Metabolic Depletion of Sphingolipids Reduces Cell Surface Population of the Human Serotonin<sub>1A</sub> Receptor due to Impaired Trafficking

Kumar, Abhishek; Sarkar, Parijat; Chattopadhyay, Amitabha

doi:10.1021/acschemneuro.1c00017

Cited by 6 publications

(12 citation statements)

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“…Use of a mycotoxin known to reduce sphingolipid content reduces the amount of cell surface localised 5HT-1A receptor [111], in line with earlier results indicating the same treatment reduced specific ligand binding and associated downstream cAMP signalling for this receptor [48]. In contrast, similar studies on the angiotensin II type 1A receptor and the bitter taste receptor, T2R14, indicated no change in receptor signalling as a result of sphingomyelin depletion [112].…”

Section: Gpcrs and Sphingolipids And Glycolipidssupporting

confidence: 79%

Insights into the Role of Membrane Lipids in the Structure, Function and Regulation of Integral Membrane Proteins

Renard

Byrne

2021

IJMS

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Membrane proteins exist within the highly hydrophobic membranes surrounding cells and organelles, playing key roles in cellular function. It is becoming increasingly clear that the membrane does not just act as an appropriate environment for these proteins, but that the lipids that make up these membranes are essential for membrane protein structure and function. Recent technological advances in cryogenic electron microscopy and in advanced mass spectrometry methods, as well as the development of alternative membrane mimetic systems, have allowed experimental study of membrane protein–lipid complexes. These have been complemented by computational approaches, exploiting the ability of Molecular Dynamics simulations to allow exploration of membrane protein conformational changes in membranes with a defined lipid content. These studies have revealed the importance of lipids in stabilising the oligomeric forms of membrane proteins, mediating protein–protein interactions, maintaining a specific conformational state of a membrane protein and activity. Here we review some of the key recent advances in the field of membrane protein–lipid studies, with major emphasis on respiratory complexes, transporters, channels and G-protein coupled receptors.

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Section: Gpcrs and Sphingolipids And Glycolipidssupporting

confidence: 79%

Insights into the Role of Membrane Lipids in the Structure, Function and Regulation of Integral Membrane Proteins

Renard

Byrne

2021

IJMS

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“…We previously showed that the N‐terminal myc‐tagged serotonin 1A receptors expressed in HEK‐293 cells retain all characteristics of the native serotonin 1A receptor in terms of ligand binding, G‐protein coupling and cellular signaling 43,61 . To monitor endocytosis of the serotonin 1A receptor in HEK‐5‐HT 1A R cells, we used a quantitative flow cytometric assay, previously developed by us 43–46,54 . This assay allows quantitative measurement of receptor endocytosis by quantifying the cell surface population of serotoinin 1A receptors from a large number of cells using an anti‐myc antibody conjugated to Alexa Fluor 488 against the N‐terminal myc‐tag of the receptor.…”

Section: Resultsmentioning

confidence: 99%

Metabolic depletion of sphingolipids inhibits agonist‐induced endocytosis of the serotonin_1Areceptor

Kumar

Sarkar

Chattopadhyay

2022

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G protein-coupled receptors (GPCRs) are vital cellular signaling machinery and currently represent $40% drug targets. Endocytosis of GPCRs is an important process that allows stringent spatiotemporal control over receptor population on the cell surface. Although the role of proteins in GPCR endocytosis is well addressed, the contribution of membrane lipids in this process is rather unexplored. Sphingolipids are essential functional lipids in higher eukaryotes and are implicated in several neurological functions. To understand the role of sphingolipids in GPCR endocytosis, we subjected cells expressing human serotonin 1A receptors (an important neurotransmitter GPCR involved in cognitive and behavioral functions) to metabolic sphingolipid depletion using fumonisin B 1 , an inhibitor of sphingolipid biosynthetic pathway. Our results, using flow cytometric analysis and confocal microscopic imaging, show that sphingolipid depletion inhibits agonist-induced endocytosis of the serotonin 1A receptor in a concentration-dependent manner, which was restored when sphingolipid levels were replenished. We further show that there was no change in the internalization of transferrin, a marker for clathrin-mediated endocytosis, under sphingolipiddepleted condition, highlighting the specific requirement of sphingolipids for endocytosis of serotonin 1A receptors. Our results reveal the regulatory role of sphingolipids in GPCR endocytosis and highlight the importance of neurotransmitter receptor trafficking in health and disease.endocytosis, fumonisin B 1 , G protein-coupled receptor, serotonin 1A receptor, sphingolipids, sphingomyelin | INTRODUCTIONG protein-coupled receptors (GPCRs) represent the largest class of integral membrane proteins and have a characteristic seven transmembrane domain architecture involved in signal transduction across the plasma membrane. [1][2][3][4] Because of their versatile role in multiple physiological functions, GPCRs are major drug targets across all clinical areas. [4][5][6][7] The intracellular signaling responses mediated by GPCRs are initiated at the plasma membrane and the downstream signaling cascades are spatiotemporally regulated in a stringent manner.In this context, endocytosis offers a key desensitization mechanism, by internalization of the receptor from the plasma membrane into the cellular interior, to restrict GPCR signaling responses in a strict spatiotemporal regime. [8][9][10] Emerging evidences have suggested that endocytosis of GPCRs could simultaneously facilitate non-canonical receptor-mediated signaling even from the intracellular receptor pool. [11][12][13][14] Endocytosis is concerted by a number of membraneassociated and cytoplasmic proteins that help in cargo assembly, Abbreviations: BCA, bicinchoninic acid; FB 1 , fumonisin B 1 ; FCS, fetal calf serum; GPCR, G protein-coupled receptor; HEK-5-HT 1A R, HEK-293 cells stably expressing N-terminal myctagged human serotonin 1A receptors; MTT, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide; PBS, phosphate buffered...

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“…These results highlight the complexity involved in these two processes and emphasize the relevance of the actual methodology used to deplete cholesterol (for more details, see Sarkar et al, 2017 ; Kumar and Chattopadhyay, 2021a ). In addition, we showed that inhibition of sphingolipid biosynthesis could affect trafficking of the serotonin 1A receptor ( Kumar et al, 2021 ).…”

Section: Endocytosis and Cellular Trafficking: New Paradigm In Slos R...mentioning

confidence: 99%

Smith-Lemli-Opitz syndrome: A pathophysiological manifestation of the Bloch hypothesis

Chattopadhyay

Sharma²

2023

Front. Mol. Biosci.

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The biosynthesis of cholesterol, an essential component of higher eukaryotic membranes, was worked out by Konrad Bloch (and Feodor Lynen) in the 1960s and they received the Nobel Prize around that time in recognition of their pioneering contributions. An elegant consequence of this was a hypothesis proposed by Konrad Bloch (the Bloch hypothesis) which suggests that each subsequent intermediate in the cholesterol biosynthesis pathway is superior in supporting membrane function in higher eukaryotes relative to its precursor. In this review, we discuss an autosomal recessive metabolic disorder, known as Smith-Lemli-Opitz syndrome (SLOS), associated with a defect in the Kandutsch-Russell pathway of cholesterol biosynthesis that results in accumulation of the immediate precursor of cholesterol in its biosynthetic pathway (7-dehydrocholesterol) and an altered cholesterol to total sterol ratio. Patients suffering from SLOS have several developmental, behavioral and cognitive abnormalities for which no drug is available yet. We characterize SLOS as a manifestation of the Bloch hypothesis and review its molecular etiology and current treatment. We further discuss defective Hedgehog signaling in SLOS and focus on the role of the serotonin1A receptor, a representative neurotransmitter receptor belonging to the GPCR family, in SLOS. Notably, ligand binding activity and cellular signaling of serotonin1A receptors are impaired in SLOS-like condition. Importantly, cellular localization and intracellular trafficking of the serotonin1A receptor (which constitute an important determinant of a GPCR cellular function) are compromised in SLOS. We highlight some of the recent developments and emerging concepts in SLOS pathobiology and suggest that novel therapies based on trafficking defects of target receptors could provide new insight into treatment of SLOS.

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Metabolic Depletion of Sphingolipids Reduces Cell Surface Population of the Human Serotonin_1A Receptor due to Impaired Trafficking

Cited by 6 publications

References 72 publications

Insights into the Role of Membrane Lipids in the Structure, Function and Regulation of Integral Membrane Proteins

Insights into the Role of Membrane Lipids in the Structure, Function and Regulation of Integral Membrane Proteins

Metabolic depletion of sphingolipids inhibits agonist‐induced endocytosis of the serotonin_1Areceptor

Smith-Lemli-Opitz syndrome: A pathophysiological manifestation of the Bloch hypothesis

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Metabolic Depletion of Sphingolipids Reduces Cell Surface Population of the Human Serotonin1A Receptor due to Impaired Trafficking

Cited by 6 publications

References 72 publications

Insights into the Role of Membrane Lipids in the Structure, Function and Regulation of Integral Membrane Proteins

Insights into the Role of Membrane Lipids in the Structure, Function and Regulation of Integral Membrane Proteins

Metabolic depletion of sphingolipids inhibits agonist‐induced endocytosis of the serotonin1Areceptor

Smith-Lemli-Opitz syndrome: A pathophysiological manifestation of the Bloch hypothesis

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Product

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Metabolic Depletion of Sphingolipids Reduces Cell Surface Population of the Human Serotonin_1A Receptor due to Impaired Trafficking

Metabolic depletion of sphingolipids inhibits agonist‐induced endocytosis of the serotonin_1Areceptor