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Cell Reports Oct 2023Loss of transcription-coupled histone H3 lysine 36 trimethylation (H3K36me3) contributes to shorter lifespans in eukaryotes. However, the molecular mechanism of the...
Loss of transcription-coupled histone H3 lysine 36 trimethylation (H3K36me3) contributes to shorter lifespans in eukaryotes. However, the molecular mechanism of the decline of H3K36me3 during aging remains poorly understood. Here, we report that the degradation of the methyltransferase Set2 is the cause of decreased H3K36me3 levels during chronological aging in budding yeast. We show that Set2 protein degradation during cellular senescence and chronological aging is mainly mediated by the ubiquitin-conjugating E2 enzyme Ubc3 and the E3 ligase Bre1. Lack of Bre1 or abolishment of the ubiquitination stabilizes Set2 protein, sustains H3K36me3 levels at the aging-related gene loci, and upregulates their gene expression, thus leading to extended chronological lifespan. We further illustrate that Gcn5-mediated Set2 acetylation is a prerequisite for Bre1-catalyzed Set2 polyubiquitination and proteolysis during aging. We propose that two sequential post-translational modifications regulate Set2 homeostasis, suggesting a potential strategy to target the Gcn5-Bre1-Set2 axis for intervention of longevity.
Topics: Histones; Methylation; Methyltransferases; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins; Ubiquitin-Conjugating Enzymes; Aging
PubMed: 37796660
DOI: 10.1016/j.celrep.2023.113186 -
The Journal of Cell Biology Jul 2023During autophagy, rapid membrane assembly expands small phagophores into large double-membrane autophagosomes. Theoretical modeling predicts that the majority of...
During autophagy, rapid membrane assembly expands small phagophores into large double-membrane autophagosomes. Theoretical modeling predicts that the majority of autophagosomal phospholipids are derived from highly efficient non-vesicular phospholipid transfer (PLT) across phagophore-ER contacts (PERCS). Currently, the phagophore-ER tether Atg2 is the only PLT protein known to drive phagophore expansion in vivo. Here, our quantitative live-cell imaging analysis reveals a poor correlation between the duration and size of forming autophagosomes and the number of Atg2 molecules at PERCS of starving yeast cells. Strikingly, we find that Atg2-mediated PLT is non-rate limiting for autophagosome biogenesis because membrane tether and the PLT protein Vps13 localizes to the rim and promotes the expansion of phagophores in parallel with Atg2. In the absence of Vps13, the number of Atg2 molecules at PERCS determines the duration and size of forming autophagosomes with an apparent in vivo transfer rate of ∼200 phospholipids per Atg2 molecule and second. We propose that conserved PLT proteins cooperate in channeling phospholipids across organelle contact sites for non-rate-limiting membrane assembly during autophagosome biogenesis.
Topics: Autophagosomes; Phospholipids; Endoplasmic Reticulum; Autophagy; Saccharomyces cerevisiae; Autophagy-Related Proteins; Saccharomyces cerevisiae Proteins
PubMed: 37115156
DOI: 10.1083/jcb.202211039 -
Cells Nov 2023The TEM8 protein represents an emerging biomarker in many solid tumor histologies. Given the various roles it plays in oncogenesis, including but not limited to... (Review)
Review
The TEM8 protein represents an emerging biomarker in many solid tumor histologies. Given the various roles it plays in oncogenesis, including but not limited to angiogenesis, epithelial-to-mesenchymal transition, and cell migration, TEM8 has recently served and will continue to serve as the target of novel oncologic therapies. We review herein the role of TEM8 in oncogenesis. We review its normal function, highlight the additional roles it plays in the tumor microenvironment, and synthesize pre-clinical and clinical data currently available. We underline the protein's prognostic and predictive abilities in various solid tumors by (1) highlighting its association with more aggressive disease biology and poor clinical outcomes and (2) assessing its associated clinical trial landscape. Finally, we offer future directions for clinical studies involving TEM8, including incorporating pre-clinical agents into clinical trials and combining previously tested oncologic therapies with currently available treatments, such as immunotherapy.
Topics: Humans; Receptors, Cell Surface; Neoplasm Proteins; Microfilament Proteins; Neoplasms; Cell Transformation, Neoplastic; Carcinogenesis; Biology; Tumor Microenvironment
PubMed: 37998358
DOI: 10.3390/cells12222623 -
Science Advances Nov 2023Parkinson's disease (PD) is characterized by the pathologic aggregation and prion-like propagation of α-synuclein (α-syn). Emerging evidence shows that fungal...
Parkinson's disease (PD) is characterized by the pathologic aggregation and prion-like propagation of α-synuclein (α-syn). Emerging evidence shows that fungal infections increase the incidence of PD. However, the molecular mechanisms by which fungi promote the onset of PD are poorly understood. Here, we show that nasal infection with () in α-syn A53T transgenic mice accelerates the aggregation of α-syn. Furthermore, we found that Sup35, a prion protein from , is the key factor initiating α-syn pathology induced by . Sup35 interacts with α-syn and accelerates its aggregation in vitro. Notably, injection of Sup35 fibrils into the striatum of wild-type mice led to α-syn pathology and PD-like motor impairment. The Sup35-seeded α-syn fibrils showed enhanced seeding activity and neurotoxicity compared with pure α-syn fibrils in vitro and in vivo. Together, these observations indicate that the yeast prion protein Sup35 initiates α-syn pathology in PD.
Topics: Animals; Mice; alpha-Synuclein; Mice, Transgenic; Parkinson Disease; Prion Proteins; Prions; Saccharomyces cerevisiae
PubMed: 37910610
DOI: 10.1126/sciadv.adj1092 -
Platelets Dec 2024Protein S (PS) is a vital endogenous anticoagulant. It plays a crucial role in regulating coagulation by acting as a cofactor for the activated protein C (APC) and... (Review)
Review
Protein S (PS) is a vital endogenous anticoagulant. It plays a crucial role in regulating coagulation by acting as a cofactor for the activated protein C (APC) and tissue factor pathway inhibitor (TFPI) pathways. Additionally, it possesses direct anticoagulant properties by impeding the intrinsic tenase and prothrombinase complexes. Protein S oversees the coagulation process in both the initiation and propagation stages through these roles. The significance of protein S in regulating blood clotting can be inferred from the significant correlation between deficits in protein S and an elevated susceptibility to venous thrombosis. This is likely because activated protein C and tissue factor pathway inhibitor exhibit low efficacy as anticoagulants when no cofactors exist. The precise biochemical mechanisms underlying the roles of protein S cofactors have yet to be fully elucidated. Nevertheless, recent scientific breakthroughs have significantly enhanced comprehension findings for these functions. The diagnosis of protein S deficiency, both from a technical and genetic standpoint, is still a subject of debate due to the complex structural characteristics of the condition. This paper will provide an in-depth review of the molecular structure of protein S and its hemostatic effects. Furthermore, we shall address the insufficiency of protein S and its methods of diagnosis and treatment.
Topics: Humans; Anticoagulants; Protein C; Blood Coagulation; Hemostatics
PubMed: 38602463
DOI: 10.1080/09537104.2024.2337907 -
The EMBO Journal Aug 2023CDC14, originally identified as crucial mediator of mitotic exit in budding yeast, belongs to the family of dual-specificity phosphatases (DUSPs) that are present in... (Review)
Review
CDC14, originally identified as crucial mediator of mitotic exit in budding yeast, belongs to the family of dual-specificity phosphatases (DUSPs) that are present in most eukaryotes. Contradicting data have sparked a contentious discussion whether a cell cycle role is conserved in the human paralogs CDC14A and CDC14B but possibly masked due to redundancy. Subsequent studies on CDC14A and CDC14B double knockouts in human and mouse demonstrated that CDC14 activity is dispensable for mitotic progression in higher eukaryotes and instead suggested functional specialization. In this review, we provide a comprehensive overview of our current understanding of how faithful cell division is linked to phosphorylation and dephosphorylation and compare functional similarities and divergences between the mitotic phosphatases CDC14, PP2A, and PP1 from yeast and higher eukaryotes. Furthermore, we review the latest discoveries on CDC14B, which identify this nuclear phosphatase as a key regulator of gene expression and reveal its role in neuronal development. Finally, we discuss CDC14B functions in meiosis and possible implications in other developmental processes.
Topics: Humans; Animals; Mice; Saccharomyces cerevisiae; Protein Tyrosine Phosphatases; Cell Division; Cell Cycle; Dual-Specificity Phosphatases; Cell Cycle Proteins; Phosphorylation; Mitosis; Saccharomyces cerevisiae Proteins
PubMed: 37493185
DOI: 10.15252/embj.2023114364 -
Nature Sep 2023The presequence translocase of the mitochondrial inner membrane (TIM23) represents the major route for the import of nuclear-encoded proteins into mitochondria. About...
The presequence translocase of the mitochondrial inner membrane (TIM23) represents the major route for the import of nuclear-encoded proteins into mitochondria. About 60% of more than 1,000 different mitochondrial proteins are synthesized with amino-terminal targeting signals, termed presequences, which form positively charged amphiphilic α-helices. TIM23 sorts the presequence proteins into the inner membrane or matrix. Various views, including regulatory and coupling functions, have been reported on the essential TIM23 subunit Tim17 (refs. ). Here we mapped the interaction of Tim17 with matrix-targeted and inner membrane-sorted preproteins during translocation in the native membrane environment. We show that Tim17 contains conserved negative charges close to the intermembrane space side of the bilayer, which are essential to initiate presequence protein translocation along a distinct transmembrane cavity of Tim17 for both classes of preproteins. The amphiphilic character of mitochondrial presequences directly matches this Tim17-dependent translocation mechanism. This mechanism permits direct lateral release of transmembrane segments of inner membrane-sorted precursors into the inner membrane.
Topics: Mitochondria; Mitochondrial Membranes; Mitochondrial Precursor Protein Import Complex Proteins; Protein Transport; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins
PubMed: 37527780
DOI: 10.1038/s41586-023-06477-8 -
Thrombosis Research Oct 2023Thrombin, the enzyme which converts fibrinogen into a fibrin clot, is produced by the prothrombinase complex, composed of factor Xa (FXa) and factor Va (FVa)....
INTRODUCTION
Thrombin, the enzyme which converts fibrinogen into a fibrin clot, is produced by the prothrombinase complex, composed of factor Xa (FXa) and factor Va (FVa). Down-regulation of this process is critical, as excess thrombin can lead to life-threatening thrombotic events. FXa and FVa are inhibited by the anticoagulants tissue factor pathway inhibitor alpha (TFPIα) and activated protein C (APC), respectively, and their common cofactor protein S (PS). However, prothrombinase is resistant to either of these inhibitory systems in isolation.
MATERIALS AND METHODS
We hypothesized that these anticoagulants function best together, and tested this hypothesis using purified proteins and plasma-based systems.
RESULTS
In plasma, TFPIα had greater anticoagulant activity in the presence of APC and PS, maximum PS activity required both TFPIα and APC, and antibodies against TFPI and APC had an additive procoagulant effect, which was mimicked by an antibody against PS alone. In purified protein systems, TFPIα dose-dependently inhibited thrombin activation by prothrombinase, but only in the presence of APC, and this activity was enhanced by PS. Conversely, FXa protected FVa from cleavage by APC, even in the presence of PS, and TFPIα reversed this protection. However, prothrombinase assembled on platelets was still protected from inhibition, even in the presence of TFPIα, APC, and PS.
CONCLUSIONS
We propose a model of prothrombinase inhibition through combined targeting of both FXa and FVa, and that this mechanism enables down-regulation of thrombin activation outside of a platelet clot. Platelets protect prothrombinase from inhibition, however, supporting a procoagulant environment within the clot.
Topics: Humans; Anticoagulants; Blood Coagulation; Factor V; Factor Va; Factor Xa; Protein C; Protein S; Thrombin; Thromboplastin
PubMed: 37660436
DOI: 10.1016/j.thromres.2023.08.012 -
The EMBO Journal Dec 2023Ceramide synthases (CerS) catalyze ceramide formation via N-acylation of a sphingoid base with a fatty acyl-CoA and are attractive drug targets for treating numerous...
Ceramide synthases (CerS) catalyze ceramide formation via N-acylation of a sphingoid base with a fatty acyl-CoA and are attractive drug targets for treating numerous metabolic diseases and cancers. Here, we present the cryo-EM structure of a yeast CerS complex, consisting of a catalytic Lac1 subunit and a regulatory Lip1 subunit, in complex with C26-CoA substrate. The CerS holoenzyme exists as a dimer of Lac1-Lip1 heterodimers. Lac1 contains a hydrophilic reaction chamber and a hydrophobic tunnel for binding the CoA moiety and C26-acyl chain of C26-CoA, respectively. Lip1 interacts with both the transmembrane region and the last luminal loop of Lac1 to maintain the proper acyl chain binding tunnel. A lateral opening on Lac1 serves as a potential entrance for the sphingoid base substrate. Our findings provide a template for understanding the working mechanism of eukaryotic ceramide synthases and may facilitate the development of therapeutic CerS modulators.
Topics: Ceramides; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins; Oxidoreductases; Membrane Proteins
PubMed: 37953642
DOI: 10.15252/embj.2023114889 -
Journal of the American Society For... May 2024This paper sheds light on the meaning of hydrogen/deuterium exchange-mass spectrometry (HDX-MS) data. HDX-MS data provide not structural information but dynamic...
This paper sheds light on the meaning of hydrogen/deuterium exchange-mass spectrometry (HDX-MS) data. HDX-MS data provide not structural information but dynamic information on an analyte protein. First, the reaction mechanism of backbone amide HDX reaction is considered and the correlation between the parameters from an X-ray crystal structure and the protection factors of HDX reactions of cytochrome is evaluated. The presence of H-bonds in a protein structure has a strong influence on HDX rates which represent protein dynamics, while the solvent accessibility only weakly affects the HDX rates. Second, the energy diagrams of the HDX reaction at each residue in the presence and absence of perturbation are described. Whereas the free energy change upon mutation can be directly measured by the HDX rates, the free energy change upon ligand binding may be complicated due to the presence of unbound analyte protein in the protein-ligand mixture. Third, the meanings of HDX and other biophysical techniques are explained using a hypothetical protein folding well. The shape of the protein folding well describes the protein dynamics and provides Boltzmann distribution of open and closed states which yield HDX protection factors, while a protein's crystal structure represents a snapshot near the bottom of the well. All biophysical data should be consistent yet provide different information because they monitor different parts of the same protein folding well.
Topics: Crystallography, X-Ray; Cytochromes c; Deuterium Exchange Measurement; Hydrogen Bonding; Hydrogen Deuterium Exchange-Mass Spectrometry; Models, Molecular; Protein Conformation; Protein Folding; Proteins; Thermodynamics
PubMed: 38639434
DOI: 10.1021/jasms.4c00044