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Biomolecules Jan 2024The purpose of this review is to succinctly examine the methodologies used in lipid raft research in the brain and to highlight the drawbacks of some investigative... (Review)
Review
The purpose of this review is to succinctly examine the methodologies used in lipid raft research in the brain and to highlight the drawbacks of some investigative approaches. Lipid rafts are biochemically and biophysically different from the bulk membrane. A specific lipid environment within membrane domains provides a harbor for distinct raftophilic proteins, all of which in concert create a specialized platform orchestrating various cellular processes. Studying lipid rafts has proved to be arduous due to their elusive nature, mobility, and constant dynamic reorganization to meet the cellular needs. Studying neuronal lipid rafts is particularly cumbersome due to the immensely complex regional molecular architecture of the central nervous system. Biochemical fractionation, performed with or without detergents, is still the most widely used method to isolate lipid rafts. However, the differences in solubilization when various detergents are used has exposed a dire need to find more reliable methods to study particular rafts. Biochemical methods need to be complemented with other approaches such as live-cell microscopy, imaging mass spectrometry, and the development of specific non-invasive fluorescent probes to obtain a more complete image of raft dynamics and to study the spatio-temporal expression of rafts in live cells.
Topics: Detergents; Membrane Microdomains; Brain
PubMed: 38397393
DOI: 10.3390/biom14020156 -
Archives of Biochemistry and Biophysics Apr 2024G-protein-coupled receptors (GPCRs) are the largest family of membrane proteins, regulate a plethora of physiological responses and are the therapeutic target for 30-40%... (Review)
Review
G-protein-coupled receptors (GPCRs) are the largest family of membrane proteins, regulate a plethora of physiological responses and are the therapeutic target for 30-40% of clinically-prescribed drugs. They are integral membrane proteins deeply embedded in the plasma membrane where they activate intracellular signalling via coupling to G-proteins and β-arrestin. GPCRs are in intimate association with the bilayer lipids and that lipid environment regulates the signalling functions of GPCRs. This complex lipid 'landscape' is both heterogeneous and dynamic. GPCR function is modulated by bulk membrane properties including membrane fluidity, microdomains, curvature, thickness and asymmetry but GPCRs are also regulated by specific lipid:GPCR binding, including cholesterol and anionic lipids. Understanding the molecular mechanisms whereby GPCR signalling is regulated by lipids is a very active area of research currently. A major advance in membrane protein research in recent years was the application of poly(styrene-co-maleic acid) (SMA) copolymers. These spontaneously generate SMA lipid particles (SMALPs) encapsulating membrane protein in a nano-scale disc of cell membrane, thereby removing the historical need for detergent and preserving lipid:GPCR interaction. The focus of this review is how GPCR-SMALPs are increasing our understanding of GPCR structure and function at the molecular level. Furthermore, an increasing number of 'second generation' SMA-like copolymers have been reported recently. These are reviewed from the context of increasing our understanding of GPCR molecular mechanisms. Moreover, their potential as a novel platform for downstream biophysical and structural analyses is assessed and looking ahead, the translational application of SMA-like copolymers to GPCR drug discovery programmes in the future is considered.
Topics: Cell Membrane; Lipids; Membrane Proteins; Receptors, G-Protein-Coupled
PubMed: 38395122
DOI: 10.1016/j.abb.2024.109946 -
Frontiers in Cell and Developmental... 2024Digestive system malignancies, including cancers of the esophagus, pancreas, stomach, liver, and colorectum, are the leading causes of cancer-related deaths worldwide... (Review)
Review
Digestive system malignancies, including cancers of the esophagus, pancreas, stomach, liver, and colorectum, are the leading causes of cancer-related deaths worldwide due to their high morbidity and poor prognosis. The lack of effective early diagnosis methods is a significant factor contributing to the poor prognosis for these malignancies. Tetraspanins (Tspans) are a superfamily of 4-transmembrane proteins (TM4SF), classified as low-molecular-weight glycoproteins, with 33 Tspan family members identified in humans to date. They interact with other membrane proteins or TM4SF members to form a functional platform on the cytoplasmic membrane called Tspan-enriched microdomain and serve multiple functions including cell adhesion, migration, propagation and signal transduction. In this review, we summarize the various roles of Tspans in the progression of digestive system tumors and the underlying molecular mechanisms in recent years. Generally, the expression of CD9, CD151, Tspan1, Tspan5, Tspan8, Tspan12, Tspan15, and Tspan31 are upregulated, facilitating the migration and invasion of digestive system cancer cells. Conversely, Tspan7, CD82, CD63, Tspan7, and Tspan9 are downregulated, suppressing digestive system tumor cell metastasis. Furthermore, the connection between Tspans and the metastasis of malignant bone tumors is reviewed. We also summarize the potential role of Tspans as novel immunotherapy targets and as an approach to overcome drug resistance. Finally, we discuss the potential clinical value and therapeutic targets of Tspans in the treatments of digestive system malignancies and provide some guidance for future research.
PubMed: 38389703
DOI: 10.3389/fcell.2024.1343894 -
MBio Mar 2024The cellular junctional architecture remodeling by adhesion protein-heat shock protein 60 (LAP-Hsp60) interaction for () passage through the epithelial barrier is...
UNLABELLED
The cellular junctional architecture remodeling by adhesion protein-heat shock protein 60 (LAP-Hsp60) interaction for () passage through the epithelial barrier is incompletely understood. Here, using the gerbil model, permissive to internalin (Inl) A/B-mediated pathways like in humans, we demonstrate that crosses the intestinal villi at 48 h post-infection. In contrast, the single isogenic ( or Δ) or double (Δ) mutant strains show significant defects. LAP promotes translocation via endocytosis of cell-cell junctional complex in enterocytes that do not display luminal E-cadherin. In comparison, InlA facilitates translocation at cells displaying apical E-cadherin during cell extrusion and mucus expulsion from goblet cells. LAP hijacks caveolar endocytosis to traffic integral junctional proteins to the early and recycling endosomes. Pharmacological inhibition in a cell line and genetic knockout of caveolin-1 in mice prevents LAP-induced intestinal permeability, junctional endocytosis, and translocation. Furthermore, LAP-Hsp60-dependent tight junction remodeling is also necessary for InlA access to E-cadherin for intestinal barrier crossing in InlA-permissive hosts.
IMPORTANCE
() is a foodborne pathogen with high mortality (20%-30%) and hospitalization rates (94%), particularly affecting vulnerable groups such as pregnant women, fetuses, newborns, seniors, and immunocompromised individuals. Invasive listeriosis involves 's internalin (InlA) protein binding to E-cadherin to breach the intestinal barrier. However, non-functional InlA variants have been identified in isolates, suggesting InlA-independent pathways for translocation. Our study reveals that adhesion protein (LAP) and InlA cooperatively assist entry into the gut lamina propria in a gerbil model, mimicking human listeriosis in early infection stages. LAP triggers caveolin-1-mediated endocytosis of critical junctional proteins, transporting them to early and recycling endosomes, facilitating passage through enterocytes. Furthermore, LAP-Hsp60-mediated junctional protein endocytosis precedes InlA's interaction with basolateral E-cadherin, emphasizing LAP and InlA's cooperation in enhancing intestinal translocation. This understanding is vital in combating the severe consequences of infection, including sepsis, meningitis, encephalitis, and brain abscess.
Topics: Infant, Newborn; Female; Mice; Pregnancy; Humans; Animals; Listeria monocytogenes; Listeria; Caveolin 1; Caveolae; Gerbillinae; Bacterial Proteins; Listeriosis; Cadherins
PubMed: 38376160
DOI: 10.1128/mbio.02821-23 -
The Journal of Cell Biology May 2024Different membrane microdomain compositions provide unique environments that can regulate signaling receptor function. We identify microdomains on the endosome membrane...
Different membrane microdomain compositions provide unique environments that can regulate signaling receptor function. We identify microdomains on the endosome membrane of Drosophila endosomes, enriched in lipid-raft or clathrin/ESCRT-0, which are associated with Notch activation by distinct, ligand-independent mechanisms. Transfer of Notch between microdomains is regulated by Deltex and Suppressor of deltex ubiquitin ligases and is limited by a gate-keeper role for ESCRT complexes. Ubiquitination of Notch by Deltex recruits it to the clathrin/ESCRT-0 microdomain and enhances Notch activation by an ADAM10-independent/TRPML-dependent mechanism. This requirement for Deltex is bypassed by the downregulation of ESCRT-III. In contrast, while ESCRT-I depletion also activates Notch, it does so by an ADAM10-dependent/TRPML-independent mechanism and Notch is retained in the lipid raft-like microdomain. In the absence of such endosomal perturbation, different activating Notch mutations also localize to different microdomains and are activated by different mechanisms. Our findings demonstrate the interplay between Notch regulators, endosomal trafficking components, and Notch genetics, which defines membrane locations and activation mechanisms.
Topics: Animals; ADAM10 Protein; Clathrin; Down-Regulation; Drosophila; Drosophila Proteins; Endosomal Sorting Complexes Required for Transport; Endosomes; Transient Receptor Potential Channels; Receptors, Notch; Ubiquitination; Membrane Proteins; Membrane Microdomains
PubMed: 38358349
DOI: 10.1083/jcb.202211041 -
Pharmacological Research Mar 2024The uncontrolled bacterial infection-induced cytokine storm and sequential immunosuppression are commonly observed in septic patients, which indicates that the...
The uncontrolled bacterial infection-induced cytokine storm and sequential immunosuppression are commonly observed in septic patients, which indicates that the activation of phagocytic cells and the efficient and timely elimination of bacteria are crucial for combating bacterial infections. However, the role of dysregulated immune cells and their disrupted function in sepsis remains unclear. Here, we found that macrophages exhibited the impaired endocytosis capabilities in sepsis by Single-cell RNA sequencing and bulk RNA sequencing. Caveolae protein Caveolin-1 (Cav-1) of macrophages was inactivated by SHP2 rapidly during Escherichia coli (E.coli) infection. Allosteric inhibitor of SHP2 effectively maintains Cav-1 phosphorylation to enhance macrophage to endocytose and eliminate bacteria. Additionally, TLR4 endocytosis of macrophage was also enhanced upon E.coli infection by SHP099, inducing an increased and rapidly resolved inflammatory response. In vivo, pretreatment or posttreatment with inhibitor of SHP2 significantly reduced the bacterial burden in organs and mortality of mice subjected E.coli infection or CLP-induced sepsis. The cotreatment of inhibitor of SHP2 with an antibiotic conferred complete protection against mortality in mice. Our findings suggest that Cav-1-mediated endocytosis and bacterial elimination may play a critical role in the pathogenesis of sepsis, highlighting inhibitor of SHP2 as a potential therapeutic agent for sepsis.
Topics: Animals; Humans; Mice; Bacteria; Caveolae; Endocytosis; Escherichia coli Infections; Macrophages; Protein Tyrosine Phosphatase, Non-Receptor Type 11; Sepsis
PubMed: 38320736
DOI: 10.1016/j.phrs.2024.107096 -
MSphere Feb 2024Noelia Lander works on cell signaling in American trypanosomes and studies the role of cyclic adenosine monophosphate (cAMP) microdomains in environmental sensing and...
Noelia Lander works on cell signaling in American trypanosomes and studies the role of cyclic adenosine monophosphate (cAMP) microdomains in environmental sensing and differentiation. In this mSphere of Influence, Dr. Lander reflects on three research articles in different eukaryotic models that had impacted on the way she thinks about the regulation of cAMP signals in , the etiologic agent of Chagas disease. The articles "FRET biosensor uncovers cAMP nano-domains at β-adrenergic targets that dictate precise tuning of cardiac contractility" (N. C. Surdo, M. Berrera, A. Koschinski, M. Brescia, et al., Nat Commun 8:15031, 2017, https://doi.org/10.1038/ncomms15031), "Cyclic AMP signaling and glucose metabolism mediate pH taxis by African trypanosomes" (S. Shaw, S. Knüsel, D. Abbühl, A. Naguleswaran, et al., Nat Commun 13:603, 2022, https://doi.org/10.1038/s41467-022-28293-w), and "Encystation stimuli sensing is mediated by adenylate cyclase AC2-dependent cAMP signaling in " (H. W. Shih, G. C. M. Alas, and A. R. Paredez, Nat Commun 14:7245, 2023, https://doi.org/10.1038/s41467-023-43028-1) influenced her current hypothesis that cAMP signals are generated in response to environmental cues leading to changes in membrane fluidity at the flagellar tip and the contractile vacuole complex of , structures where cAMP mediates key cellular processes for developmental progression.
Topics: Female; United States; Humans; Trypanosoma cruzi; Cyclic AMP
PubMed: 38315033
DOI: 10.1128/msphere.00635-23 -
Experimental & Molecular Medicine Feb 2024Autophagy is an essential quality control mechanism for maintaining organellar functions in eukaryotic cells. Defective autophagy in pancreatic beta cells has been shown... (Review)
Review
Autophagy is an essential quality control mechanism for maintaining organellar functions in eukaryotic cells. Defective autophagy in pancreatic beta cells has been shown to be involved in the progression of diabetes through impaired insulin secretion under glucolipotoxic stress. The underlying mechanism reveals the pathologic role of the hyperactivation of mechanistic target of rapamycin (mTOR), which inhibits lysosomal biogenesis and autophagic processes. Moreover, accumulating evidence suggests that oxidative stress induces Ca depletion in the endoplasmic reticulum (ER) and cytosolic Ca overload, which may contribute to mTOR activation in perilysosomal microdomains, leading to autophagic defects and β-cell failure due to lipotoxicity. This review delineates the antagonistic regulation of autophagic flux by mTOR and AMP-dependent protein kinase (AMPK) at the lysosomal membrane, and both of these molecules could be activated by perilysosomal calcium signaling. However, aberrant and persistent Ca elevation upon lipotoxic stress increases mTOR activity and suppresses autophagy. Therefore, normalization of autophagy is an attractive therapeutic strategy for patients with β-cell failure and diabetes.
Topics: Humans; Calcium; Insulin-Secreting Cells; Adenylate Kinase; Autophagy; Diabetes Mellitus; TOR Serine-Threonine Kinases
PubMed: 38297165
DOI: 10.1038/s12276-024-01161-x -
Journal of Molecular Medicine (Berlin,... Mar 2024Amyotrophic lateral sclerosis (ALS) is an age-dependent neurodegenerative disease affecting motor neurons in the spinal cord and brainstem whose etiopathogenesis remains...
Amyotrophic lateral sclerosis (ALS) is an age-dependent neurodegenerative disease affecting motor neurons in the spinal cord and brainstem whose etiopathogenesis remains unclear. Recent studies have linked major neurodegenerative diseases with altered function of multimolecular lipid-protein complexes named lipid rafts. In the present study, we have isolated lipid rafts from the anterior horn of the spinal cords of controls and ALS individuals and analysed their lipid composition. We found that ALS affects levels of different fatty acids, lipid classes and related ratios and indexes. The most significant changes affected the contents of n-9/n-7 monounsaturated fatty acids and arachidonic acid, the main n-6 long-chain polyunsaturated fatty acid (LCPUFA), which were higher in ALS lipid rafts. Paralleling these findings, ALS lipid rafts lower saturates-to-unsaturates ratio compared to controls. Further, levels of cholesteryl ester (SE) and anionic-to-zwitterionic phospholipids ratio were augmented in ALS lipid rafts, while sulfatide contents were reduced. Further, regression analyses revealed augmented SE esterification to (mono)unsaturated fatty acids in ALS, but to saturates in controls. Overall, these changes indicate that lipid rafts from ALS spinal cord undergo destabilization of the lipid structure, which might impact their biophysical properties, likely leading to more fluid membranes. Indeed, estimations of membrane microviscosity confirmed less viscous membranes in ALS, as well as more mobile yet smaller lipid rafts compared to surrounding membranes. Overall, these results demonstrate that the changes in ALS lipid rafts are unrelated to oxidative stress, but to anomalies in lipid metabolism and/or lipid raft membrane biogenesis in motor neurons. KEY MESSAGES: The lipid matrix of multimolecular membrane complexes named lipid rafts are altered in human spinal cord in sporadic amyotrophic lateral sclerosis (ALS). Lipid rafts from ALS spinal cord contain higher levels of n-6 LCPUFA (but not n-3 LCPUFA), n-7/n-9 monounsaturates and lower saturates-to-unsaturates ratio. ALS lipid rafts display increased contents of cholesteryl esters, anomalous anionic-to-zwitterionic phospholipids and phospholipid remodelling and reduced sulphated and total sphingolipid levels, compared to control lipid rafts. Destabilization of the lipid structure of lipid raft affects their biophysical properties and leads to more fluid, less viscous membrane microdomains. The changes in ALS lipid rafts are unlikely related to increased oxidative stress, but to anomalies in lipid metabolism and/or raft membrane biogenesis in motor neurons.
Topics: Humans; Amyotrophic Lateral Sclerosis; Neurodegenerative Diseases; Lipids; Membrane Microdomains
PubMed: 38285093
DOI: 10.1007/s00109-024-02419-7 -
Cells Jan 2024Tetraspanins, a superfamily of small integral membrane proteins, are characterized by four transmembrane domains and conserved protein motifs that are configured into a... (Review)
Review
Tetraspanins, a superfamily of small integral membrane proteins, are characterized by four transmembrane domains and conserved protein motifs that are configured into a unique molecular topology and structure in the plasma membrane. They act as key organizers of the plasma membrane, orchestrating the formation of specialized microdomains called "tetraspanin-enriched microdomains (TEMs)" or "tetraspanin nanodomains" that are essential for mediating diverse biological processes. TSPAN8 is one of the earliest identified tetraspanin members. It is known to interact with a wide range of molecular partners in different cellular contexts and regulate diverse molecular and cellular events at the plasma membrane, including cell adhesion, migration, invasion, signal transduction, and exosome biogenesis. The functions of cell-surface TSPAN8 are governed by ER targeting, modifications at the Golgi apparatus and dynamic trafficking. Intriguingly, limited evidence shows that TSPAN8 can translocate to the nucleus to act as a transcriptional regulator. The transcription of is tightly regulated and restricted to defined cell lineages, where it can serve as a molecular marker of stem/progenitor cells in certain normal tissues as well as tumors. Importantly, the oncogenic roles of TSPAN8 in tumor development and cancer metastasis have gained prominence in recent decades. Here, we comprehensively review the current knowledge on the molecular characteristics and regulatory mechanisms defining TSPAN8 functions, and discuss the potential and significance of TSPAN8 as a biomarker and therapeutic target across various epithelial cancers.
Topics: Humans; Tetraspanins; Neoplasms; Membrane Proteins; Cell Membrane; Cell Adhesion
PubMed: 38275818
DOI: 10.3390/cells13020193