Lee T C, Malone B, Blank M L, Fitzgerald V, Snyder F
Medical Sciences Division, Oak Ridge Associated Universities, Tennessee 37831-0117.
J Biol Chem. 1990 Jun 5;265(16):9181-7.
We have established previously that 1-alkyl-2-acetyl-sn-glycerol (alkylacetyl-G) can be converted into at least six metabolites by rabbit platelets, including alkylacetyl-sn-(glycero-3-phosphocholine) (-GPC), i.e. platelet-activating factor (PAF) and 1-alkyl-2-acyl-sn- (alkylacyl)-GPC. Since part of the biological functions of alkylacetyl-G can be explained by its metabolic conversion to PAF and also to alkylacyl-GPC as an inactive storage precursor of PAF, the present study focused on the regulation of the synthesis of PAF and alkylacyl-GPC from alkylacetyl-G. Our results document the presence of a specific dithiothreitol (DTT)-insensitive cholinephosphotransferase in saponin-permeabilized rabbit platelets and show that DTT potentiates the production of PAF from alkylacetyl-G but inhibits the formation of phosphatidylcholine from diolein. We also demonstrated that the availability of CDP-choline controls the generation of PAF from alkylacetyl-G. Furthermore, when CTP: phosphocholine cytidylyltransferase is activated to produce more CDP-choline through the translocation of this enzyme from the cytosol to membranes by incubating the rabbit platelets with 0.2 mM sodium oleate, the production of PAF from alkylacetyl-G is increased 5-fold. More importantly, our experiments reveal the presence of two metabolic pathways that are responsible for the synthesis of alkylacyl-GPC from alkylacetyl-G, with each producing a unique molecular species composition of the stored PAF precursor, alkylacyl-GPC. The latter is enriched in polyunsaturates (70.7-78.5% 20:4) when formed through the remodeling pathway of PAF cycle via alkylacetyl-G (DTT-insensitive cholinephosphotransferase)----alkylacetyl-GPC----alkyllyso-GPC---- alkylacyl-GPC . Alkylacyl-GPC containing saturated species (71.8% 16:0) is generated by the retroconversion/de novo pathway according to the reaction scheme of alkylacetyl-G----alkyl-G----alkyllyso-glycero-3-phosphate (-GP)----alkylacyl-GP----alkylacyl-G (DTT-sensitive cholinephosphotransferase)----alkylacyl-GPC. Inactivation of PAF through the remodeling/PAF cycle can generate alkylacyl-GPC at both low (1.75 x 10(-7) M) and high (10(-6) M) concentrations of PAF whereas the conversion of alkylacetyl-G to alkylacyl-GPC via PAF through the remodeling pathway only occurs at a low concentration (1.75 x 10(-7) M). At a high concentration (10(-6) M), alkylacetyl-G is converted to alkylacyl-GPC via the retroconversion/de novo route. These data suggest that the formation of PAF by the DTT-insensitive cholinephosphotransferase activity limits the amounts of alkylacyl-GPC produced from alkylacetyl-G through this remodeling pathway (PAF cycle).(ABSTRACT TRUNCATED AT 400 WORDS)
我们之前已经证实，1-烷基-2-乙酰基-sn-甘油（烷基乙酰基-G）可被兔血小板转化为至少六种代谢产物，包括烷基乙酰基-sn-（甘油-3-磷酸胆碱）（-GPC），即血小板活化因子（PAF）和1-烷基-2-酰基-sn-（烷基酰基）-GPC。由于烷基乙酰基-G的部分生物学功能可通过其代谢转化为PAF以及作为PAF无活性储存前体的烷基酰基-GPC来解释，因此本研究聚焦于从烷基乙酰基-G合成PAF和烷基酰基-GPC的调控。我们的结果证明，在皂素通透的兔血小板中存在一种对二硫苏糖醇（DTT）不敏感的胆碱磷酸转移酶，并且表明DTT可增强从烷基乙酰基-G生成PAF的过程，但抑制由二油精生成磷脂酰胆碱的过程。我们还证明，CDP-胆碱的可用性控制着从烷基乙酰基-G生成PAF的过程。此外，当通过用0.2 mM油酸钠孵育兔血小板，使CTP：磷酸胆碱胞苷转移酶活化，通过该酶从胞质溶胶向膜的转位产生更多的CDP-胆碱时，从烷基乙酰基-G生成PAF的量增加了5倍。更重要的是，我们的实验揭示了存在两条负责从烷基乙酰基-G合成烷基酰基-GPC的代谢途径，每条途径产生储存的PAF前体烷基酰基-GPC的独特分子种类组成。当通过PAF循环的重塑途径经由烷基乙酰基-G（对DTT不敏感的胆碱磷酸转移酶）----烷基乙酰基-GPC----烷基溶血-GPC----烷基酰基-GPC形成时，后者富含多不饱和脂肪酸（70.7 - 78.5% 20:4）。根据烷基乙酰基-G----烷基-G----烷基溶血甘油-3-磷酸（-GP）----烷基酰基-GP----烷基酰基-G（对DTT敏感的胆碱磷酸转移酶）----烷基酰基-GPC的反应方案，含有饱和种类（71.8% 16:0）的烷基酰基-GPC通过逆向转化/从头合成途径生成。通过重塑/PAF循环使PAF失活可在低（1.75×10⁻⁷ M）和高（10⁻⁶ M）浓度的PAF下生成烷基酰基-GPC，而通过重塑途径经由PAF将烷基乙酰基-G转化为烷基酰基-GPC仅在低浓度（1.75×10⁻⁷ M）下发生。在高浓度（10⁻⁶ M）时，烷基乙酰基-G通过逆向转化/从头合成途径转化为烷基酰基-GPC。这些数据表明，对DTT不敏感的胆碱磷酸转移酶活性形成PAF限制了通过该重塑途径（PAF循环）从烷基乙酰基-G产生的烷基酰基-GPC的量。（摘要截断于400字）
