bims-lypmec Biomed News
on Lysosomal positioning and metabolism in cardiomyocytes
Issue of 2024–12–29
eleven papers selected by
Satoru Kobayashi, New York Institute of Technology
  1. The Cullin3-Rbx1-KLHL9 E3 ubiquitin ligase complex ubiquitinates Rheb and supports amino acid-induced mTORC1 activation.
  2. Impairment of lipid homeostasis causes lysosomal accumulation of endogenous protein aggregates through ESCRT disruption.
  3. Tagless LysoIP for immunoaffinity enrichment of native lysosomes from clinical samples.
  4. A Molecular Logic Gate Enables Unconventional Super-resolution Same-Day Imaging of Lysosome Membrane in Live Cells.
  5. NOVEL APPROACHES IN TARGETING MEMBRANE AND SECRETED PROTEINS FOR LYSOSOMAL DEGRADATION.
  6. Reconsidering the selectivity of bulk autophagy: cargo hitchhiking specifies cargo for degradation.
  7. Spns1-dependent endocardial lysosomal function drives valve morphogenesis through Notch1-signaling.
  8. Wide-ranging cellular functions of ion channels and lipid scramblases in the structurally related TMC, TMEM16 and TMEM63 families.
  9. Research progress on the relationship between free fatty acid profile and type 2 diabetes complicated by coronary heart disease.
  10. Calmodulin enhances mTORC1 signaling by preventing TSC2-Rheb binding.
  11. Prediction of metabolic subphenotypes of type 2 diabetes via continuous glucose monitoring and machine learning.
  1. Cell Rep. 2024 Dec 20. pii: S2211-1247(24)01452-9. [Epub ahead of print]44(1): 115101
    The Cullin3-Rbx1-KLHL9 E3 ubiquitin ligase complex ubiquitinates Rheb and supports amino acid-induced mTORC1 activation.
    Yao Yao, Sungki Hong, Shota Yoshida, Vinamra Swaroop, Brady Curtin, Ken Inoki.
      Mechanistic target of rapamycin complex 1 (mTORC1) is recruited to the lysosomal membrane by the active Rag heterodimer, where mTORC1 interacts with active Rheb for its activation. It has been shown that polyubiquitination of Rheb is crucial for enhancing its interaction with mTORC1 on the lysosome. However, the specific ubiquitin ligases for Rheb, which promotes mTORC1 activation, remain elusive. We report that the CUL3-RBX1-KLHL9 E3 ubiquitin ligase complex is translocated to the lysosome and ubiquitinates Rheb in response to amino acid stimulation. KLHL9 serves as an essential adaptor for CUL3-RBX1 to target Rheb on the lysosome. Deleting either CUL3, RBX1, or KLHL9 diminishes Rheb ubiquitination and reduces amino acid-induced mTORC1 activation without impacting lysosomal mTORC1 localization or Akt activity. Thus, the CUL3-RBX1-KLHL9 complex functions as a mTORC1 activator by acting as an E3 ubiquitin ligase for Rheb and supports amino acid-induced mTORC1 activation.
    Keywords:  CP: Cell biology; CUL3; KLHL9; Rheb; lysosome; mTORC1; ubiquitination
    DOI:  https://doi.org/10.1016/j.celrep.2024.115101
  2. Elife. 2024 Dec 23. pii: RP86194. [Epub ahead of print]12
    Impairment of lipid homeostasis causes lysosomal accumulation of endogenous protein aggregates through ESCRT disruption.
    John Yong, Jacqueline E Villalta, Ngoc Vu, Matthew A Kukurugya, Niclas Olsson, Magdalena Preciado López, Julia R Lazzari-Dean, Kayley Hake, Fiona E McAllister, Bryson D Bennett, Calvin H Jan.
      Protein aggregation increases during aging and is a pathological hallmark of many age-related diseases. Protein homeostasis (proteostasis) depends on a core network of factors directly influencing protein production, folding, trafficking, and degradation. Cellular proteostasis also depends on the overall composition of the proteome and numerous environmental variables. Modulating this cellular proteostasis state can influence the stability of multiple endogenous proteins, yet the factors contributing to this state remain incompletely characterized. Here, we performed genome-wide CRISPRi screens to elucidate the modulators of proteostasis state in mammalian cells, using a fluorescent dye to monitor endogenous protein aggregation. These screens identified known components of the proteostasis network and uncovered a novel link between protein and lipid homeostasis. Increasing lipid uptake and/or disrupting lipid metabolism promotes the accumulation of sphingomyelins and cholesterol esters and drives the formation of detergent-insoluble protein aggregates at the lysosome. Proteome profiling of lysosomes revealed ESCRT accumulation, suggesting disruption of ESCRT disassembly, lysosomal membrane repair, and microautophagy. Lipid dysregulation leads to lysosomal membrane permeabilization but does not otherwise impact fundamental aspects of lysosomal and proteasomal functions. Together, these results demonstrate that lipid dysregulation disrupts ESCRT function and impairs proteostasis.
    Keywords:  CRISPR; ESCRT; aggregation; cell biology; human; lipid dysregulation; lysosome; proteostasis
    DOI:  https://doi.org/10.7554/eLife.86194
  3. J Clin Invest. 2024 Dec 26. pii: e183592. [Epub ahead of print]
    Tagless LysoIP for immunoaffinity enrichment of native lysosomes from clinical samples.
    Daniel Saarela, Pawel Lis, Sara Gomes, Raja S Nirujogi, Wentao Dong, Eshaan S Rawat, Sophie Glendinning, Karolina Zeneviciute, Enrico Bagnoli, Rotimi Fasimoye, Cindy Lin, Kwamina Nyame, Fanni A Boros, Friederike Zunke, Frederic Lamoliatte, Sadik Elshani, Matthew Jaconelli, Judith Jm Jans, Margriet A Huisman, Christian Posern, Lena M Westermann, Angela Schulz, Peter M van Hasselt, Dario R Alessi, Monther Abu-Remaileh, Esther M Sammler.
      Lysosomes are implicated in a wide spectrum of human diseases including monogenic lysosomal storage disorders (LSDs), age-associated neurodegeneration and cancer. Profiling lysosomal content using tag-based lysosomal immunoprecipitation (LysoTagIP) in cell and animal models has substantially moved the field forward, but studying lysosomal dysfunction in human patients remains challenging. Here, we report the development of the 'tagless LysoIP' method, designed to enable the rapid enrichment of lysosomes, via immunoprecipitation, using the endogenous integral lysosomal membrane protein TMEM192, directly from clinical samples and human cell lines (e.g., induced pluripotent stem cell derived neurons). Isolated lysosomes were intact and suitable for subsequent multimodal omics analyses. To validate our approach, we applied the tagless LysoIP to enrich lysosomes from peripheral blood mononuclear cells derived from fresh blood of healthy donors and patients with CLN3 disease, an autosomal recessive neurodegenerative LSD. Metabolic profiling of isolated lysosomes revealed massive accumulation of glycerophosphodiesters (GPDs) in patients' lysosomes. Interestingly, a patient with a milder phenotype and genotype displayed lower accumulation of lysosomal GPDs, consistent with their potential role as disease biomarkers. Altogether, the tagless LysoIP provides a framework to study native lysosomes from patient samples, identify disease biomarkers, and discover human-relevant disease mechanisms.
    Keywords:  Cell biology; Genetic diseases; Lysosomes; Neurodegeneration; Neuroscience
    DOI:  https://doi.org/10.1172/JCI183592
  4. Anal Chem. 2024 Dec 25.
    A Molecular Logic Gate Enables Unconventional Super-resolution Same-Day Imaging of Lysosome Membrane in Live Cells.
    Yangzi Xie, Yuan Luo, Wen Li, Yi Zhou, Yongfan Men, Fufeng Liu, Fan Pan, Lintao Cai, Zhe Jiao, Pengfei Zhang.
      Lysosomes are acidic membrane-bound organelles that aid digestion, excretion, and cell renewal. The lysosomal membranes are essential for maintaining lysosomal functions and cellular homeostasis. In this work, we developed a molecular "NOR" logic gate, SIATFluor-580L, by introducing malachite green into the spirocyclic rhodamine. SIATFluor-580L requires restriction of molecular rotation of the malachite green motif (Input 1, tight membrane structure) and a large amount of H+ ions to convert the spirocyclic rhodamine into the zwitterionic form (Input 2, acidic environment) to produce a fluorescent product (Output), providing a fluorogenic probe to visualize the lysosomal membrane dynamics in living cells with subdiffraction resolution by using HyVolution (also known as Lightning), an unconventional and inexpensive super-resolution imaging approach based on a basic confocal optical system.
    DOI:  https://doi.org/10.1021/acs.analchem.4c05907
  5. Chembiochem. 2024 Dec 23. e202400887
    NOVEL APPROACHES IN TARGETING MEMBRANE AND SECRETED PROTEINS FOR LYSOSOMAL DEGRADATION.
    Mohamed Eldeeb, Grace Hohman, Michael Shahid.
      Protein degradation is pivotal for all biochemical aspects of cellular function. In mammalian cells, protein degradation is mediated mainly by the ubiquitin proteasome system (UPS) and the autophagic-lysosomal system (ALS). Over the last two decades, different types of targeted protein degradation approaches have been developed including proteolysis targeting chimeras (PROTACs) and lysosome targeting chimeras (LYTACs), which employ the UPS to degrade intracellular proteins and the ALS to degrade extracellular and membrane proteins respectively. Nevertheless, current targeted membrane protein degradation approaches face some inherent challenges including limited target protein degradation efficacy and cell type specific applicability. Herein, we highlight a recent development of novel targeted membrane protein degradation modalities that exhibit wide-applicability and high protein degradation efficiency. These novel membrane protein degraders hold tremendous promise as new pharmacological and biochemical tools in targeting membrane and secretory proteins for lysosomal degradation.
    Keywords:  Lysosome; Protein degradation; Proteolysis; autophagy; membrane proteins
    DOI:  https://doi.org/10.1002/cbic.202400887
  6. Autophagy. 2024 Dec 27.
    Reconsidering the selectivity of bulk autophagy: cargo hitchhiking specifies cargo for degradation.
    Eigo Takeda, Alexander I May, Yoshinori Ohsumi.
      Bulk macroautophagy/autophagy, typically induced by starvation, is generally thought to non-selectively isolate cytosolic components for degradation. However, a detailed analysis of bulk autophagy cargo has not been conducted. We recently employed mass spectrometry to analyze the contents of isolated autophagic bodies. In this process, we uncovered Hab1 (Highly enriched in Autophagic Bodies 1), a novel protein, that is preferentially delivered via autophagy. Hab1 is a receptor protein that binds Atg8-PE and ribosomes at its N- and C-termini, respectively. We found that ribosome-bound Hab1 is preferentially delivered to the vacuole by "'hitchhiking'" on phagophores/isolation membranes formed during bulk autophagy. This hitchhiking mechanism confers selectivity to bulk autophagy.
    Keywords:  Atg8; Hab1; Saccharomyces cerevisiae; hitchhiking; ribosome
    DOI:  https://doi.org/10.1080/15548627.2024.2447209
  7. iScience. 2024 Dec 20. 27(12): 111406
    Spns1-dependent endocardial lysosomal function drives valve morphogenesis through Notch1-signaling.
    Myra N Chávez, Prateek Arora, Marco Meer, Ines J Marques, Alexander Ernst, Rodrigo A Morales Castro, Nadia Mercader.
      Autophagy-lysosomal degradation is a conserved homeostatic process considered to be crucial for cardiac morphogenesis. However, both its cell specificity and functional role during heart development remain unclear. Here, we introduced zebrafish models to visualize autophagic vesicles in vivo and track their temporal and cellular localization in the larval heart. We observed a significant accumulation of autolysosomal and lysosomal vesicles in the atrioventricular and bulboventricular regions and their respective valves. We addressed the role of lysosomal degradation based on the Spinster homolog 1 (spns1) mutant (not really started, nrs). n rs larvae displayed morphological and functional cardiac defects, including abnormal endocardial organization, impaired valve formation and retrograde blood flow. Single-nuclear transcriptome analyses revealed endocardial-specific differences in lysosome-related genes and alterations of notch1-signalling. Endocardial-specific overexpression of spns1 and notch1 rescued features of valve formation and function. Altogether, our results reveal a cell-autonomous role of lysosomal processing during cardiac valve formation affecting notch1-signalling.
    Keywords:  Cell biology; Developmental biology; Model organism; Molecular biology; Transcriptomics
    DOI:  https://doi.org/10.1016/j.isci.2024.111406
  8. Nat Struct Mol Biol. 2024 Dec 23.
    Wide-ranging cellular functions of ion channels and lipid scramblases in the structurally related TMC, TMEM16 and TMEM63 families.
    Lily Yeh Jan, Yuh Nung Jan.
      Calcium (Ca2+)-activated ion channels and lipid scramblases in the transmembrane protein 16 (TMEM16) family are structurally related to mechanosensitive ion channels in the TMEM63 and transmembrane channel-like (TMC) families. Members of this structurally related superfamily share similarities in gating transitions and serve a wide range of physiological functions, which is evident from their disease associations. The TMEM16, TMEM63 and TMC families include members with important functions in the cell membrane and/or intracellular organelles such as the endoplasmic reticulum, membrane contact sites, endosomes and lysosomes. Moreover, some members of the TMEM16 family and the TMC family perform dual functions of ion channel and lipid scramblase, leading to intriguing physiological implications. In addition to their physiological functions such as mediating phosphatidylserine exposure and facilitation of extracellular vesicle generation and cell fusion, scramblases are involved in the entry and replication of enveloped viruses. Comparisons of structurally diverse scramblases may uncover features in the lipid-scrambling mechanisms that are likely shared by scramblases.
    DOI:  https://doi.org/10.1038/s41594-024-01444-x
  9. Front Endocrinol (Lausanne). 2024 ;15 1503704
    Research progress on the relationship between free fatty acid profile and type 2 diabetes complicated by coronary heart disease.
    Xiuyan Liu, Ming Gong, Na Wu.
      Patients with type 2 diabetes mellitus (T2DM) have a 2 to 3 times higher risk of cardiovascular disease compared to non-diabetic individuals, and cardiovascular disease has consistently been a leading cause of death among diabetic patients. Therefore, preventing cardiovascular disease in diabetic patients remains a significant challenge. In addition to classic indicators such as cholesterol and lipoproteins, previous studies have demonstrated that plasma level of free fatty acid (FFA) is closely related to the occurrence of atherosclerosis, particularly in T2DM patients. In recent years, with further research and advancements in testing technologies, the FFA profile has garnered widespread attention. The FFA profile includes many different types of FFAs, and changes in the plasma FFA profile and concentrations in T2DM patients may lead to the development of insulin resistance, causing damage to vascular endothelial cells and promoting the occurrence and progression of atherosclerosis. Furthermore, some FFAs have shown potential in predicting cardiovascular complications in T2DM and are associated with the severity of these complications. Here, we aim to review the changes in the FFA profile in T2DM and discuss the relationship between the FFA profile and the occurrence of vascular complications in T2DM.
    Keywords:  atherosclerosis; coronary heart disease; free fatty acid profile; polyunsaturated fatty acids; type 2 diabetes
    DOI:  https://doi.org/10.3389/fendo.2024.1503704
  10. J Biol Chem. 2024 Dec 21. pii: S0021-9258(24)02624-3. [Epub ahead of print] 108122
    Calmodulin enhances mTORC1 signaling by preventing TSC2-Rheb binding.
    Yuna Amemiya, Yuichiro Ioi, Makoto Araki, Kenji Kontani, Masatoshi Maki, Hideki Shibata, Terunao Takahara.
      The mechanistic target of rapamycin complex 1 (mTORC1) functions as a master regulator of cell growth and proliferation. We previously demonstrated that intracellular calcium ion (Ca2+) concentration modulates the mTORC1 pathway via binding of the Ca2+ sensor protein calmodulin (CaM) to tuberous sclerosis complex 2 (TSC2), a critical negative regulator of mTORC1. However, the precise molecular mechanism by which Ca2+/CaM modulates mTORC1 activity remains unclear. Here, we performed a binding assay based on nano-luciferase reconstitution, a method for detecting weak interactions between TSC2 and its target, Ras homolog enriched in brain (Rheb), an activator of mTORC1. CaM inhibited the binding of TSC2 to Rheb in a Ca2+-dependent manner. Live-cell imaging analysis indicated increased interaction between the CaM-binding region of TSC2 and CaM in response to elevated intracellular Ca2+ levels. Furthermore, treatment with carbachol, an acetylcholine analog, elevated intracellular Ca2+ levels, and activated mTORC1. Notably, carbachol-induced activation of mTORC1 was inhibited by CaM inhibitors, corroborating the role of Ca2+/CaM in promoting the mTORC1 pathway. Consistent with the effect of Ca2+/CaM on the TSC2-Rheb interaction, increased intracellular Ca2+ concentration promoted the dissociation of TSC2 from lysosomes without affecting Akt-dependent phosphorylation of TSC2, suggesting that the regulatory mechanism of TSC2 by Ca2+/CaM is distinct from the previously established action mechanism of TSC2. Collectively, our findings offer mechanistic insights into TSC2-Rheb regulation mediated by Ca2+/CaM, which links Ca2+ signaling to mTORC1 activation.
    Keywords:  Rheb; calcium; calmodulin; mechanistic target of rapamycin complex 1; tuberous sclerosis complex
    DOI:  https://doi.org/10.1016/j.jbc.2024.108122
  11. Nat Biomed Eng. 2024 Dec 23.
    Prediction of metabolic subphenotypes of type 2 diabetes via continuous glucose monitoring and machine learning.
    Ahmed A Metwally, Dalia Perelman, Heyjun Park, Yue Wu, Alokkumar Jha, Seth Sharp, Alessandra Celli, Ekrem Ayhan, Fahim Abbasi, Anna L Gloyn, Tracey McLaughlin, Michael P Snyder.
      The classification of type 2 diabetes and prediabetes does not consider heterogeneity in the pathophysiology of glucose dysregulation. Here we show that prediabetes is characterized by metabolic heterogeneity, and that metabolic subphenotypes can be predicted by the shape of the glucose curve measured via a continuous glucose monitor (CGM) during standardized oral glucose-tolerance tests (OGTTs) performed in at-home settings. Gold-standard metabolic tests in 32 individuals with early glucose dysregulation revealed dominant or co-dominant subphenotypes (muscle or hepatic insulin-resistance phenotypes in 34% of the individuals, and β-cell-dysfunction or impaired-incretin-action phenotypes in 40% of them). Machine-learning models trained with glucose time series from OGTTs from the 32 individuals predicted the subphenotypes with areas under the curve (AUCs) of 95% for muscle insulin resistance, 89% for β-cell deficiency and 88% for impaired incretin action. With CGM-generated glucose curves obtained during at-home OGTTs, the models predicted the muscle-insulin-resistance and β-cell-deficiency subphenotypes of 29 individuals with AUCs of 88% and 84%, respectively. At-home identification of metabolic subphenotypes via a CGM may aid the risk stratification of individuals with early glucose dysregulation.
    DOI:  https://doi.org/10.1038/s41551-024-01311-6
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