoffmann, Ling Hai, Miroslaw Tarnawski, Gabriela Pino, Krasimir Slanchev, Ilya Kolb, Claudio Acuna, Lisa M Fenk, Herwig Baier, Julien Hiblot, Kai Johnsson.

Recordings of the physiological history of cells provide insights into biological processes, yet obtaining such recordings is a challenge. To address this, we introduce a method to record transient cellular events for later analysis. We designed proteins that become labeled in the presence of both a specific cellular activity and a fluorescent substrate. The recording period is set by the presence of the substrate, whereas the cellular activity controls the degree of the labeling. The use of distinguishable substrates enabled the recording of successive periods of activity. We recorded protein-protein interactions, G protein-coupled receptor activation, and increases in intracellular calcium. Recordings of elevated calcium levels allowed selections of cells from heterogeneous populations for transcriptomic analysis and tracking of neuronal activities in flies and zebrafish.

DOI: https://doi.org/10.1126/science.adg0812
Biochem J. 2024 Feb 21. pii: BCJ20240015. [Epub ahead of print]

Reporter cell lines to screen for inhibitors or regulators of the KRAS-RAF-MEK1/2-ERK1/2 pathway.

Laura Weatherdon, Kate Stuart, Megan A Cassidy, Alberto Moreno de la Gándara, Hanneke Okkenhaug, Markus Muellener, Grahame Mckenzie, Simon J Cook, Rebecca Gilley.

  The RAS-regulated RAF-MEK1/2-ERK1/2 signalling pathway is activated in cancer due to mutations in RAS proteins (especially KRAS), BRAF, CRAF, MEK1 and MEK2. Whilst inhibitors of KRASG12C (lung adenocarcinoma) and BRAF and MEK1/2 (melanoma and colorectal cancer) are clinically approved, acquired resistance remains a problem. Consequently, the search for new inhibitors (especially of RAS proteins), new inhibitor modalities and regulators of this pathway, which may be new drug targets, continues and increasingly involves cell-based screens with small molecules or genetic screens such as RNAi, CRISPR or Protein Interference. Here we describe cell lines that exhibit doxycycline-dependent expression KRASG12V or BRAFV600E and harbour a stably integrated EGR1:EmGFP reporter gene that can be detected by flow cytometry, high-content microscopy or immunoblotting. KRASG12V or BRAFV600E-driven EmGFP expression is inhibited by MEK1/2 or ERK1/2 inhibitors (MEKi and ERKi). BRAFi inhibit BRAFV600E-driven EmGFP expression but enhance the response to KRASG12V, recapitulating paradoxical activation of wild type RAF proteins. In addition to small molecules, expression of iDab6, encoding a RAS-specific antibody fragment inhibited KRASG12V- but not BRAFV600E-driven EmGFP expression. Finally, substitution of EmGFP for a bacterial nitroreductase gene allowed KRASG12V or BRAFV600E to drive cell death in the presence of a pro-drug, which may allow selection of pathway inhibitors that promote survival. These cell lines should prove useful for cell-based screens to identify new regulators of KRAS- or BRAF-dependent ERK1/2 signalling (drug target discovery) as well as screening or triaging 'hits' from drug discovery screens.

Keywords:  BRAF; ERK1/2; Gene reporters; KRAS; MEK1/2; small molecules

DOI:  https://doi.org/10.1042/BCJ20240015
Sci Adv. 2024 Feb 23. 10(8): eadj7944

Phase-shifting optothermal microscopy enables live-cell mid-infrared hyperspectral imaging of large cell populations at high confluency.

Tao Yuan, Lucas Riobo, Francesca Gasparin, Vasilis Ntziachristos, Miguel A Pleitez.

Rapid live-cell hyperspectral imaging at large fields of view (FOVs) and high cell confluency remains challenging for conventional vibrational spectroscopy-based microscopy technologies. At the same time, imaging at high cell confluency and large FOVs is important for proper cell function and statistical significance of measurements, respectively. Here, we introduce phase-shifting mid-infrared optothermal microscopy (PSOM), which interprets molecular-vibrational information as the optical path difference induced by mid-infrared absorption and can take snapshot vibrational images over broad excitation areas at high live-cell confluency. By means of phase-shifting, PSOM suppresses noise to a quarter of current optothermal microscopy modalities to allow capturing live-cell vibrational images at FOVs up to 50 times larger than state of the art. PSOM also reduces illumination power flux density (PFD) down to four orders of magnitude lower than other conventional vibrational microscopy methods, such as coherent anti-Stokes Raman scattering (CARS), thus considerably decreasing the risk of cell photodamage.

DOI: https://doi.org/10.1126/sciadv.adj7944
J Struct Biol. 2024 Feb 15. pii: S1047-8477(24)00007-8. [Epub ahead of print] 108067

ColabSeg: An interactive tool for editing, processing, and visualizing membrane segmentations from cryo-ET data.

Marc Siggel, Rasmus K Jensen, Valentin J Maurer, Julia Mahamid, Jan Kosinski.

  Cellular cryo-electron tomography (cryo-ET) has emerged as a key method to unravel the spatial and structural complexity of cells in their near-native state at unprecedented molecular resolution. To enable quantitative analysis of the complex shapes and morphologies of lipid membranes, the noisy three-dimensional (3D) volumes must be segmented. Despite recent advances, this task often requires considerable user intervention to curate the resulting segmentations. Here, we present ColabSeg, a Python-based tool for processing, visualizing, cleaning, and fitting membrane segmentations from cryo-ET data for downstream analysis. ColabSeg makes many well-established algorithms for point-cloud processing easily available to the broad community of structural biologists for applications in cryo-ET through its graphical user interface (GUI). We demonstrate the usefulness of the tool with a range of use cases and biological examples. Finally, for a large Mycoplasma pneumoniae dataset of 50 tomograms, we show how ColabSeg enables high-throughput membrane segmentation, which can be used as valuable training data for fully automated convolutional neural network (CNN)-based segmentation.

Keywords:  Cryo-electron tomography (CryoET); electron microscopy; image analysis; lipid membranes; segmentation

DOI:  https://doi.org/10.1016/j.jsb.2024.108067
bioRxiv. 2024 Feb 07. pii: 2024.02.06.579216. [Epub ahead of print]

mTORC1 activity oscillates throughout the cell cycle promoting mitotic entry and differentially influencing autophagy induction.

Jay N Joshi, Ariel D Lerner, Frank Scallo, Alexandra N Grumet, Paul Matteson, James H Millonig, Alexander J Valvezan.

Mechanistic Target of Rapamycin Complex 1 (mTORC1) is a master metabolic regulator that stimulates anabolic cell growth while suppressing catabolic processes such as autophagy. mTORC1 is active in most, if not all, proliferating eukaryotic cells. However, it remains unclear whether and how mTORC1 activity changes from one cell cycle phase to another. Here we tracked mTORC1 activity through the complete cell cycle and uncover oscillations in its activity. We find that mTORC1 activity peaks in S and G2, and is lowest in mitosis and G1. We further demonstrate that multiple mechanisms are involved in controlling this oscillation. The interphase oscillation is mediated through the TSC complex, an upstream negative regulator of mTORC1, but is independent of major known regulatory inputs to the TSC complex, including Akt, Mek/Erk, and CDK4/6 signaling. By contrast, suppression of mTORC1 activity in mitosis does not require the TSC complex, and instead involves CDK1-dependent control of the subcellular localization of mTORC1 itself. Functionally, we find that in addition to its well-established role in promoting progression through G1, mTORC1 also promotes progression through S and G2, and is important for satisfying the Wee1- and Chk1-dependent G2/M checkpoint to allow entry into mitosis. We also find that low mTORC1 activity in G1 sensitizes cells to autophagy induction in response to partial mTORC1 inhibition or reduced nutrient levels. Together these findings demonstrate that mTORC1 is differentially regulated throughout the cell cycle, with important phase-specific functional consequences in proliferating cells.

DOI: https://doi.org/10.1101/2024.02.06.579216
Mol Oncol. 2024 Feb 20.

ITIH5 as a multifaceted player in pancreatic cancer suppression, impairing tyrosine kinase signaling, cell adhesion and migration.

Jennifer Kosinski, Antonio Sechi, Johanna Hain, Sophia Villwock, Stefanie Anh Ha, Maximilian Hauschulz, Michael Rose, Florian Steib, Nadina Ortiz-Brüchle, Lara Heij, Sanne L Maas, Emiel P C van der Vorst, Thomas Knoesel, Annelore Altendorf-Hofmann, Ronald Simon, Guido Sauter, Jan Bednarsch, Danny Jonigk, Edgar Dahl.

  Inter-alpha-trypsin inhibitor heavy chain 5 (ITIH5) has been identified as a metastasis suppressor gene in pancreatic cancer. Here, we analyzed ITIH5 promoter methylation and protein expression in The Cancer Genome Atlas (TCGA) dataset and three tissue microarray cohorts (n = 618), respectively. Cellular effects, including cell migration, focal adhesion formation and protein tyrosine kinase activity, induced by forced ITIH5 expression in pancreatic cancer cell lines were studied in stable transfectants. ITIH5 promoter hypermethylation was associated with unfavorable prognosis, while immunohistochemistry demonstrated loss of ITIH5 in the metastatic setting and worsened overall survival. Gain-of-function models showed a significant reduction in migration capacity, but no alteration in proliferation. Focal adhesions in cells re-expressing ITIH5 exhibited a smaller and more rounded phenotype, typical for slow-moving cells. An impressive increase of acetylated alpha-tubulin was observed in ITIH5-positive cells, indicating more stable microtubules. In addition, we found significantly decreased activities of kinases related to focal adhesion. Our results indicate that loss of ITIH5 in pancreatic cancer profoundly affects its molecular profile: ITIH5 potentially interferes with a variety of oncogenic signaling pathways, including the PI3K/AKT pathway. This may lead to altered cell migration and focal adhesion formation. These cellular alterations may contribute to the metastasis-inhibiting properties of ITIH5 in pancreatic cancer.

Keywords:  C2TSG; ITIH5; focal adhesions; metastasis suppressor; pancreatic cancer; tumor cell migration

DOI:  https://doi.org/10.1002/1878-0261.13609
Nat Chem. 2024 Feb 21.

Asymmetric oligomerization state and sequence patterning can tune multiphase condensate miscibility.

Ushnish Rana, Ke Xu, Amal Narayanan, Mackenzie T Walls, Athanassios Z Panagiotopoulos, José L Avalos, Clifford P Brangwynne.

Endogenous biomolecular condensates, composed of a multitude of proteins and RNAs, can organize into multiphasic structures with compositionally distinct phases. This multiphasic organization is generally understood to be critical for facilitating their proper biological function. However, the biophysical principles driving multiphase formation are not completely understood. Here we use in vivo condensate reconstitution experiments and coarse-grained molecular simulations to investigate how oligomerization and sequence interactions modulate multiphase organization in biomolecular condensates. We demonstrate that increasing the oligomerization state of an intrinsically disordered protein results in enhanced immiscibility and multiphase formation. Interestingly, we find that oligomerization tunes the miscibility of intrinsically disordered proteins in an asymmetric manner, with the effect being more pronounced when the intrinsically disordered protein, exhibiting stronger homotypic interactions, is oligomerized. Our findings suggest that oligomerization is a flexible biophysical mechanism that cells can exploit to tune the internal organization of biomolecular condensates and their associated biological functions.

DOI: https://doi.org/10.1038/s41557-024-01456-6
Eur J Nucl Med Mol Imaging. 2024 Feb 19.

Magnetic-optical dual-modality imaging monitoring chemotherapy efficacy of pancreatic ductal adenocarcinoma with a low-dose fibronectin-targeting Gd-based contrast agent.

Wenjia Zhang, Xiaolong Liang, Xinyu Zhang, Wei Tong, Guangyuan Shi, Haozhuo Guo, Zhengyu Jin, Jie Tian, Yang Du, Huadan Xue.

   PURPOSE: Pancreatic ductal adenocarcinoma (PDAC) is a lethal hypovascular tumor surrounded by dense fibrosis. Albumin-bound paclitaxel and gemcitabine (AG) chemotherapy is the mainstay of PDAC treatment through depleting peritumoral fibrosis and killing tumor cells; however, it remains challenging due to the lack of a noninvasive imaging method evaluating fibrotic changes during AG chemotherapy. In this study, we developed a dual-modality imaging platform that enables noninvasive, dynamic, and quantitative assessment of chemotherapy-induced fibrotic changes through near-infrared fluorescence molecular imaging (FMI) and magnetic resonance imaging (MRI) using an extradomain B fibronectin (EDB-FN)-targeted imaging probe (ZD2-Gd-DOTA-Cy7).
METHODS: The ZD2-Gd-DOTA-Cy7 probe was constructed by conjugating a peptide (Cys-TVRTSAD) to Gd-DOTA and the near-infrared dye Cy7. PDAC murine xenograft models were intravenously injected with ZD2-Gd-DOTA-Cy7 at a Gd concentration of 0.05 mmol/kg or free Cy7 and Gd-DOTA as control. The normalized tumor background ratio (TBR) on FMI and the T1 reduction ratio on MRI were quantitatively analyzed. For models receiving AG chemotherapy or saline, MRI/FMI was performed before and after treatment. Histological analyses were performed for validation.
RESULTS: The ZD2-Gd-DOTA-Cy7 concentration showed a linear correlation with the fluorescence intensity and T1 relaxation time in vitro. The optimal imaging time was 30 min after injection of the ZD2-Gd-DOTA-Cy7 (0.05 mmol/kg), only half of the clinic dosage of gadolinium. Additionally, ZD2-Gd-DOTA-Cy7 generated a 1.44-fold and 1.90-fold robust contrast enhancement compared with Cy7 (P < 0.05) and Gd-DOTA (P < 0.05), respectively. For AG chemotherapy monitoring, the T1 reduction ratio and normalized TBR in the fibrotic tumor areas were significantly increased by 1.99-fold (P < 0.05) and 1.78-fold (P < 0.05), respectively, in the control group compared with those in the AG group.
CONCLUSION: MRI/FMI with a low dose of ZD2-Gd-DOTA-Cy7 enables sensitive imaging of PDAC and the quantitative assessment of fibrotic changes during AG chemotherapy, which shows potential clinical applications for precise diagnosis, post-treatment monitoring, and disease management.

Keywords:  Fibronectin; Fluorescence molecular imaging; Magnetic resonance imaging; Molecular imaging; Pancreatic cancer

DOI:  https://doi.org/10.1007/s00259-024-06617-w
EMBO J. 2024 Feb 23.

NINJ1 induces plasma membrane rupture and release of damage-associated molecular pattern molecules during ferroptosis.

Saray Ramos, Ella Hartenian, José Carlos Santos, Philipp Walch, Petr Broz.

  Ferroptosis is a regulated form of necrotic cell death caused by iron-dependent accumulation of oxidized phospholipids in cellular membranes, culminating in plasma membrane rupture (PMR) and cell lysis. PMR is also a hallmark of other types of programmed necrosis, such as pyroptosis and necroptosis, where it is initiated by dedicated pore-forming cell death-executing factors. However, whether ferroptosis-associated PMR is also actively executed by proteins or driven by osmotic pressure remains unknown. Here, we investigate a potential ferroptosis role of ninjurin-1 (NINJ1), a recently identified executor of pyroptosis-associated PMR. We report that NINJ1 oligomerizes during ferroptosis, and that Ninj1-deficiency protects macrophages and fibroblasts from ferroptosis-associated PMR. Mechanistically, we find that NINJ1 is dispensable for the initial steps of ferroptosis, such as lipid peroxidation, channel-mediated calcium influx, and cell swelling. In contrast, NINJ1 is required for early loss of plasma membrane integrity, which precedes complete PMR. Furthermore, NINJ1 mediates the release of cytosolic proteins and danger-associated molecular pattern (DAMP) molecules from ferroptotic cells, suggesting that targeting NINJ1 could be a therapeutic option to reduce ferroptosis-associated inflammation.

Keywords:  Cell Death; Ferroptosis; Inflammation; Ninjurin-1; Plasma Membrane Rupture

DOI:  https://doi.org/10.1038/s44318-024-00055-y
bioRxiv. 2024 Feb 08. pii: 2024.02.06.579096. [Epub ahead of print]

Longitudinal Fragility Phenotyping Predicts Lifespan and Age-Associated Morbidity in C57BL/6 and Diversity Outbred Mice.

Alison Luciano, Laura Robinson, Gaven Garland, Bonnie Lyons, Ron Korstanje, Andrea Di Francesco, Gary A Churchill.

Aging studies in mammalian models often depend on natural lifespan data as a primary outcome. Tools for lifespan prediction could accelerate these studies and reduce the need for veterinary intervention. Here, we leveraged large-scale longitudinal frailty and lifespan data on two genetically distinct mouse cohorts to evaluate noninvasive strategies to predict life expectancy in mice. We applied a modified frailty assessment, the Fragility Index, derived from existing frailty indices with additional deficits selected by veterinarians. We developed an ensemble machine learning classifier to predict imminent mortality (95% proportion of life lived [95PLL]). Our algorithm represented improvement over previous predictive criteria but fell short of the level of reliability that would be needed to make advanced prediction of lifespan and thus accelerate lifespan studies. Highly sensitive and specific frailty-based predictive endpoint criteria for aged mice remain elusive. While frailty-based prediction falls short as a surrogate for lifespan, it did demonstrate significant predictive power and as such must contain information that could be used to inform the conclusion of aging experiments. We propose a frailty-based measure of healthspan as an alternative target for aging research and demonstrate that lifespan and healthspan criteria reveal distinct aspects of aging in mice.

DOI: https://doi.org/10.1101/2024.02.06.579096
Trends Cancer. 2024 Feb 19. pii: S2405-8033(24)00009-8. [Epub ahead of print]

Fusion genes in pancreatic tumors.

Anastasios Gkountakos, Aatur D Singhi, C Benedikt Westphalen, Aldo Scarpa, Claudio Luchini.

  Gene fusions and rearrangements play a crucial role in tumor biology. They are rare events typically detected in KRAS wild-type (WT) pancreatic tumors. Their identification can inform clinical management by enabling precision oncology, as fusions involving BRAF, FGFR2, RET, NTRK, NRG1, and ALK represent actionable targets in KRAS-WT cancers, and serve diagnostic purposes since fusions involving PRKACA/B represent the diagnostic hallmark of intraductal oncocytic papillary neoplasms (IOPNs). Although they are rare, the therapeutic and diagnostic importance of these genomic events should not be underestimated, highlighting the need for quality-ensured molecular diagnostics in the management of cancer. Herein we review the existing literature on the role of fusion genes in pancreatic tumors and their clinical potential as effective biomarkers and therapeutic targets.

Keywords:  fusion; fusion gene; fusions; pancreatic cancer; pancreatic neoplasms; precision oncology

DOI:  https://doi.org/10.1016/j.trecan.2024.01.009
Cancer Epidemiol Biomarkers Prev. 2024 Feb 22.

Sleep and risk of pancreatic cancer in the UK Biobank.

Joshua R Freeman, Pedro F Saint-Maurice, Ting Zhang, Charles E Matthews, Rachael Z Stolzenberg-Solomon.

BACKGROUND: Light at night, which may cause circadian disruption, is a potential pancreatic cancer risk factor. However, evidence from related exposures such as poor sleep health and shift work remains inconclusive and sparsely investigated.
METHODS: We evaluated associations between self-reported typical sleep duration, chronotype, shift work, insomnia symptoms, snoring, and daytime sleeping and pancreatic ductal adenocarcinomas (PDAC) incidence among 475,286 United Kingdom (UK) Biobank participants. We used Cox proportional hazards models to estimate hazard ratios (HR) and 95% confidence intervals (CI) adjusting for age, sex, body mass index, smoking status, duration, and frequency, alcohol intake, diabetes status, race, and employment/shift work.
RESULTS: Over 14 years of follow-up, 1,079 adults were diagnosed with PDAC. There were no associations observed between sleep characteristics, including sleep duration (<7 vs. 7-<9 hours; HR: 1.03, 95% CI 0.90-1.19; ≥9 hours; HR: 1.00 [0.81-1.24]), evening chronotype ("definitely" an evening person vs. "definitely" a morning person; HR: 0.99 [0.77-1.29]), shift work, insomnia symptoms, snoring, or daytime sleep and PDAC risk.
CONCLUSIONS: Self-reported typical sleep characteristics and shift work were not associated with PDAC risk.
IMPACT: Considering the role of light at night and shift work in circadian disruption and cancer risk, it is plausible that poor sleep health among a general population may be related to cancer risk through similar sleep and circadian disrupting processes. This work may suggest that typical sleep characteristics and shift work are not associated with PDAC, though additional work is needed to confirm these findings.

DOI: https://doi.org/10.1158/1055-9965.EPI-23-0983
J Exp Clin Cancer Res. 2024 Feb 21. 43(1): 53

PARP1-targeted fluorescence molecular endoscopy as novel tool for early detection of esophageal dysplasia and adenocarcinoma.

Sabrina Marcazzan, Marcos J Braz Carvalho, Nghia T Nguyen, Julia Strangmann, Julia Slotta-Huspenina, Anna Tenditnaya, Markus Tschurtschenthaler, Jonas Rieder, Andrea Proaño-Vasco, Vasilis Ntziachristos, Katja Steiger, Dimitris Gorpas, Michael Quante, Susanne Kossatz.

   BACKGROUND: Esophageal cancer is one of the 10 most common cancers worldwide and its incidence is dramatically increasing. Despite some improvements, the current surveillance protocol with white light endoscopy and random untargeted biopsies collection (Seattle protocol) fails to diagnose dysplastic and cancerous lesions in up to 50% of patients. Therefore, new endoscopic imaging technologies in combination with tumor-specific molecular probes are needed to improve early detection. Herein, we investigated the use of the fluorescent Poly (ADP-ribose) Polymerase 1 (PARP1)-inhibitor PARPi-FL for early detection of dysplastic lesions in patient-derived organoids and transgenic mouse models, which closely mimic the transformation from non-malignant Barrett's Esophagus (BE) to invasive esophageal adenocarcinoma (EAC).
METHODS: We determined PARP1 expression via immunohistochemistry (IHC) in human biospecimens and mouse tissues. We also assessed PARPi-FL uptake in patient- and mouse-derived organoids. Following intravenous injection of 75 nmol PARPi-FL/mouse in L2-IL1B (n = 4) and L2-IL1B/IL8Tg mice (n = 12), we conducted fluorescence molecular endoscopy (FME) and/or imaged whole excised stomachs to assess PARPi-FL accumulation in dysplastic lesions. L2-IL1B/IL8Tg mice (n = 3) and wild-type (WT) mice (n = 2) without PARPi-FL injection served as controls. The imaging results were validated by confocal microscopy and IHC of excised tissues.
RESULTS: IHC on patient and murine tissue revealed similar patterns of increasing PARP1 expression in presence of dysplasia and cancer. In human and murine organoids, PARPi-FL localized to PARP1-expressing epithelial cell nuclei after 10 min of incubation. Injection of PARPi-FL in transgenic mouse models of BE resulted in the successful detection of lesions via FME, with a mean target-to-background ratio > 2 independently from the disease stage. The localization of PARPi-FL in the lesions was confirmed by imaging of the excised stomachs and confocal microscopy. Without PARPi-FL injection, identification of lesions via FME in transgenic mice was not possible.
CONCLUSION: PARPi-FL imaging is a promising approach for clinically needed improved detection of dysplastic and malignant EAC lesions in patients with BE. Since PARPi-FL is currently evaluated in a phase 2 clinical trial for oral cancer detection after topical application, clinical translation for early detection of dysplasia and EAC in BE patients via FME screening appears feasible.

Keywords:  Animal Models; Dysplasia; Esophageal Adenocarcinoma; Fluorescence Imaging; Fluorescence Molecular Endoscopy; PARP1

DOI:  https://doi.org/10.1186/s13046-024-02963-7