bims-cagime Biomed News
on Cancer, aging and metabolism
Issue of 2024–08–25
25 papers selected by
Kıvanç Görgülü, Technical University of Munich



  1. Nat Metab. 2024 Aug 19.
      Metastases arise from subsets of cancer cells that disseminate from the primary tumour1,2. The ability of cancer cells to thrive in a new tissue site is influenced by genetic and epigenetic changes that are important for disease initiation and progression, but these factors alone do not predict if and where cancers metastasize3,4. Specific cancer types metastasize to consistent subsets of tissues, suggesting that primary tumour-associated factors influence where cancers can grow. We find primary and metastatic pancreatic tumours have metabolic similarities and that the tumour-initiating capacity and proliferation of both primary-derived and metastasis-derived cells is favoured in the primary site relative to the metastatic site. Moreover, propagating cells as tumours in the lung or the liver does not enhance their relative ability to form large tumours in those sites, change their preference to grow in the primary site, nor stably alter aspects of their metabolism relative to primary tumours. Primary liver and lung cancer cells also exhibit a preference to grow in their primary site relative to metastatic sites. These data suggest cancer tissue of origin influences both primary and metastatic tumour metabolism and may impact where cancer cells can metastasize.
    DOI:  https://doi.org/10.1038/s42255-024-01105-9
  2. J Am Chem Soc. 2024 Aug 20.
      The lateral organization of proteins and lipids in the plasma membrane is fundamental to regulating a wide range of cellular processes. Compartmentalized ordered membrane domains enriched with specific lipids, often termed lipid rafts, have been shown to modulate the physicochemical and mechanical properties of membranes and to drive protein sorting. Novel methods and tools enabling the visualization, characterization, and/or manipulation of membrane compartmentalization are crucial to link the properties of the membrane with cell functions. Flipper, a commercially available fluorescent membrane tension probe, has become a reference tool for quantitative membrane tension studies in living cells. Here, we report on a so far unidentified property of Flipper, namely, its ability to photosensitize singlet oxygen (1O2) under blue light when embedded into lipid membranes. This in turn results in the production of lipid hydroperoxides that increase membrane tension and trigger phase separation. In biological membranes, the photoinduced segregated domains retain the sorting ability of intact phase-separated membranes, directing raft and nonraft proteins into ordered and disordered regions, respectively, in contrast to radical-based photo-oxidation reactions that disrupt raft protein partitioning. The dual tension reporting and photosensitizing abilities of Flipper enable simultaneous visualization and manipulation of the mechanical properties and lateral organization of membranes, providing a powerful tool to optically control lipid raft formation and to explore the interplay between membrane biophysics and cell function.
    DOI:  https://doi.org/10.1021/jacs.4c08580
  3. Proc Natl Acad Sci U S A. 2024 Aug 27. 121(35): e2322755121
      The mechanistic target of rapamycin complex 1 (mTORC1) pathway regulates cell growth and metabolism in response to many environmental cues, including nutrients. Amino acids signal to mTORC1 by modulating the guanine nucleotide loading states of the heterodimeric Rag GTPases, which bind and recruit mTORC1 to the lysosomal surface, its site of activation. The Rag GTPases are tethered to the lysosome by the Ragulator complex and regulated by the GATOR1, GATOR2, and KICSTOR multiprotein complexes that localize to the lysosomal surface through an unknown mechanism(s). Here, we show that mTORC1 is completely insensitive to amino acids in cells lacking the Rag GTPases or the Ragulator component p18. Moreover, not only are the Rag GTPases and Ragulator required for amino acids to regulate mTORC1, they are also essential for the lysosomal recruitment of the GATOR1, GATOR2, and KICSTOR complexes, which stably associate and traffic to the lysosome as the "GATOR" supercomplex. The nucleotide state of RagA/B controls the lysosomal association of GATOR, in a fashion competitively antagonized by the N terminus of the amino acid transporter SLC38A9. Targeting of Ragulator to the surface of mitochondria is sufficient to relocalize the Rags and GATOR to this organelle, but not to enable the nutrient-regulated recruitment of mTORC1 to mitochondria. Thus, our results reveal that the Rag-Ragulator complex is the central organizer of the physical architecture of the mTORC1 nutrient-sensing pathway and underscore that mTORC1 activation requires signal transduction on the lysosomal surface.
    Keywords:  biochemistry; mTOR signaling; nutrient sensing
    DOI:  https://doi.org/10.1073/pnas.2322755121
  4. EMBO Rep. 2024 Aug 16.
      One of the key events in autophagy is the formation of a double-membrane phagophore, and many regulatory mechanisms underpinning this remain under investigation. WIPI2b is among the first proteins to be recruited to the phagophore and is essential for stimulating autophagy flux by recruiting the ATG12-ATG5-ATG16L1 complex, driving LC3 and GABARAP lipidation. Here, we set out to investigate how WIPI2b function is regulated by phosphorylation. We studied two phosphorylation sites on WIPI2b, S68 and S284. Phosphorylation at these sites plays distinct roles, regulating WIPI2b's association with ATG16L1 and the phagophore, respectively. We confirm WIPI2b is a novel ULK1 substrate, validated by the detection of endogenous phosphorylation at S284. Notably, S284 is situated within an 18-amino acid stretch, which, when in contact with liposomes, forms an amphipathic helix. Phosphorylation at S284 disrupts the formation of the amphipathic helix, hindering the association of WIPI2b with membranes and autophagosome formation. Understanding these intricacies in the regulatory mechanisms governing WIPI2b's association with its interacting partners and membranes, holds the potential to shed light on these complex processes, integral to phagophore biogenesis.
    Keywords:  Amphipathic Helix; Autophagosome; Autophagy; Kinase; WIPIs
    DOI:  https://doi.org/10.1038/s44319-024-00215-5
  5. J Physiol. 2024 Aug 21.
      Oxidative stress contributes to the loss of skeletal muscle mass and function in cancer cachexia. However, this outcome may be mitigated by an improved endogenous antioxidant defence system. Here, using the well-established oxidative stress-inducing muscle atrophy model of Lewis lung carcinoma (LLC) in 13-week-old male C57BL/6J mice, we demonstrate that extracellular superoxide dismutase (EcSOD) levels increase in the cachexia-prone extensor digitorum longus muscle. LLC transplantation significantly increased interleukin-1β (IL-1β) expression and release from extensor digitorum longus muscle fibres. Moreover, IL-1β treatment of C2C12 myotubes increased NBR1, p62 phosphorylation at Ser351, Nrf2 nuclear translocation and EcSOD protein expression. Additional studies in vivo indicated that intramuscular IL-1β injection is sufficient to stimulate EcSOD expression, which is prevented by muscle-specific knockout of p62 and Nrf2 (i.e. in p62 skmKO and Nrf2 skmKO mice, respectively). Finally, since an increase in circulating IL-1β may lead to unwanted outcomes, we demonstrate that targeting this pathway at p62 is sufficient to drive muscle EcSOD expression in an Nrf2-dependent manner. In summary, cancer cachexia increases EcSOD expression in extensor digitorum longus muscle via muscle-derived IL-1β-induced upregulation of p62 phosphorylation and Nrf2 activation. These findings provide further mechanistic evidence for the therapeutic potential of p62 and Nrf2 to mitigate cancer cachexia-induced muscle atrophy. KEY POINTS: Oxidative stress plays an important role in muscle atrophy during cancer cachexia. EcSOD, which mitigates muscle loss during oxidative stress, is upregulated in 13-week-old male C57BL/6J mice of extensor digitorum longus muscles during cancer cachexia. Using mouse and cellular models, we demonstrate that cancer cachexia promotes muscle EcSOD protein expression via muscle-derived IL-1β-dependent stimulation of the NBR1-p62-Nrf2 signalling pathway. These results provide further evidence for the potential therapeutic targeting of the NBR1-p62-Nrf2 signalling pathway downstream of IL-1β to mitigate cancer cachexia-induced muscle atrophy.
    Keywords:  NBR1; Nrf2; SQSTM1/p62; extracellular superoxide dismutase; interleukin‐1β; muscle atrophy; skeletal muscle
    DOI:  https://doi.org/10.1113/JP286460
  6. BMC Neurosci. 2024 Aug 22. 25(1): 37
       BACKGROUND: Adipose and muscle tissue wasting outlines the cachectic process during tumor progression. The sympathetic nervous system (SNS) is known to promote tumor progression and research suggests that it might also contribute to cancer-associated cachexia (CAC) energetic expenditure through fat wasting.
    METHODS: We sympathectomized L5178Y-R tumor-bearing male BALB/c mice by intraperitoneally administering 6-hydroxydopamine to evaluate morphometric, inflammatory, and molecular indicators of CAC and tumor progression.
    RESULTS: Tumor burden was associated with cachexia indicators, including a 10.5% body mass index (BMI) decrease, 40.19% interscapular, 54% inguinal, and 37.17% visceral adipose tissue loss, a 12% food intake decrease, and significant (p = 0.038 and p = 0.0037) increases in the plasmatic inflammatory cytokines IL-6 and IFN-γ respectively. Sympathectomy of tumor-bearing mice was associated with attenuated BMI and visceral adipose tissue loss, decreased interscapular Ucp-1 gene expression to basal levels, and 2.6-fold reduction in Mmp-9 relative gene expression, as compared with the unsympathectomized mice control group.
    CONCLUSION: The SNS contributes to CAC-associated morphometric and adipose tissue alterations and promotes tumor progression in a murine model.
    Keywords:  Adrenergic system; Cancer cachexia; Inflammation; Lymphoma; Sympathectomy; Sympathetic nervous system
    DOI:  https://doi.org/10.1186/s12868-024-00887-8
  7. J Biol Chem. 2024 Aug 20. pii: S0021-9258(24)02198-7. [Epub ahead of print] 107697
      To elucidate the dynamic evolution of cancer cell characteristics within the tumor microenvironment (TME), we developed an integrative approach combining single-cell tracking, cell fate simulation, and three-dimensional (3D) TME modeling. We began our investigation by analyzing the spatiotemporal behavior of individual cancer cells in cultured pancreatic (MiaPaCa2) and cervical (HeLa) cancer cell lines, with a focus on the α2-6 sialic acid (α2-6Sia) modification on glycans, which is associated with cell stemness. Our findings revealed that MiaPaCa2 cells exhibited significantly higher levels of α2-6Sia modification, correlating with enhanced reproductive capabilities, whereas HeLa cells showed less prevalence of this modification. To accommodate the in vivo variability of α2-6Sia levels, we employed a cell fate simulation algorithm that digitally generates cell populations based on our observed data while varying the level of sialylation, thereby simulating cell growth patterns. Subsequently, we performed a 3D TME simulation with these deduced cell populations, considering the microenvironment that could impact cancer cell growth. Immune cell landscape information derived from 193 cervical and 172 pancreatic cancer cases was used to estimate the degree of the positive or negative impact. Our analysis suggests that the deduced cells generated based on the characteristics of MiaPaCa2 cells are less influenced by the immune cell landscape within the TME compared to those of HeLa cells, highlighting that the fate of cancer cells is shaped by both the surrounding immune landscape and the intrinsic characteristics of the cancer cells.
    Keywords:  3-dimensional tumor microenvironment simulation; Sambucus nigra lectin; Single-cell tracking; cancer cell heterogeneity; cancer cell lines; cell fate simulation; cervical cancer; live cell imaging; pancreatic cancer; stemness; tumor microenvironment; α2-6 sialic acid modification on glycans
    DOI:  https://doi.org/10.1016/j.jbc.2024.107697
  8. Nature. 2024 Aug;632(8027): 987-988
      
    Keywords:  Biochemistry; Cell biology
    DOI:  https://doi.org/10.1038/d41586-024-02528-w
  9. Mol Cancer Ther. 2024 Aug 20.
      Despite advances in immune checkpoint inhibitors (ICIs), chemotherapy remains the standard therapy for patients with pancreatic ductal adenocarcinoma (PDAC). As the combinations of chemotherapy, including the FOLFIRINOX (5-fluorouracil (5FU), irinotecan, and oxaliplatin) regimen, and ICIs have failed to demonstrate clinical benefit in patients with metastatic PDAC tumors, there is increasing interest in identifying therapeutic approaches to potentiate ICI efficacy in PDAC patients. In this study, we report that neoadjuvant FOLFRINOX-treated human PDAC tumors exhibit increased MEK/ERK activation. We also show elevated MEK/ERK signaling in ex vivo PDAC slice cultures and cell lines treated with a combination of 5FU (F), irinotecan (I), and oxaliplatin (O) (FIO). In addition, we find that the KPC-FIO cells, established from repeated treatment of mouse PDAC cell lines with 6-8 cycles of FIO, display enhanced ERK phosphorylation and demonstrate increased sensitivity to MEK inhibition in vitro and in vivo. Significantly, the KPC-FIO cells develop tumors with a pro-inflammatory immune profile similar to human PDAC tumors following neoadjuvant FOLFIRINOX treatment. Furthermore, we found that the MEK inhibitor Trametinib enables additional infiltration of highly functional CD8+ T cells into the KPC-FIO tumors and potentiates the efficacy of anti-PD-1 antibody in syngeneic mouse models. Our findings provide a rationale for combining Trametinib and anti-PD-1 antibodies in PDAC patients following neoadjuvant or short-term FOLFIRINOX treatment to achieve effective anti-tumor responses.
    DOI:  https://doi.org/10.1158/1535-7163.MCT-23-0833
  10. STAR Protoc. 2024 Aug 22. pii: S2666-1667(24)00346-0. [Epub ahead of print]5(3): 103181
      Here, we present a protocol to comprehensively quantify autophagy initiation using the readout of the microtubule associated protein 1 light chain 3 beta (LC3B) Förster's resonance energy transfer (FRET) biosensor. We describe steps for cell seeding, transfection, FRET/FLIM (fluorescence lifetime imaging microscopy) imaging, and image analysis. This protocol can be useful in any physiology- or disease-related paradigm where the LC3B biosensor can be expressed to determine whether autophagy has been initiated or is stalled. The analysis pipeline presented here can be applied to any other genetically encoded FRET sensor imaged using FRET/FLIM. For complete details on the use and execution of this protocol, please refer to Gökerküçük et al.1.
    Keywords:  Cell-based Assays; Microscopy; Molecular/Chemical Probes; Single Cell
    DOI:  https://doi.org/10.1016/j.xpro.2024.103181
  11. Am J Gastroenterol. 2024 Aug 20.
       INTRODUCTION: Recent associative studies have linked intra-pancreatic fat deposition (IPFD) with risk of pancreatitis, but the causal relationship remains unclear.
    METHODS: Utilizing Mendelian randomization, we evaluated the causal association between genetically predicted IPFD and pancreatitis. This approach utilized genetic variants from genome-wide association studies of IPFD (n=25,617), acute pancreatitis (n=6,787 cases/361,641 controls), and chronic pancreatitis (n=3,875 cases/361,641 controls).
    RESULTS: Genetically predicted IPFD was significantly associated with acute pancreatitis (OR per 1-SD increase: 1.40[95%CI:1.12-1.76], p=0.0032) and chronic pancreatitis (OR:1.64[95%CI:1.13-2.39], p=0.0097).
    DISCUSSION: Our findings support a causal role of IPFD in pancreatitis, suggesting that reducing IPFD could lower the risk of pancreatitis.
    DOI:  https://doi.org/10.14309/ajg.0000000000003048
  12. Cell. 2024 Aug 11. pii: S0092-8674(24)00825-0. [Epub ahead of print]
      During wound healing, different pools of stem cells (SCs) contribute to skin repair. However, how SCs become activated and drive the tissue remodeling essential for skin repair is still poorly understood. Here, by developing a mouse model allowing lineage tracing and basal cell lineage ablation, we monitor SC fate and tissue dynamics during regeneration using confocal and intravital imaging. Analysis of basal cell rearrangements shows dynamic transitions from a solid-like homeostatic state to a fluid-like state allowing tissue remodeling during repair, as predicted by a minimal mathematical modeling of the spatiotemporal dynamics and fate behavior of basal cells. The basal cell layer progressively returns to a solid-like state with re-epithelialization. Bulk, single-cell RNA, and epigenetic profiling of SCs, together with functional experiments, uncover a common regenerative state regulated by the EGFR/AP1 axis activated during tissue fluidization that is essential for skin SC activation and tissue repair.
    Keywords:  AP1 transcription factor; Voronoi model; intravital; lineage ablation; regenerative state; skin; stem cells; tissue fluidity; tissue repair; wound healing
    DOI:  https://doi.org/10.1016/j.cell.2024.07.031
  13. Autophagy. 2024 Aug 23.
      Macroautophagy/autophagy enables lysosomal degradation of a diverse array of intracellular material. This process is essential for normal cellular function and its dysregulation is implicated in many diseases. Given this, there is much interest in understanding autophagic mechanisms of action in order to determine how it can be best targeted therapeutically. In mitophagy, the selective degradation of mitochondria via autophagy, mitochondria first need to be primed with signals that allow the recruitment of the core autophagy machinery to drive the local formation of an autophagosome around the target mitochondrion. To determine how the recruitment of different core autophagy components can drive mitophagy, we took advantage of the mito-QC mitophagy assay (an outer mitochondrial membrane-localized tandem mCherry-GFP tag). By tagging autophagy proteins with an anti-mCherry (or anti-GFP) nanobody, we could recruit them to mitochondria and simultaneously monitor levels of mitophagy. We found that targeting ULK1, ATG16L1 and the different Atg8-family proteins was sufficient to induce mitophagy. Mitochondrial recruitment of ULK1 and the Atg8-family proteins induced a conventional mitophagy pathway, requiring RB1CC1/FIP200, PIK3C3/VPS34 activity and ATG5. Surprisingly, the mitophagy pathway upon recruitment of ATG16L1 proceeded independently of ATG5, although it still required RB1CC1 and PIK3C3/VPS34 activity. In this latter pathway, mitochondria were alternatively delivered to lysosomes via uptake into early endosomes.
    Keywords:  ATG16L1; Atg8; ULK1; nanobody; targeted organelle degradation
    DOI:  https://doi.org/10.1080/15548627.2024.2395149
  14. Nat Cell Biol. 2024 Aug 19.
      Cells migrating through complex three-dimensional environments experience considerable physical challenges, including tensile stress and compression. To move, cells need to resist these forces while also squeezing the large nucleus through confined spaces. This requires highly coordinated cortical contractility. Microtubules can both resist compressive forces and sequester key actomyosin regulators to ensure appropriate activation of contractile forces. Yet, how these two roles are integrated to achieve nuclear transmigration in three dimensions is largely unknown. Here, we demonstrate that compression triggers reinforcement of a dedicated microtubule structure at the rear of the nucleus by the mechanoresponsive recruitment of cytoplasmic linker-associated proteins, which dynamically strengthens and repairs the lattice. These reinforced microtubules form the mechanostat: an adaptive feedback mechanism that allows the cell to both withstand compressive force and spatiotemporally organize contractility signalling pathways. The microtubule mechanostat facilitates nuclear positioning and coordinates force production to enable the cell to pass through constrictions. Disruption of the mechanostat imbalances cortical contractility, stalling migration and ultimately resulting in catastrophic cell rupture. Our findings reveal a role for microtubules as cellular sensors that detect and respond to compressive forces, enabling movement and ensuring survival in mechanically demanding environments.
    DOI:  https://doi.org/10.1038/s41556-024-01476-x
  15. Nature. 2024 Aug 21.
      Mitochondrial membranes define distinct structural and functional compartments. Cristae of the inner mitochondrial membrane (IMM) function as independent bioenergetic units that undergo rapid and transient remodelling, but the significance of this compartmentalized organization is unknown1. Using super-resolution microscopy, here we show that cytosolic IMM vesicles, devoid of outer mitochondrial membrane or mitochondrial matrix, are formed during resting state. These vesicles derived from the IMM (VDIMs) are formed by IMM herniation through pores formed by voltage-dependent anion channel 1 in the outer mitochondrial membrane. Live-cell imaging showed that lysosomes in proximity to mitochondria engulfed the herniating IMM and, aided by the endosomal sorting complex required for transport machinery, led to the formation of VDIMs in a microautophagy-like process, sparing the remainder of the organelle. VDIM formation was enhanced in mitochondria undergoing oxidative stress, suggesting their potential role in maintenance of mitochondrial function. Furthermore, the formation of VDIMs required calcium release by the reactive oxygen species-activated, lysosomal calcium channel, transient receptor potential mucolipin 1, showing an interorganelle communication pathway for maintenance of mitochondrial homeostasis. Thus, IMM compartmentalization could allow for the selective removal of damaged IMM sections via VDIMs, which should protect mitochondria from localized injury. Our findings show a new pathway of intramitochondrial quality control.
    DOI:  https://doi.org/10.1038/s41586-024-07835-w
  16. Lancet Gastroenterol Hepatol. 2024 Aug 16. pii: S2468-1253(24)00197-3. [Epub ahead of print]
       BACKGROUND: A standardised dose-reduction strategy has not been established for the widely used gemcitabine plus nab-paclitaxel regimen in patients with metastatic pancreatic ductal adenocarcinoma. We aimed to investigate the efficacy and tolerability of alternating treatment cycles of nab-paclitaxel-gemcitabine combination therapy and gemcitabine alone versus continuous treatment with the nab-paclitaxel-gemcitabine combination.
    METHODS: ALPACA was a randomised, open-label, phase 2 trial conducted at 29 study centres across Germany. Patients aged 18 years or older with a histologically or cytologically confirmed diagnosis of metastatic pancreatic ductal adenocarcinoma who had not been previously treated for advanced disease were enrolled. After an induction phase with three cycles of nab-paclitaxel-gemcitabine combination therapy (nab-paclitaxel 125 mg/m2 and gemcitabine 1000 mg/m2 administered intravenously on days 1, 8, and 15 of each 28-day cycle), patients were randomly assigned (1:1) by stratified permuted block randomisation either to continue treatment with standard nab-paclitaxel-gemcitabine or to receive alternating cycles of nab-paclitaxel-gemcitabine and gemcitabine alone. Patients and investigators were not masked to treatment allocation. Randomisation was done centrally by the study statistician using a computer-generated randomisation list, and was stratified by Karnofsky Performance Status and presence of liver metastases. The primary endpoint was the derivation of an unbiased point estimate and an associated confidence interval with a confidence coefficient of 80% for the hazard ratio (HR) for overall survival after randomisation, without testing a specific hypothesis, analysed by intention to treat in all patients who started randomised treatment. Safety was analysed according to treatment received. This trial is registered with ClinicalTrials.gov, NCT02564146, and is completed.
    FINDINGS: Between May 27, 2016, and May 27, 2021, 325 patients were enrolled. Following three cycles of induction treatment, 174 patients were randomly assigned: 85 to continue receiving standard nab-paclitaxel-gemcitabine, of whom 79 started treatment, and 89 to the alternating treatment schedule, of whom 88 started treatment. Of the 167 patients who started randomised treatment, 88 (53%) were female and 79 (47%) were male. Median overall survival after randomisation was 10·4 months (80% CI 9·2-12·0) in the group that received standard treatment and 10·5 months (10·2-11·1) in the group that received alternating treatment (HR 0·90, 80% CI 0·72-1·13; p=0·56). The most common adverse events of any grade were peripheral neuropathy (59 [74%] of 80 patients in the continuous treatment group vs 53 [62%] of 85 patients in the alternating treatment group) and fatigue (43 [54%] vs 44 [52%]). Treatment-emergent serious adverse events after randomisation occurred in 40 (50%) patients in the continuous treatment group and in 28 (33%) in the alternating treatment group. Fewer treatment-emergent adverse events of grade 3 or higher occurred in patients treated with alternating cycles compared with those receiving standard therapy, especially for peripheral neuropathy (17 [21%] patients in the continuous treatment group vs 12 [14%] in the alternating treatment group) and infections (16 [20%] vs nine [11%]). There were two treatment-related deaths after randomisation, both in the continuous treatment group (one multiple organ dysfunction syndrome, not treated after randomisation, and one interstitial lung disease).
    INTERPRETATION: Our findings suggest that a dose-reduced regimen with alternating cycles of nab-paclitaxel-gemcitabine and gemcitabine alone after three induction cycles is associated with similar overall survival to that for standard treatment with nab-paclitaxel-gemcitabine, but with improved tolerability. We therefore propose that a switch to the alternating schedule could be considered in a clinical setting for patients with metastatic pancreatic cancer who have at least stable disease after three cycles of nab-paclitaxel-gemcitabine treatment.
    FUNDING: Celgene/Bristol Myers Squibb.
    DOI:  https://doi.org/10.1016/S2468-1253(24)00197-3
  17. Sci Adv. 2024 Aug 23. 10(34): eadm9195
      Eukaryotic cells show an astounding ability to remodel their shape and cytoskeleton and to migrate through pores and constrictions smaller than their nuclear diameter. However, the relation of nuclear deformation and migration dynamics in confinement remains unclear. Here, we study the mechanics and dynamics of mesenchymal cancer cell nuclei transitioning through three-dimensional compliant hydrogel channels. We find a biphasic dependence of migration speed and transition frequency on channel width, peaking at widths comparable to the nuclear diameter. Using confocal imaging and hydrogel bead displacement, we determine nuclear deformations and corresponding forces during confined migration. The nucleus deforms reversibly with a reduction in volume during confinement. With decreasing channel width, the nuclear shape during transmigration changes biphasically, concomitant with the transitioning dynamics. Our proposed physical model explains the observed nuclear shapes and transitioning dynamics in terms of the cytoskeletal force generation adapting from purely pulling-based to a combined pulling- and pushing-based mechanism with increasing nuclear confinement.
    DOI:  https://doi.org/10.1126/sciadv.adm9195
  18. ACS Sens. 2024 Aug 21.
      In this study, we introduce a new separation of phases-based activity reporter of kinase (SPARK) for AMP-activated kinase (AMPK), named AMPK-SPARK, which reports the AMPK activation by forming bright fluorescent clusters. Furthermore, we introduce a dual reporter system, named GCaMP-AMPK-SPARK, by incorporating a single-fluorescent protein (FP)-based Ca2+ biosensor, GCaMP6f, into our initial design, enabling simultaneous monitoring of Ca2+ levels and AMPK activity. This system offers the essential quality of information by single-channel fluorescence microscopy without the need for coexpression of different biosensors and elaborate filter layouts to overcome spectral limitations. We used AMPK-SPARK to map endogenous AMPK activity in different cell types and visualized the dynamics of AMPK activation in response to various stimuli. Using GCaMP-AMPK-SPARK, we revealed cell-to-cell heterogeneities in AMPK activation by Ca2+ mobilization. We anticipate that this dual reporter strategy can be employed to study the intricate interplays between different signaling networks and kinase activities.
    Keywords:  AMPK reporter; Ca2+ biosensor; GCaMP; SPARK; fluorescence microscopy; multiplexing; phase separation-based activity reporter of kinase
    DOI:  https://doi.org/10.1021/acssensors.4c01058
  19. Transl Res. 2024 Aug 16. pii: S1931-5244(24)00147-6. [Epub ahead of print]
      Pancreatic ductal adenocarcinoma (PDAC) is often diagnosed at metastatic stage and typically treated with fluorouracil, leucovorin, irinotecan and oxaliplatin (FOLFIRINOX). Few patients benefit from this treatment. Molecular subtypes are prognostic in particularly resectable PDAC and might predict treatment response. This study aims to correlate molecular subtypes in metastatic PDAC with FOLFIRINOX responses using real-world data, providing assistance in counselling patients. We collected 131 RNA-sequenced metastatic biopsies and applied a network-based meta-analysis using published PDAC classifiers. Subsequent survival analysis was performed using the most suitable classifier. For validation, we developed an immunohistochemistry (IHC) classifier using GATA6 and keratin-17 (KRT17), and applied it to 86 formalin-fixed paraffin-embedded samples of advanced PDAC. Lastly, GATA6 knockdown models were generated in PDAC organoids and cell lines. We showed that the PurIST classifier was the most suitable classifier. With this classifier, classical tumors had longer PFS and OS than basal-like tumors (PFS: 216 vs. 78 days, p = 0.0002; OS: 251 vs. 195 days, p = 0.049). The validation cohort showed a similar trend. Importantly, IHC GATA6low patients had significantly shorter survival with FOLFIRINOX (323 vs. 746 days, p = 0.006), but no difference in non-treated patients (61 vs. 54 days, p = 0.925). This suggests that GATA6 H-score predicts therapy response. GATA6 knockdown models did not lead to increased FOLFIRINOX responsiveness. These data suggest a predictive role for subtyping (transcriptomic and GATA6 IHC), though no direct causal relationship was found between GATA6 expression and chemoresistance. GATA6 immunohistochemistry should be seamlessly added to current diagnostics and integrated into upcoming clinical trials.
    Keywords:  carcinoma, pancreatic ductal; drug resistance, neoplasm; folfirinox; gene expression profiling; immunohistochemistry
    DOI:  https://doi.org/10.1016/j.trsl.2024.08.002
  20. Nature. 2024 Aug 21.
      For over a century, fasting regimens have improved health, lifespan and tissue regeneration in diverse organisms, including humans1-6. However, how fasting and post-fast refeeding affect adult stem cells and tumour formation has yet to be explored in depth. Here we demonstrate that post-fast refeeding increases intestinal stem cell (ISC) proliferation and tumour formation; post-fast refeeding augments the regenerative capacity of Lgr5+ ISCs, and loss of the tumour suppressor gene Apc in post-fast-refed ISCs leads to a higher tumour incidence in the small intestine and colon than in the fasted or ad libitum-fed states, demonstrating that post-fast refeeding is a distinct state. Mechanistically, we discovered that robust mTORC1 induction in post-fast-refed ISCs increases protein synthesis via polyamine metabolism to drive these changes, as inhibition of mTORC1, polyamine metabolite production or protein synthesis abrogates the regenerative or tumorigenic effects of post-fast refeeding. Given our findings, fast-refeeding cycles must be carefully considered and tested when planning diet-based strategies for regeneration without increasing cancer risk, as post-fast refeeding leads to a burst in stem-cell-driven regeneration and tumorigenicity.
    DOI:  https://doi.org/10.1038/s41586-024-07840-z
  21. Cell Rep. 2024 Aug 22. pii: S2211-1247(24)01013-1. [Epub ahead of print]43(9): 114662
      Ferroptosis is a form of nonapoptotic cell death characterized by iron-dependent peroxidation of polyunsaturated phospholipids. However, much remains unknown about the regulators of ferroptosis. Here, using CRISPR-Cas9-mediated genetic screening, we identify protein arginine methyltransferase 1 (PRMT1) as a crucial promoter of ferroptosis. We find that PRMT1 decreases the expression of solute carrier family 7 member 11 (SLC7A11) to limit the abundance of intracellular glutathione (GSH). Moreover, we show that PRMT1 interacts with ferroptosis suppressor protein 1 (FSP1), a GSH-independent ferroptosis suppressor, to inhibit the membrane localization and enzymatic activity of FSP1 through arginine dimethylation at R316, thus reducing CoQ10H2 content and inducing ferroptosis sensitivity. Importantly, genetic depletion or pharmacological inhibition of PRMT1 in mice prevents ferroptotic events in the liver and improves the overall survival under concanavalin A (ConA) exposure. Hence, our findings suggest that PRMT1 is a key regulator of ferroptosis and a potential target for antiferroptosis therapeutics.
    Keywords:  ADMA; CP: Immunology; CP: Metabolism; FSP1; PRMT1; SLC7A11; acute liver injury; ferroptosis
    DOI:  https://doi.org/10.1016/j.celrep.2024.114662
  22. Biophysicist (Rockv). 2024 ;5(1): 1-14
      Epithelial mechanics and mechanobiology have become 2 important research fields in life sciences and bioengineering. These fields investigate how physical factors induced by cell adhesion and collective behaviors can directly regulate biologic processes, such as organ development and disease progression. Cell mechanics and mechanobiology thus make exciting biophysics education topics to illustrate how fundamental physics principles play a role in regulating cell biology. However, the field currently lacks hands-on activities that engage students in learning science and outreach programs in these topics. One such area is the development of robust hands-on modules that allow students to observe features of cell shape and mechanics and connect them to fundamental physics principles. Here, we demonstrate a workflow that engages students in studying epithelial cell mechanics by using commercial histology slides of frog skin. We show that by using recently developed artificial intelligence-based image-segmentation tools, students can easily quantify different cell morphologic features in a high-throughput manner. Using our workflow, students can reproduce 2 essential findings in cell mechanics: the common gamma distribution of normalized cell aspect ratio in jammed epithelia and the constant ratio between the nuclear and cellular area. Importantly, because the only required instrument for this active learning module is a readily available light microscope and a computer, our module is relatively low cost, as well as portable. These features make the module scalable for students at various education levels and outreach programs. This highly accessible education module provides a fun and engaging way to introduce students to the world of epithelial tissue mechanics.
    Keywords:  first-year undergraduate; hands-on learning; high/middle school laboratories; interdisciplinary
    DOI:  https://doi.org/10.35459/tbp.2023.000253
  23. J Chem Phys. 2024 Aug 28. pii: 085103. [Epub ahead of print]161(8):
      Biological cell membranes are primarily comprised of a diverse lipid bilayer with multiple phospholipid (lipid) types, each of which is comprised of a hydrophilic headgroup and two hydrophobic hydrocarbon tails. The lipid type determines the molecular structure of head and tail groups, which can affect membrane mechanics at nanoscale and subsequently cell viability under mechanical loading. Hence, using molecular dynamics simulations, the current study investigated seven membrane phospholipids and the effect of their structural differences on physical deformation, mechanoporation damage, and mechanical failure of the membranes under tension. The inspected phospholipids showed similar yield stresses and strains, as well as pore evolution and damage, but significantly different failure strains. In general, failure occurred at a lower strain for lipids with a larger equilibrium area per lipid. The obtained results suggest that larger headgroup structure, greater degree of unsaturation, and tail-length asymmetry influenced the phospholipids' ability to pack against each other, increased the fluidity and equilibrium area per lipid of the membrane, and resulted in lower failure strain. Overall, this study provides insights on how different phospholipid structures affect membrane physical responses at the molecular level and serves as a reference for future studies of more complex membrane systems with intricate biophysical properties.
    DOI:  https://doi.org/10.1063/5.0214893
  24. Database (Oxford). 2024 Aug 19. pii: baae078. [Epub ahead of print]2024
      Genetically engineered mouse models (GEMMs) are vital for elucidating gene function and disease mechanisms. An overwhelming number of GEMM lines have been generated, but endeavors to collect and organize the information of these GEMMs are seriously lagging behind. Only a few databases are developed for the information of current GEMMs, and these databases lack biological descriptions of allele compositions, which poses a challenge for nonexperts in mouse genetics to interpret the genetic information of these mice. Moreover, these databases usually do not provide information on human diseases related to the GEMM, which hinders the dissemination of the insights the GEMM provides as a human disease model. To address these issues, we developed an algorithm to annotate all the allele compositions that have been reported with Python programming and have developed the genetically modified mice information database (GMMID; http://www.gmmid.cn), a user-friendly database that integrates information on GEMMs and related diseases from various databases, including National Center for Biotechnology Information, Mouse Genome Informatics, Online Mendelian Inheritance in Man, International Mouse Phenotyping Consortium, and Jax lab. GMMID provides comprehensive genetic information on >70 055 alleles, 65 520 allele compositions, and ∼4000 diseases, along with biologically meaningful descriptions of alleles and allele combinations. Furthermore, it provides spatiotemporal visualization of anatomical tissues mentioned in these descriptions, shown alongside the allele compositions. Compared to existing mouse databases, GMMID considers the needs of researchers across different disciplines and presents obscure genetic information in an intuitive and easy-to-understand format. It facilitates users in obtaining complete genetic information more efficiently, making it an essential resource for cross-disciplinary researchers. Database URL: http://www.gmmid.cn.
    DOI:  https://doi.org/10.1093/database/baae078
  25. Cell Stem Cell. 2024 Aug 13. pii: S1934-5909(24)00257-1. [Epub ahead of print]
      Regeneration is a heroic biological process that restores tissue architecture and function in the face of day-to-day cell loss or the aftershock of injury. Capacities and mechanisms for regeneration can vary widely among species, organs, and injury contexts. Here, we describe "hallmarks" of regeneration found in diverse settings of the animal kingdom, including activation of a cell source, initiation of regenerative programs in the source, interplay with supporting cell types, and control of tissue size and function. We discuss these hallmarks with an eye toward major challenges and applications of regenerative biology.
    Keywords:  regeneration
    DOI:  https://doi.org/10.1016/j.stem.2024.07.007