The Project Leader's laboratory was the first to describe the inducible PGS2 cDNA, protein, and gene. Subsequent work demonstrated that non-steroidal anti-inflammatory drugs exert much of their anti-inflammatory effects by inhibiting PGS2 enzyme activity. The applicant also made the first inroads into the apparent paradox presented by the presence, in the same cell, of the constitutive prostaglandin synthase (PGS1) and inducible prostaglandin synthase (PGS2). The applicant and his colleagues showed that arachidonic acid generated by phospholipase (PL) activation in response to mitogens, inflammatory stimuli, and other agents is often sequestered from the constitutive PGS1 enzyme present in cells and can only be converted to prostaglandins via the PGS2 enzyme induced in response to these stimuli. In work performed in this AAIDCRC, Dr. Herschman and his co-workers made the singular discovery that prostaglandin production in activated mast cells differs from prostaglandin production in nearly all other cells. Specifically, production of prostaglandin D2 production in activated mast cells occurs in two phases, an early phase that couples a distinct PL to PGS1 and a late phase in which a second PL is coupled to PGS2. The applicant now plans the following: (1) to identify the phospholipase A2 (PLA2) enzymes that provide arachidonate to PGS1 and PGS2, (2) to determine the subcellular localizatio of the PGS1, PGS2, type V soluble (s)PLA2 and type IV cytoplasmic (c)PLA2 proteins in mast cells, to understand the basis for arachidonic acid channeling in prostaglandin synthesis, (3) to determine the amino acid sequences of PGS1 and PGS2 that are responsible for differential temporal accessibility to arachidonic acid and PGS subcellular localization in mast cells, and (4) to determine the cis-acting elements, transcription factors, and signal transduction pathways that mediate PGS2 expression in mast cells. The applicants also discovered that phenanthrenes profoundly inhibits prostaglandin production in activated mast cells and in endotoxin-stimulated macrophages. The investigators now plan (5) to determine how phenanthrene inhibits prostaglandin induction in mast cells and macrophages, and (6) to determine whether phenanthrene enhances leukotriene synthesis in activated mast cells following IgE receptor aggregation. The applicants work will (1) provide new insight into fundamental mechanisms of prostaglandin synthesis in cells that mediate allergic response; (2) identify new targets for pharmacologic modulation of prostaglandin synthesis; and (3) investigate mechanisms by which PAH may modulate airway inflammation.
