Conversion of alkylacetylglycerol to platelet-activating factor in HL-60 cells and subcellular localization of the mediator

Journal Article · Thu Feb 01 00:00:00 EST 1990 · Archives of Biochemistry and Biophysics; (USA)

DOI:https://doi.org/10.1016/0003-9861(90)90756-O· OSTI ID:7026874

Vallari, D S; Record, M; Snyder, F ^[1]

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1. Oak Ridge Associated Universities, TN (USA)

A human promyelocytic leukemia (HL-60) cell line was used to investigate the conversion of 1-alkyl-2-acetyl-sn-glycerol (alkylacetyl-G) to platelet-activating factor (PAF; 1-alkyl-2-acetyl-sn-glycero-3-phosphocholine) by intact cells and in subcellular fractions in order to examine the fate of PAF synthesized de novo. Lipid extracts obtained from undifferentiated HL-60 cells incubated with (3H)alkylacetyl-G contained 2-4% of the label as (3H)PAF; several related metabolites were also detected. The yield of (3H)PAF could be dramatically increased by pretreating the cells with either oleic acid, an activator of CTP:phosphocholine cytidylyltransferase, or phenylmethylsulfonyl fluoride, an inhibitor of PAF acetylhydrolase. These results, together with a kinetic study of (3H)alkylacetyl-G metabolism, indicate the sequential participation of a cholinephosphotransferase for the conversion of (3H)-alkylacetyl-G to PAF and acetylhydrolase and transacylase activities in the remodeling pathway that metabolize the newly formed (3H)PAF to 1-(3H)alkyl-2-acyl(long chain)-sn-glycero-3-phosphocholine. The dithiothreitol-insensitive cholinephosphotransferase activity capable of converting alkylacetyl-G to PAF was localized in subcellular fractions that contain CDP-choline:1,2-dioleoyl-sn-glycerol cholinephosphotransferase (dithiothreitol-sensitive), as well as marker enzyme activities for the endoplasmic reticulum and Golgi membranes. Subcellular localization analyses also indicated that the majority of newly formed (3H)PAF and a large portion of its deacetylated metabolite were associated with the plasma membrane-containing fractions, whereas most of the 1-(3H)alkyl-2-acyl(long chain)-sn-glycero-3- phosphocholine was present in the intracellular organelles. Incubations of HL-60 cells with exogenous (3H)PAF produced a similar subcellular distribution of metabolites.

• OSTI ID: 7026874
• Journal Information: Archives of Biochemistry and Biophysics; (USA), Vol. 276:2; ISSN 0003-9861
• Country of Publication: United States
• Language: English

Related Subjects

59 BASIC BIOLOGICAL SCIENCES
BLOOD COAGULATION FACTORS
BIOSYNTHESIS
GLYCEROL
METABOLISM
PHOSPHOTRANSFERASES
ENZYME ACTIVITY
BIOLOGICAL MARKERS
BIOLOGICAL PATHWAYS
ENDOPLASMIC RETICULUM
ETHERS
LEUKEMIA
MAN
METABOLITES
PHOSPHOLIPIDS
SUBCELLULAR DISTRIBUTION
TRACER TECHNIQUES
TRITIUM COMPOUNDS
TUMOR CELLS
ALCOHOLS
ANIMAL CELLS
ANIMALS
CELL CONSTITUENTS
COAGULANTS
DISEASES
DISTRIBUTION
DRUGS
ENZYMES
ESTERS
HEMATOLOGIC AGENTS
HEMIC DISEASES
HYDROGEN COMPOUNDS
HYDROXY COMPOUNDS
IMMUNE SYSTEM DISEASES
ISOTOPE APPLICATIONS
LIPIDS
MAMMALS
NEOPLASMS
ORGANIC COMPOUNDS
ORGANIC OXYGEN COMPOUNDS
ORGANIC PHOSPHORUS COMPOUNDS
ORGANOIDS
PHOSPHORUS-GROUP TRANSFERASES
PRIMATES
PROTEINS
SYNTHESIS
TRANSFERASES
VERTEBRATES
550501* - Metabolism- Tracer Techniques