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The Journal of Biological Chemistry Mar 2023Diacylglycerol kinase (DGK) α, which is a key enzyme in the progression of cancer and, in contrast, in T-cell activity attenuation, preferentially produces saturated...
Diacylglycerol kinase (DGK) α, which is a key enzyme in the progression of cancer and, in contrast, in T-cell activity attenuation, preferentially produces saturated fatty acid (SFA)- and/or monounsaturated fatty acid (MUFA)-containing phosphatidic acids (PAs), such as 16:0/16:0-, 16:0/18:0-, and 16:1/16:1-PA, in melanoma cells. In the present study, we searched for the target proteins of 16:0/16:0-PA in melanoma cells and identified heat shock protein (HSP) 27, which acts as a molecular chaperone and contributes to cancer progression. HSP27 more strongly interacted with PA than other phospholipids, including phosphatidylcholine, phosphatidylserine, phosphatidylglycerol, cardiolipin, phosphatidylinositol, phosphatidylinositol 4-monophosphate, and phosphatidylinositol 4,5-bisphosphate. Moreover, HSP27 is more preferentially bound to SFA- and/or MUFA-containing PAs, including 16:0/16:0- and 16:0/18:1-PAs, than PUFA-containing PAs, including 18:0/20:4- and 18:0/22:6-PA. Furthermore, HSP27 and constitutively active DGKα expressed in COS-7 cells colocalized in a DGK activity-dependent manner. Notably, 16:0/16:0-PA, but not phosphatidylcholine or 16:0/16:0-phosphatidylserine, induced oligomer dissociation of HSP27, which enhances its chaperone activity. Intriguingly, HSP27 protein was barely detectable in Jurkat T cells, while the protein band was intensely detected in AKI melanoma cells. Taken together, these results strongly suggest that SFA- and/or MUFA-containing PAs produced by DGKα selectively target HSP27 and regulate its cancer-progressive function in melanoma cells but not in T cells.
Topics: Humans; Fatty Acids; Fatty Acids, Monounsaturated; HSP27 Heat-Shock Proteins; Phosphatidic Acids; Phosphatidylserines; Phosphatidylinositols; Phosphatidylcholines; Melanoma
PubMed: 36791913
DOI: 10.1016/j.jbc.2023.103019 -
The Plant Journal : For Cell and... Nov 2022Cytosolic glyceraldehyde-3-phosphate dehydrogenase (GAPC) is a glycolytic enzyme, but undergoes stress-induced nuclear translocation for moonlighting. We previously...
Cytosolic glyceraldehyde-3-phosphate dehydrogenase (GAPC) is a glycolytic enzyme, but undergoes stress-induced nuclear translocation for moonlighting. We previously reported that in response to heat stress, GAPC accumulated in the nucleus to modulate transcription and thermotolerance. Here we show a cellular and molecular mechanism that mediates heat-induced nuclear translocation of cytosolic GAPC in Arabidopsis thaliana. Heat-induced GAPC nuclear accumulation and plant heat tolerance were reduced in Arabidopsis phospholipase D (PLD) knockout mutants of pldδ and pldα1pldδ, but not of pldα1. These changes were restored to wild type by genetic complementation with active PLDδ, but not with catalytically inactive PLDδ. GAPC overexpression enhanced the seedling thermotolerance and the expression of heat-inducible genes, but this effect was abolished in the pldδ background. Heat stress elevated the levels of the PLD product phosphatidic acid (PA) in the nucleus in wild type, but not in pldδ plants. Lipid labeling demonstrated the heat-induced nuclear co-localization of PA and GAPC, which was impaired by zinc, which inhibited the PA-GAPC interaction, and by the membrane trafficking inhibitor brefeldin A (BFA). The GAPC nuclear accumulation and seedling thermotolerance were also decreased by treatment with zinc or BFA. Our data suggest that PLDδ and PA are critical for the heat-induced nuclear translocation of GAPC. We propose that PLDδ-produced PA mediates the process via lipid-protein interaction and that the lipid mediation acts as a cellular conduit linking stress perturbations at cell membranes to nuclear functions in plants coping with heat stress.
Topics: Arabidopsis; Phosphatidic Acids; Arabidopsis Proteins; Phospholipases; Phospholipase D; Glyceraldehyde-3-Phosphate Dehydrogenases; Seedlings; Brefeldin A; Zinc
PubMed: 36111506
DOI: 10.1111/tpj.15981 -
The Journal of Biological Chemistry Apr 2022The simple structure of phosphatidic acid (PA) belies its complex biological functions as both a key phospholipid biosynthetic intermediate and a potent signaling...
The simple structure of phosphatidic acid (PA) belies its complex biological functions as both a key phospholipid biosynthetic intermediate and a potent signaling molecule. In the latter role, PA controls processes including vesicle trafficking, actin dynamics, cell growth, and migration. However, experimental methods to decode the pleiotropy of PA are sorely lacking. Because PA metabolism and trafficking are rapid, approaches to accurately visualize and manipulate its levels require high spatiotemporal precision. Here, we describe recent efforts to create a suite of chemical tools that enable imaging and perturbation of PA signaling. First, we describe techniques to visualize PA production by phospholipase D (PLD) enzymes, which are major producers of PA, called Imaging Phospholipase D Activity with Clickable Alcohols via Transphosphatidylation (IMPACT). IMPACT harnesses the ability of endogenous PLD enzymes to accept bioorthogonally tagged alcohols in transphosphatidylation reactions to generate functionalized reporter lipids that are subsequently fluorescently tagged via click chemistry. Second, we describe two light-controlled approaches for precisely manipulating PA signaling. Optogenetic PLDs use light-mediated heterodimerization to recruit a bacterial PLD to desired organelle membranes, and photoswitchable PA analogs contain azobenzene photoswitches in their acyl tails, enabling molecular shape and bioactivity to be controlled by light. We highlight select applications of these tools for studying GPCR-G signaling, discovering regulators of PLD signaling, tracking intracellular lipid transport pathways, and elucidating new oncogenic signaling roles for PA. We envision that these chemical tools hold promise for revealing many new insights into lipid signaling pathways.
Topics: Alcohols; Click Chemistry; Optogenetics; Phosphatidic Acids; Phospholipase D; Signal Transduction
PubMed: 35276134
DOI: 10.1016/j.jbc.2022.101810 -
Biochimie Dec 2014Phosphatidic acid (PA) is the simplest phospholipid naturally existing in all-living organisms. It constitutes only a minor fraction of the total cell lipids but has... (Review)
Review
Phosphatidic acid (PA) is the simplest phospholipid naturally existing in all-living organisms. It constitutes only a minor fraction of the total cell lipids but has attracted considerable attention being both a lipid second messenger and a modulator of membrane shape. The pleiotropic functions of PA are the direct consequence of its very simple chemical structure consisting of only two acyl chains linked by ester bonds to two adjacent hydroxyl groups of glycerol, whose remaining hydroxyl group is esterified with a phosphomonoester group. Hence the small phosphate head group of PA gives it the shape of a cone providing flexibility and negative curvatures in the context of a lipid bilayer. In addition, the negatively charged phosphomonoester headgroup of PA is unique because it can potentially carry one or two negative charges playing a role in the recruitment of positively charged molecules to biomembranes. In consequence, PA has been proposed to play various key cellular functions. In the brain, a fine balance between cell growth, migration and differentiation, and cell death is required to sculpt the nervous system during development. In this review, we will summarize the various functions that have been proposed for PA in neuronal development.
Topics: Animals; Axons; Cell Membrane; Cytoskeleton; Dendrites; Humans; Models, Neurological; Neurons; Phosphatidic Acids; Signal Transduction
PubMed: 25111738
DOI: 10.1016/j.biochi.2014.07.026 -
International Journal of Molecular... Feb 2021Phosphatidic acid (PA) is a bioactive phospholipid capable of regulating key biological functions, including neutrophil respiratory burst, chemotaxis, or cell growth and...
Phosphatidic acid (PA) is a bioactive phospholipid capable of regulating key biological functions, including neutrophil respiratory burst, chemotaxis, or cell growth and differentiation. However, the mechanisms whereby PA exerts these actions are not completely understood. In this work, we show that PA stimulates myoblast proliferation, as determined by measuring the incorporation of [H]thymidine into DNA and by staining the cells with crystal violet. PA induced the rapid phosphorylation of Akt and ERK1/2, and pretreatment of the cells with specific small interferin RNA (siRNA) to silence the genes encoding these kinases, or with selective pharmacologic inhibitors, blocked PA-stimulated myoblast proliferation. The mitogenic effects of PA were abolished by the preincubation of the myoblasts with pertussis toxin, a Gi protein inhibitor, suggesting the implication of Gi protein-coupled receptors in this action. Although some of the effects of PA have been associated with its possible conversion to lysoPA (LPA), treatment of the myoblasts with PA for up to 60 min did not produce any significant amount of LPA in these cells. Of interest, pharmacological blockade of the LPA receptors 1 and 2, or specific siRNA to silence the genes encoding these receptors, abolished PA-stimulated myoblast proliferation. Moreover, PA was able to compete with LPA for binding to LPA receptors, suggesting that PA can act as a ligand of LPA receptors. It can be concluded that PA stimulates myoblast proliferation through interaction with LPA1 and LPA2 receptors and the subsequent activation of the PI3K/Akt and MEK/ERK1-2 pathways, independently of LPA formation.
Topics: Animals; Cell Differentiation; Cell Line; Cell Proliferation; Chemotaxis; DNA; Lysophospholipids; Mice; Myoblasts; Phosphatidic Acids; Phosphatidylinositol 3-Kinases; Phosphorylation; Protein Binding; RNA, Small Interfering; Receptors, Lysophosphatidic Acid; Signal Transduction
PubMed: 33535610
DOI: 10.3390/ijms22031452 -
The Journal of Biological Chemistry Nov 2017Phosphatidic acid (PA) and the conserved integral ER membrane protein Scs2p regulate localization of the transcriptional repressor Opi1p, which controls expression of... (Review)
Review
Phosphatidic acid (PA) and the conserved integral ER membrane protein Scs2p regulate localization of the transcriptional repressor Opi1p, which controls expression of phospholipid biosynthesis genes, but the mechanisms conducting Opi1p localization are not fully understood. A new study suggests the existence of a distinct pool of PA in the ER that is required for regulation of Opi1p localization and thus phospholipid metabolism in yeast.
Topics: Cell Nucleus; Endoplasmic Reticulum; Gene Expression Regulation, Bacterial; Mitochondria; Models, Biological; Phosphatidic Acids; Phospholipids; Phosphorylation; Protein Processing, Post-Translational; Protein Transport; Repressor Proteins; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins
PubMed: 29127205
DOI: 10.1074/jbc.H117.809970 -
Handbook of Experimental Pharmacology 2020Lipids play a vital role in numerous cellular functions starting from a structural role as major constituents of membranes to acting as signaling intracellular or... (Review)
Review
Lipids play a vital role in numerous cellular functions starting from a structural role as major constituents of membranes to acting as signaling intracellular or extracellular entities. Accordingly, it has been known for decades that lipids, especially those coming from diet, are important to maintain normal physiological functions and good health. On the other side, the exact molecular nature of these beneficial or deleterious lipids, as well as their precise mode of action, is only starting to be unraveled. This recent improvement in our knowledge is largely resulting from novel pharmacological, molecular, cellular, and genetic tools to study lipids in vitro and in vivo. Among these important lipids, phosphatidic acid plays a unique and central role in a great variety of cellular functions. This review will focus on the proposed functions of phosphatidic acid generated by phospholipase D in the last steps of regulated exocytosis with a specific emphasis on hormonal and neurotransmitter release and its potential impact on different neurological diseases.
Topics: Exocytosis; Humans; Nervous System Diseases; Phosphatidic Acids; Phospholipase D; Signal Transduction
PubMed: 30570690
DOI: 10.1007/164_2018_180 -
Nature Communications Nov 2021AGPATs (1-acylglycerol-3-phosphate O-acyltransferases) catalyze the acylation of lysophosphatidic acid to form phosphatidic acid (PA), a key step in the...
AGPATs (1-acylglycerol-3-phosphate O-acyltransferases) catalyze the acylation of lysophosphatidic acid to form phosphatidic acid (PA), a key step in the glycerol-3-phosphate pathway for the synthesis of phospholipids and triacylglycerols. AGPAT2 is the only AGPAT isoform whose loss-of-function mutations cause a severe form of human congenital generalized lipodystrophy. Paradoxically, AGPAT2 deficiency is known to dramatically increase the level of its product, PA. Here, we find that AGPAT2 deficiency impairs the biogenesis and growth of lipid droplets. We show that AGPAT2 deficiency compromises the stability of CDP-diacylglycerol (DAG) synthases (CDSs) and decreases CDS activity in both cell lines and mouse liver. Moreover, AGPAT2 and CDS1/2 can directly interact and form functional complexes, which promote the metabolism of PA along the CDP-DAG pathway of phospholipid synthesis. Our results provide key insights into the regulation of metabolic flux during lipid synthesis and suggest substrate channelling at a major branch point of the glycerol-3-phosphate pathway.
Topics: Acyltransferases; Animals; Biosynthetic Pathways; Cell Line; Cytidine Diphosphate Diglycerides; Diacylglycerol Cholinephosphotransferase; Fatty Acids; Humans; Lipid Droplets; Lipogenesis; Liver; Mice; Multienzyme Complexes; Oleic Acid; Phosphatidic Acids
PubMed: 34824276
DOI: 10.1038/s41467-021-27279-4 -
Biomolecules Sep 2020Phosphatidic acid (PA) is a glycerophospholipid intermediate in the triglyceride synthesis pathway that has incredibly important structural functions as a component of... (Review)
Review
Phosphatidic acid (PA) is a glycerophospholipid intermediate in the triglyceride synthesis pathway that has incredibly important structural functions as a component of cell membranes and dynamic effects on intracellular and intercellular signaling pathways. Although there are many pathways to synthesize and degrade PA, a family of PA phosphohydrolases (lipin family proteins) that generate diacylglycerol constitute the primary pathway for PA incorporation into triglycerides. Previously, it was believed that the pool of PA used to synthesize triglyceride was distinct, compartmentalized, and did not widely intersect with signaling pathways. However, we now know that modulating the activity of lipin 1 has profound effects on signaling in a variety of cell types. Indeed, in most tissues except adipose tissue, lipin-mediated PA phosphohydrolase activity is far from limiting for normal rates of triglyceride synthesis, but rather impacts critical signaling cascades that control cellular homeostasis. In this review, we will discuss how lipin-mediated control of PA concentrations regulates metabolism and signaling in mammalian organisms.
Topics: Adipose Tissue; Glycerophospholipids; Humans; Metabolic Networks and Pathways; Muscle, Skeletal; Phosphatidate Phosphatase; Phosphatidic Acids; Protein Conformation; Triglycerides
PubMed: 33003344
DOI: 10.3390/biom10101386 -
BMC Biology Jun 2020Lymphocytes circulate between peripheral lymphoid tissues via blood and lymphatic systems, and chemokine-induced migration is important in trafficking lymphocytes to...
BACKGROUND
Lymphocytes circulate between peripheral lymphoid tissues via blood and lymphatic systems, and chemokine-induced migration is important in trafficking lymphocytes to distant sites. The small GTPase Rap1 is important in mediating lymphocyte motility, and Rap1-GEFs are involved in chemokine-mediated Rap1 activation. Here, we describe the roles and mechanisms of Rap1-GEFs in lymphocyte trafficking.
RESULTS
In this study, we show that RA-GEF-1 and 2 (also known as Rapgef2 and 6) are key guanine nucleotide exchange factors (GEF) for Rap1 in lymphocyte trafficking. Mice harboring T cell-specific knockouts of Rapgef2/6 demonstrate defective homing and egress of T cells. Sphingosine-1-phosphate (S1P) as well as chemokines activates Rap1 in a RA-GEF-1/2-dependent manner, and their deficiency in T cells impairs Mst1 phosphorylation, cell polarization, and chemotaxis toward S1P gradient. On the other hand, B cell-specific knockouts of Rapgef2/6 impair chemokine-dependent retention of B cells in the bone marrow and passively facilitate egress. Phospholipase D2-dependent production of phosphatidic acid by these chemotactic factors determines spatial distribution of Rap1-GTP subsequent to membrane localization of RA-GEFs and induces the development of front membrane. On the other hand, basal de-phosphorylation of RA-GEFs is necessary for chemotactic factor-dependent increase in GEF activity for Rap1.
CONCLUSIONS
We demonstrate here that subcellular distribution and activation of RA-GEFs are key factors for a directional movement of lymphocytes and that phosphatidic acid is critical for membrane translocation of RA-GEFs with chemokine stimulation.
Topics: Animals; Cell Line; Cell Movement; Female; Guanine Nucleotide Exchange Factors; Humans; Lymphocytes; Male; Mice; Phosphatidic Acids; Phosphorylation
PubMed: 32600317
DOI: 10.1186/s12915-020-00809-0