Cunningham T W, Lips D L, Bansal V S, Caldwell K K, Mitchell C A, Majerus P W
Washington University School of Medicine, Department of Internal Medicine and Biological Chemistry, St. Louis, Missouri 63110.
J Biol Chem. 1990 Dec 15;265(35):21676-83.
We have identified the structure of phosphatidylinositol 3-phosphate (PtdIns(3)P), phosphatidylinositol 3,4-bisphosphate (PtdIns(3,4)P2) and phosphatidylinositol 3,4,5-trisphosphate (PtdIns(3,4,5)P3) in human platelets. These lipids accounted for less than 2% of the total 32P incorporated into inositol phospholipids in the platelets. All three lipids were labeled in unstimulated platelets, but incorporation of 32P changed rapidly by 15 s after thrombin stimulation, suggesting that they are important in platelet activation. Specific inositol polyphosphate phosphatases were used to both identify the lipid structures and to determine the route of synthesis of these lipids. During 32P labeling and after thrombin stimulation of human platelets, as much as 60% of the total radioactivity present in PtdIns(3,4)P2 was found in the D-4 phosphate and only 35% in the D-3 phosphate indicating that PtdIns(3)P is the precursor of PtdIns(3,4)P2. In addition, the D-5 and D-4 phosphates of PtdIns(3,4,5)P3 each contained 35-40% of the total radioactivity in the molecule compared with only 18-28% in the D-3 position, suggesting that PtdIns(3,4)P2 and not PtdIns(4,5)P2 is the major precursor of this lipid. These results define the predominant pathway for synthesis of these lipids in platelets as PtdIns----PtdIns(3)P----PtdIns(3,4)P2----PtdIns(3,4,5)P3.
我们已经确定了人血小板中磷脂酰肌醇3-磷酸（PtdIns(3)P）、磷脂酰肌醇3,4-二磷酸（PtdIns(3,4)P2）和磷脂酰肌醇3,4,5-三磷酸（PtdIns(3,4,5)P3）的结构。这些脂质占血小板中掺入32P的总肌醇磷脂的比例不到2%。所有这三种脂质在未受刺激的血小板中均有标记，但在凝血酶刺激后15秒内，32P的掺入迅速变化，这表明它们在血小板激活中起重要作用。使用特异性肌醇多磷酸磷酸酶来鉴定脂质结构并确定这些脂质的合成途径。在人血小板的32P标记期间以及凝血酶刺激后，发现PtdIns(3,4)P2中存在的总放射性中多达60%存在于D-4磷酸中，而D-3磷酸中仅占35%，这表明PtdIns(3)P是PtdIns(3,4)P2的前体。此外，PtdIns(3,4,5)P3的D-5和D-4磷酸各自在分子中含有35 - 40%的总放射性，而D-3位置仅占18 - 28%，这表明PtdIns(3,4)P2而非PtdIns(4,5)P2是这种脂质的主要前体。这些结果确定了血小板中这些脂质合成的主要途径为PtdIns→PtdIns(3)P→PtdIns(3,4)P2→PtdIns(3,4,5)P3。
