Nat Metab co-auth.: B.Thorens

Autophagy regulator ATG5 preserves cerebellar function by safeguarding its glycolytic activity

Janine Tutas 1 2Marianna Tolve 1 2Ebru Özer-Yildiz 1 2Lotte Ickert 1 2Ines Klein 3Quinn Silverman 3Filip Liebsch 4Frederik Dethloff 5Patrick Giavalisco 5Heike Endepols 6 7 8Theodoros Georgomanolis 1Bernd Neumaier 7 8Alexander Drzezga 7 9 10Guenter Schwarz 4 11Bernard Thorens 12Graziana Gatto 3Christian Frezza 1 13Natalia L Kononenko 14 15 16 17

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Abstract

Dysfunctions in autophagy, a cellular mechanism for breaking down components within lysosomes, often lead to neurodegeneration. The specific mechanisms underlying neuronal vulnerability due to autophagy dysfunction remain elusive. Here we show that autophagy contributes to cerebellar Purkinje cell (PC) survival by safeguarding their glycolytic activity. Outside the conventional housekeeping role, autophagy is also involved in the ATG5-mediated regulation of glucose transporter 2 (GLUT2) levels during cerebellar maturation. Autophagy-deficient PCs exhibit GLUT2 accumulation on the plasma membrane, along with increased glucose uptake and alterations in glycolysis. We identify lysophosphatidic acid and serine as glycolytic intermediates that trigger PC death and demonstrate that the deletion of GLUT2 in ATG5-deficient mice mitigates PC neurodegeneration and rescues their ataxic gait. Taken together, this work reveals a mechanism for regulating GLUT2 levels in neurons and provides insights into the neuroprotective role of autophagy by controlling glucose homeostasis in the brain.

Nat Commun auth.: group Fajas

CDK4 inactivation inhibits apoptosis via mitochondria-ER contact remodeling in triple-negative breast cancer

Dorian V Ziegler 1Kanishka Parashar 1Lucia Leal-Esteban 1Jaime López-Alcalá 1 2Wilson Castro 3Nadège Zanou 4Laia Martinez-Carreres 1Katharina Huber 1Xavier Pascal Berney 1María M Malagón 2 5Catherine Roger 1Marie-Agnès Berger 6Yves Gouriou 6Giulia Paone 1Hector Gallart-Ayala 7George Sflomos 8Carlos Ronchi 8Julijana Ivanisevic 7Cathrin Brisken 8 9Jennifer Rieusset 6Melita Irving 3Lluis Fajas 10 11

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Abstract

The energetic demands of proliferating cells during tumorigenesis require close coordination between the cell cycle and metabolism. While CDK4 is known for its role in cell proliferation, its metabolic function in cancer, particularly in triple-negative breast cancer (TNBC), remains unclear. Our study, using genetic and pharmacological approaches, reveals that CDK4 inactivation only modestly impacts TNBC cell proliferation and tumor formation. Notably, CDK4 depletion or long-term CDK4/6 inhibition confers resistance to apoptosis in TNBC cells. Mechanistically, CDK4 enhances mitochondria-endoplasmic reticulum contact (MERCs) formation, promoting mitochondrial fission and ER-mitochondrial calcium signaling, which are crucial for TNBC metabolic flexibility. Phosphoproteomic analysis identified CDK4’s role in regulating PKA activity at MERCs. In this work, we highlight CDK4’s role in mitochondrial apoptosis inhibition and suggest that targeting MERCs-associated metabolic shifts could enhance TNBC therapy.

Commun Biol, co-auth.:M.Quadroni

Arabidopsis conditional photosynthesis mutants abc1k1 and var2 accumulate partially processed thylakoid preproteins and are defective in chloroplast biogenesis

Joy Collombat 1Manfredo Quadroni 2Véronique Douet 1Rosa Pipitone 3Fiamma Longoni 1Felix Kessler 4

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Abstract

Photosynthetic activity is established during chloroplast biogenesis. In this study we used 680 nm red light to overexcite Photosystem II and disrupt photosynthesis in two conditional mutants (var2 and abc1k1) which reversibly arrested chloroplast biogenesis. During biogenesis, chloroplasts import most proteins associated with photosynthesis. Some of these must be inserted in or transported across the thylakoid membrane into the thylakoid lumen. They are synthesized in the cytoplasm with cleavable targeting sequences and the lumenal ones have bi-partite targeting sequences (first for the chloroplast envelope, second for the thylakoid membrane). Cleavage of these peptides is required to establish photosynthesis and a critical step of chloroplast biogenesis. We employ a combination of Western blotting and mass spectrometry to analyze proteins in var2 and abc1k1. Under red light, var2 and abc1k1 accumulated incompletely cleaved processing intermediates of thylakoid proteins. These findings correlated with colorless cotyledons, and defects in both chloroplast morphology and photosynthesis. Together the results provide evidence for the requirement of active photosynthesis for processing of photosystem-associated thylakoid proteins and concomitantly progression of chloroplast biogenesis.

Cell Mol Life Sci: co-auth.: W.Wahli

Palmitate potentiates the SMAD3-PAI-1 pathway by reducing nuclear GDF15 levels

Marta Montori-Grau 1 2 3 4Emma Barroso 1 2 3 4Javier Jurado-Aguilar 1 2 3 4Mona Peyman 1 2 3 4Walter Wahli 5 6 7Xavier Palomer 1 2 3 4Manuel Vázquez-Carrera 8 9 10 11

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Abstract

Nuclear growth differentiation factor 15 (GDF15) reduces the binding of the mothers’ against decapentaplegic homolog (SMAD) complex to its DNA-binding elements. However, the stimuli that control this process are unknown. Here, we examined whether saturated fatty acids (FA), particularly palmitate, regulate nuclear GDF15 levels and the activation of the SMAD3 pathway in human skeletal myotubes and mouse skeletal muscle, where most insulin-stimulated glucose use occurs in the whole organism. Human LHCN-M2 myotubes and skeletal muscle from wild-type and Gdf15-/- mice fed a standard (STD) or a high-fat (HFD) diet were subjected to a series of studies to investigate the involvement of lipids in nuclear GDF15 levels and the activation of the SMAD3 pathway. The saturated FA palmitate, but not the monounsaturated FA oleate, increased the expression of GDF15 in human myotubes and, unexpectedly, decreased its nuclear levels. This reduction was prevented by the nuclear export inhibitor leptomycin B. The decrease in nuclear GDF15 levels caused by palmitate was accompanied by increases in SMAD3 protein levels and in the expression of its target gene SERPINE1, which encodes plasminogen activator inhibitor 1 (PAI-1). HFD-fed Gdf15-/- mice displayed aggravated glucose intolerance compared to HFD-fed WT mice, with increased levels of SMAD3 and PAI-1 in the skeletal muscle. The increased PAI-1 levels in the skeletal muscle of HFD-fed Gdf15-/- mice were accompanied by a reduction in one of its targets, hepatocyte growth factor (HGF)α, a cytokine involved in glucose metabolism. Interestingly, PAI-1 acts as a ligand of signal transducer and activator of transcription 3 (STAT3) and the phosphorylation of this transcription factor was exacerbated in HFD-fed Gdf15-/- mice compared to HFD-fed WT mice. At the same time, the protein levels of insulin receptor substrate 1 (IRS-1) were reduced. These findings uncover a potential novel mechanism through which palmitate induces the SMAD3-PAI-1 pathway to promote insulin resistance in skeletal muscle by reducing nuclear GDF15 levels.

EMBO J. auth.: group Gatfield

MCTS2 and distinct eIF2D roles in uORF-dependent translation regulation revealed by in vitro re-initiation assays

Romane Meurs 1Mara De Matos 1Adrian Bothe 2Nicolas Guex 3Tobias Weber 4Aurelio A Teleman 4Nenad Ban 2David Gatfield 5

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Free article

Abstract

Ribosomes scanning from the mRNA 5′ cap to the start codon may initiate at upstream open reading frames (uORFs), decreasing protein biosynthesis. Termination at a uORF can lead to re-initiation, where 40S subunits resume scanning and initiate another translation event downstream. The noncanonical translation factors MCTS1-DENR participate in re-initiation at specific uORFs, but knowledge of other trans-acting factors or uORF features influencing re-initiation is limited. Here, we establish a cell-free re-initiation assay using HeLa lysates to address this question. Comparing in vivo and in vitro re-initiation on uORF-containing reporters, we validate MCTS1-DENR-dependent re-initiation in vitro. Using this system and ribosome profiling in cells, we found that knockdown of the MCTS1-DENR homolog eIF2D causes widespread gene deregulation unrelated to uORF translation, and thus distinct to MCTS1-DENR-dependent re-initiation regulation. Additionally, we identified MCTS2, encoded by an Mcts1 retrogene, as a DENR partner promoting re-initiation in vitro, providing a plausible explanation for clinical differences associated with DENR vs. MCTS1 mutations in humans.

Cell Commun Signal. co-auth.: W.Wahli

PPARβ/δ upregulates the insulin receptor β subunit in skeletal muscle by reducing lysosomal activity and EphB4 levels

Jue-Rui Wang # 1 2 3Javier Jurado-Aguilar # 1 2 3Emma Barroso 1 2 3Ricardo Rodríguez-Calvo 2 4Antoni Camins 1 5 6Walter Wahli 7 8Xavier Palomer 1 2 3Manuel Vázquez-Carrera 9 10 11 12

Abstract

Background: The increased degradation of the insulin receptor β subunit (InsRβ) in lysosomes contributes to the development of insulin resistance and type 2 diabetes mellitus. Endoplasmic reticulum (ER) stress contributes to insulin resistance through several mechanisms, including the reduction of InsRβ levels. Here, we examined how peroxisome proliferator-activated receptor (PPAR)β/δ regulates InsRβ levels in mouse skeletal muscle and C2C12 myotubes exposed to the ER stressor tunicamycin.

Methods: Wild-type (WT) and Ppard-/- mice, WT mice treated with vehicle or the PPARβ/δ agonist GW501516, and C2C12 myotubes treated with the ER stressor tunicamycin or different activators or inhibitors were used.

Results: Ppard-/- mice displayed reduced InsRβ protein levels in their skeletal muscle compared to wild-type (WT) mice, while the PPARβ/δ agonist GW501516 increased its levels in WT mice. Co-incubation of tunicamycin-exposed C2C12 myotubes with GW501516 partially reversed the decrease in InsRβ protein levels, attenuating both ER stress and the increase in lysosomal activity. In addition, the protein levels of the tyrosine kinase ephrin receptor B4 (EphB4), which binds to the InsRβ and facilitates its endocytosis and degradation in lysosomes, were increased in the skeletal muscle of Ppard-/- mice, with GW501516 reducing its levels in the skeletal muscle of WT mice.

Conclusions: Overall, these findings reveal that PPARβ/δ activation increases InsRβ levels by alleviating ER stress and lysosomal degradation.