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Frontiers in Cell and Developmental... 2024Lipid droplets (LDs) serve as intracellular compartments primarily dedicated to the storage of metabolic energy in the form of neutral lipids. The processes that... (Review)
Review
Lipid droplets (LDs) serve as intracellular compartments primarily dedicated to the storage of metabolic energy in the form of neutral lipids. The processes that regulate and control LD biogenesis are being studied extensively and are gaining significance due to their implications in major metabolic disorders, including type 2 diabetes and obesity. A protein of particular interest is Fat storage-Inducing Transmembrane 2 (FIT2), which affects the emergence step of LD biogenesis. Instead of properly emerging towards the cytosol, LDs in FIT2-deficient cells remain embedded within the membrane of the endoplasmic reticulum (ER). studies revealed the ability of FIT2 to bind both di- and triacylglycerol (DAG/TAG), key players in lipid storage, and its activity to cleave acyl-CoA. However, the translation of these functions to the observed embedding of LDs in FIT2 deficient cells remains to be established. To understand the role of FIT2 , we discuss the parameters that affect LD emergence. Our focus centers on the role that membrane curvature and surface tension play in LD emergence, as well as the impact that the lipid composition exerts on these key parameters. In addition, we discuss hypotheses on how FIT2 could function locally to modulate lipids at sites of LD emergence.
PubMed: 38872930
DOI: 10.3389/fcell.2024.1422032 -
Scientific Reports Jun 2024Eukaryotic membranes are compartmentalized into distinct micro- and nanodomains that rearrange dynamically in response to external and internal cues. This lateral...
Eukaryotic membranes are compartmentalized into distinct micro- and nanodomains that rearrange dynamically in response to external and internal cues. This lateral heterogeneity of the lipid bilayer and associated clustering of distinct membrane proteins contribute to the spatial organization of numerous cellular processes. Here, we show that membrane microdomains within the endoplasmic reticulum (ER) of yeast cells are reorganized during metabolic reprogramming and aging. Using biosensors with varying transmembrane domain length to map lipid bilayer thickness, we demonstrate that in young cells, microdomains of increased thickness mainly exist within the nuclear ER, while progressing cellular age drives the formation of numerous microdomains specifically in the cortical ER. Partitioning of biosensors with long transmembrane domains into these microdomains increased protein stability and prevented autophagic removal. In contrast, reporters with short transmembrane domains progressively accumulated at the membrane contact site between the nuclear ER and the vacuole, the so-called nucleus-vacuole junction (NVJ), and were subjected to turnover via selective microautophagy occurring specifically at these sites. Reporters with long transmembrane domains were excluded from the NVJ. Our data reveal age-dependent rearrangement of the lateral organization of the ER and establish transmembrane domain length as a determinant of membrane contact site localization and autophagic degradation.
Topics: Endoplasmic Reticulum; Autophagy; Saccharomyces cerevisiae; Membrane Microdomains; Cellular Senescence; Saccharomyces cerevisiae Proteins; Vacuoles; Membrane Proteins
PubMed: 38871812
DOI: 10.1038/s41598-024-64493-8 -
Redox Biology Jun 2024Oxidative stress (OS) and endoplasmic reticulum stress (ERS) are at the genesis of placental disorders observed in preeclampsia, intrauterine growth restriction, and...
Manganese porphyrin-based treatment improves fetal-placental development and protects against oxidative damage and NLRP3 inflammasome activation in a rat maternal hypothyroidism model.
Oxidative stress (OS) and endoplasmic reticulum stress (ERS) are at the genesis of placental disorders observed in preeclampsia, intrauterine growth restriction, and maternal hypothyroidism. In this regard, cationic manganese porphyrins (MnPs) comprise potent redox-active therapeutics of high antioxidant and anti-inflammatory potential, which have not been evaluated in metabolic gestational diseases yet. This study evaluated the therapeutic potential of two MnPs, [MnTE-2-PyP] (MnP I) and [MnT(5-Br-3-E-Py)P]5+ (MnP II), in the fetal-placental dysfunction of hypothyroid rats. Hypothyroidism was induced by administration of 6-Propyl-2-thiouracil (PTU) and treatment with MnPs I and II 0.1 mg/kg/day started on the 8th day of gestation (DG). The fetal and placental development, and protein and/or mRNA expression of antioxidant mediators (SOD1, CAT, GPx1), hypoxia (HIF1α), oxidative damage (8-OHdG, MDA), ERS (GRP78 and CHOP), immunological (TNFα, IL-6, IL-10, IL-1β, IL-18, NLRP3, Caspase1, Gasdermin D) and angiogenic (VEGF) were evaluated in the placenta and decidua on the 18th DG using immunohistochemistry and qPCR. ROS and peroxynitrite (PRX) were quantified by fluorometric assay, while enzyme activities of SOD, GST, and catalase were evaluated by colorimetric assay. MnPs I and II increased fetal body mass in hypothyroid rats, and MnP I increased fetal organ mass. MnPs restored the junctional zone morphology in hypothyroid rats and increased placental vascularization. MnPs blocked the increase of OS and ERS mediators caused by hypothyroidism, showing similar levels of expression of HIFα, 8-OHdG, MDA, Gpx1, GRP78, and Chop to the control. Moreover, MnPs I and/or II increased the protein expression of SOD1, Cat, and GPx1 and restored the expression of IL10, Nlrp3, and Caspase1 in the decidua and/or placenta. However, MnPs did not restore the low placental enzyme activity of SOD, CAT, and GST caused by hypothyroidism, while increased the decidual and placental protein expression of TNFα. The results show that treatment with MnPs improves the fetal-placental development and the placental inflammatory state of hypothyroid rats and protects against oxidative stress and reticular stress caused by hypothyroidism at the maternal-fetal interface.
PubMed: 38870780
DOI: 10.1016/j.redox.2024.103238 -
ELife Jun 2024Compared with lowlander migrants, native Tibetans have a higher reproductive success at high altitude though the underlying mechanism remains unclear. Here, we compared...
Compared with lowlander migrants, native Tibetans have a higher reproductive success at high altitude though the underlying mechanism remains unclear. Here, we compared the transcriptome and histology of full-term placentas between native Tibetans and Han migrants. We found that the placental trophoblast shows the largest expression divergence between Tibetans and Han, and Tibetans show decreased immune response and endoplasmic reticulum stress. Remarkably, we detected a sex-biased expression divergence, where the male-infant placentas show a greater between-population difference than the female-infant placentas. The umbilical cord plays a key role in the sex-biased expression divergence, which is associated with the higher birth weight of the male newborns of Tibetans. We also identified adaptive histological changes in the male-infant placentas of Tibetans, including larger umbilical artery wall and umbilical artery intima and media, and fewer syncytial knots. These findings provide valuable insights into the sex-biased adaptation of human populations, with significant implications for medical and genetic studies of human reproduction.
Topics: Humans; Female; Placenta; Male; Pregnancy; Fetal Development; Tibet; Infant, Newborn; Transcriptome; Altitude; Sex Factors; Sex Characteristics
PubMed: 38869160
DOI: 10.7554/eLife.89004 -
BMC Biology Jun 2024Most tail-anchored (TA) membrane proteins are delivered to the endoplasmic reticulum through a conserved posttranslational pathway. Although core mechanisms underlying...
BACKGROUND
Most tail-anchored (TA) membrane proteins are delivered to the endoplasmic reticulum through a conserved posttranslational pathway. Although core mechanisms underlying the targeting and insertion of TA proteins are well established in eukaryotes, their role in mediating TA protein biogenesis in plants remains unclear. We reported the crystal structures of algal arsenite transporter 1 (ArsA1), which possesses an approximately 80-kDa monomeric architecture and carries chloroplast-localized TA proteins. However, the mechanistic basis of ArsA2, a Get3 (guided entry of TA proteins 3) homolog in plants, for TA recognition remains unknown.
RESULTS
Here, for the first time, we present the crystal structures of the diatom Pt-Get3a that forms a distinct ellipsoid-shaped tetramer in the open (nucleotide-bound) state through crystal packing. Pulldown assay results revealed that only tetrameric Pt-Get3a can bind to TA proteins. The lack of the conserved zinc-coordination CXXC motif in Pt-Get3a potentially leads to the spontaneous formation of a distinct parallelogram-shaped dimeric conformation in solution, suggesting a new dimer state for subsequent tetramerization upon TA targeting. Pt-Get3a nonspecifically binds to different subsets of TA substrates due to the lower hydrophobicity of its α-helical subdomain, which is implicated in TA recognition.
CONCLUSIONS
Our study provides new insights into the mechanisms underlying TA protein shielding by tetrameric Get3 during targeting to the diatom's cell membrane.
Topics: Diatoms; Membrane Proteins; Protein Multimerization
PubMed: 38867239
DOI: 10.1186/s12915-024-01933-x -
Biological & Pharmaceutical Bulletin 2024Ceramide (Cer) is synthesized de novo in the bilayer of the endoplasmic reticulum and transported to the cytosolic leaflet of the trans-Golgi apparatus for sphingomyelin...
Ceramide (Cer) is synthesized de novo in the bilayer of the endoplasmic reticulum and transported to the cytosolic leaflet of the trans-Golgi apparatus for sphingomyelin (SM) synthesis. As the active site of SM synthase (SMS) is located on the luminal side of the Golgi membrane, Cer translocates to the lumen via transbilayer movement for SM synthesis. However, the mechanism of transbilayer movement is not fully understood. As the Cer-related translocases seem to localize near the SMS, the protein was identified using proximity-dependent biotin identification proteomics. Phospholipid scramblase 1 (PLSCR1), which is thought to act as a scramblase for phosphatidylserine and phosphatidylethanolamine, was identified as a protein proximal to the SMS isoforms SMS1 and SMS2. Although five isoforms of PLSCR have been reported in humans, only PLSCR1, PLSCR3, and PLSCR4 are expressed in HEK293T cells. Confocal microscopic analysis showed that PLSCR1 and PLSCR4 partially co-localized with p230, a trans-Golgi network marker, where SMS isoforms are localized. We established CRISPR/Cas9-mediated PLSCR1, PLSCR3, and PLSCR4 single-knockout cells and PLSCR1, 3, 4 triple knockout HEK293T cells. Liquid chromatography-tandem mass spectrometry revealed that the levels of species with distinct acyl chains in Cer and SM were not significantly different in single knockout cells or in the triple knockout cells compared to the wild-type cells. Our findings suggest that PLSCR1 is localized in the vicinity of SMS isoforms, however is not involved in the transbilayer movement of Cer for SM synthesis.
Topics: Humans; Phospholipid Transfer Proteins; Transferases (Other Substituted Phosphate Groups); HEK293 Cells; Sphingomyelins; Membrane Proteins; Isoenzymes; Golgi Apparatus
PubMed: 38866522
DOI: 10.1248/bpb.b24-00177 -
Life Science Alliance Aug 2024Multispanning membrane proteins are inserted into the endoplasmic reticulum membrane by the ribosome-bound multipass translocon (MPT) machinery. Based on cryo-electron...
Multispanning membrane proteins are inserted into the endoplasmic reticulum membrane by the ribosome-bound multipass translocon (MPT) machinery. Based on cryo-electron tomography and extensive subtomogram analysis, we reveal the composition and arrangement of ribosome-bound MPT components in their native membrane environment. The intramembrane chaperone complex PAT and the translocon-associated protein (TRAP) complex associate substoichiometrically with the MPT in a translation-dependent manner. Although PAT is preferentially part of MPTs bound to translating ribosomes, the abundance of TRAP is highest in MPTs associated with non-translating ribosomes. The subtomogram average of the TRAP-containing MPT reveals intermolecular contacts between the luminal domains of TRAP and an unknown subunit of the back-of-Sec61 complex. AlphaFold modeling suggests this protein is nodal modulator, bridging the luminal domains of nicalin and TRAPα. Collectively, our results visualize the variability of MPT factors in the native membrane environment dependent on the translational activity of the bound ribosome.
Topics: Ribosomes; Membrane Proteins; Endoplasmic Reticulum; Protein Biosynthesis; Cryoelectron Microscopy; SEC Translocation Channels; Molecular Chaperones; Protein Transport; Models, Molecular
PubMed: 38866426
DOI: 10.26508/lsa.202302496 -
Journal of Lipid Research Jun 2024Hypercholesterolemia is frequently intertwined with hepatosteatosis, hypertriglyceridemia, and hyperglycemia. This study is designed to assess the therapeutic efficacy...
Hypercholesterolemia is frequently intertwined with hepatosteatosis, hypertriglyceridemia, and hyperglycemia. This study is designed to assess the therapeutic efficacy of miR-206 in contrast to statins in the context of managing hypercholesterolemia in mice. We previously showed that miR-206 is a potent inhibitor of de novo lipogenesis (DNL), cholesterol synthesis and gluconeogenesis in mice. Given that these processes occur within hepatocytes, we employed a mini-circle (MC) system to deliver miR-206 specifically to hepatocytes (designated as MC-miR-206). A single intravenous injection of MC-miR-206 maintained high levels of miR-206 in the liver for at least two weeks, thereby maintaining suppression of hepatic DNL, cholesterol synthesis and gluconeogenesis. MC-miR-206 significantly reduced DNA damage, endoplasmic reticulum and oxidative stress, and hepatic toxicity. Therapeutically, both MC-miR-206 and statins significantly reduced total serum cholesterol and triglycerides as well as LDL cholesterol and VLDL cholesterol in mice maintained on the normal chow and high-fat high-cholesterol diet. MC-miR-206 reduced liver weight, hepatic triglycerides and cholesterol and blood glucose, while statins slightly increased hepatic cholesterol and blood glucose and failed to affect levels of liver weight and hepatic triglycerides. Mechanistically, miR-206 alleviated hypercholesterolemia by inhibiting hepatic cholesterol synthesis, while statins increased HMGCR activity, hepatic cholesterol synthesis and fecal neutral steroid excretion. CONCLUSIONS: MiR-206 facilitates the regression of hypercholesterolemia, hypertriglyceridemia, hyperglycemia, and hepatosteatosis. MiR-206 outperforms statins by reducing hyperglycemia, hepatic cholesterol levels, and hepatic toxicity.
PubMed: 38866328
DOI: 10.1016/j.jlr.2024.100576 -
Proceedings of the National Academy of... Jun 2024The heart beats approximately 100,000 times per day in humans, imposing substantial energetic demands on cardiac muscle. Adenosine triphosphate (ATP) is an essential...
The heart beats approximately 100,000 times per day in humans, imposing substantial energetic demands on cardiac muscle. Adenosine triphosphate (ATP) is an essential energy source for normal function of cardiac muscle during each beat, as it powers ion transport, intracellular Ca handling, and actin-myosin cross-bridge cycling. Despite this, the impact of excitation-contraction coupling on the intracellular ATP concentration ([ATP]) in myocytes is poorly understood. Here, we conducted real-time measurements of [ATP] in ventricular myocytes using a genetically encoded ATP fluorescent reporter. Our data reveal rapid beat-to-beat variations in [ATP]. Notably, diastolic [ATP] was <1 mM, which is eightfold to 10-fold lower than previously estimated. Accordingly, ATP-sensitive K (K) channels were active at physiological [ATP]. Cells exhibited two distinct types of ATP fluctuations during an action potential: net increases (Mode 1) or decreases (Mode 2) in [ATP]. Mode 1 [ATP] increases necessitated Ca entry and release from the sarcoplasmic reticulum (SR) and were associated with increases in mitochondrial Ca. By contrast, decreases in mitochondrial Ca accompanied Mode 2 [ATP] decreases. Down-regulation of the protein mitofusin 2 reduced the magnitude of [ATP] fluctuations, indicating that SR-mitochondrial coupling plays a crucial role in the dynamic control of ATP levels. Activation of β-adrenergic receptors decreased [ATP], underscoring the energetic impact of this signaling pathway. Finally, our work suggests that cross-bridge cycling is the largest consumer of ATP in a ventricular myocyte during an action potential. These findings provide insights into the energetic demands of EC coupling and highlight the dynamic nature of ATP concentrations in cardiac muscle.
Topics: Myocytes, Cardiac; Adenosine Triphosphate; Excitation Contraction Coupling; Animals; Calcium; Heart Ventricles; Action Potentials; Sarcoplasmic Reticulum; Heart Rate; Humans; KATP Channels; Myocardial Contraction; Mice
PubMed: 38865270
DOI: 10.1073/pnas.2318535121 -
Cell Reports Jun 2024Despite the consensus that accumulation of unfolded proteins in the endoplasmic reticulum (ER) lumen, i.e. ER stress, activates the unfolded protein response (UPR),...
Despite the consensus that accumulation of unfolded proteins in the endoplasmic reticulum (ER) lumen, i.e. ER stress, activates the unfolded protein response (UPR), studies under physiological and pathophysiological conditions suggest that ER stress may not always trigger the UPR, and the UPR can be activated in an ER stress-independent way. To better understand how the UPR is regulated and its relationship with ER stress requires direct detection of unfolded proteins in the ER, a method that is still lacking. Here, we report a strategy of visualizing unfolded protein accumulation in the ER lumen in living cells by employing an engineered ER stress sensor, PERK, which forms fluorescence puncta upon unfolded protein binding, in a fast and reversible way. Our reporter enables us to clarify the involvement of unfolded proteins in UPR activation under several physiological conditions and suggests that persistent unfolded protein accumulation in the ER despite UPR attenuation predicts cell death.
PubMed: 38865243
DOI: 10.1016/j.celrep.2024.114358