LYSOPHOSPHATIDIC ACID-INDUCED CA2- STRUCTURE-ACTIVITY ANALYSIS( MOBILIZATION IN HUMAN A431 CELLS )

Lysophosphatidic acid (LPA; 1-acyl-sn-glycero-3-phosphate) is a platel et-derived lipid mediator that activates its own G-protein-coupled rec eptor to trigger phospholipase C-mediated Ca2+ mobilization and other effector pathways in numerous cell types. In this study we have examin ed the structural features of LPA that are important for activation of the Ca2+-mobilizing receptor in human A431 carcinoma cells, which sho w an EC(50) for oleoyl-LPA as low as 0.2 nM. When the acyl chain at th e sn-1 position is altered, the rank order of potency is oleoyl-LPA > arachidonoyl-LPA > linolenoyl-LPA > linoleoyl-LPA > stearoyl-LPA = pal mitoyl-LPA > myristoyl-LPA. The shorter-chain species, lauroyl- and de canoyl-LPA, show little or no activity. Ether-linked LPA (1-O-hexadecy l-sn-glycero-3-phosphate) is somewhat less potent than the correspondi ng ester-linked LPA; its stereoisomer is about equally active. Deletio n of the glycerol backbone causes a 1000-fold decrease in potency. Rep lacement of the phosphate group in palmitoyl-LPA by a hydrogen- or met hyl-phosphonate moiety results in complete loss of activity. A phospho nate analogue with a methylene group replacing the oxygen at sn-3 has strongly decreased activity. All three phosphonate analogues induce ce ll lysis at doses > 15 mu M. Similarly, the methyl and ethyl esters of palmitoyl-LPA are virtually inactive and become cytotoxic at micromol ar doses. None of the LPA analogues tested has antagonist activity. Sp hingosine 1-phosphate, a putative messenger with some structural simil arities to LPA, elicits a transient rise in intracellular [Ca2+] only at micromolar doses; however, cross-desensitization experiments indica te that sphingosine 1-phosphate does not act through the LPA receptor. The results indicate that, although many features of the LPA structur e are important for optimal activity, the phosphate group is most crit ical, suggesting that this moiety is directly involved in receptor act ivation.