Phospholipases hydrolyze membrane phospholipids (see Lipases). The lipid substrates for these enzymes are usually quite specific. Hormone-regulated phospholipase C (PLC) can hydrolyze a number of membrane phospholipids, including phosphatidylchloline and phosphatidylethanolamine. Phosphatidylinositol (PI) and its phosphorylated derivatives (polyphosphoinositides) are the most common substrates involved in second messenger generation, producing soluble inositol phosphates, such as inositol trisphosphate (IP3), that are involved in calcium signaling.
Phospholipase D enzymes usually prefer phosphatidylcholine as substrate, generating phosphatidic acid and choline. Phosphatidic acid has been proposed to mediate a variety of biological responses involved in mitogenesis and protein trafficking (1).
The turnover of polyphosphoinositides occurs in a cycle of hydrolysis and resynthesis, called the "PI cycle." PI is phosphorylated sequentially on the 4′ and 5′ hydroxyl groups of the inositol ring, to produce PI-4′,5′-P2. Upon hormonal activation, PLC subsequently catalyzes the phosphodiesteratic cleavage of this lipid, resulting in the generation of diacylglycerol and IP3 in the cell. After its degradation, PI can be resynthesized by the phosphorylation of diacylglycerol to PA, via the activity of a diacylglycerol kinase and the subsequent resynthesis of PI via PI synthase. Thus, this cycle results in the generation of two potential second messengers, the membrane-associated diacylglycerol, which can activate protein kinase C, as well as IP3, which can lead to the mobilization of intracellular calcium levels.
Several forms of PI-PLC have been identified (1). These proteins all exhibit an absolute requirement for calcium and are equally capable of hydrolyzing PI and the polyphosphoinositides PIP and PI-4′,5′-P2. These proteins exist in three subfamilies, b, g, and d, each with multiple members. They have a number of conserved and divergent domains, reflecting their differential regulation. All three subclasses have a pleckstrin homology (PH) domain, which probably mediates some type of interaction with lipids in the membrane, and an EF-hand motif, presumably involved in calcium binding. Moreover, there are two additional conserved domains, designated X and Y, that contain the catalytic core of the enzyme; there is also a conserved C2 domain, also likely to be required for calcium interaction. PLCg, which is regulated by tyrosine kinase receptors, contains two SH2 domains and one SH3 domain and has been shown to undergo phosphorylation of tyrosine residues. PLCb is responsible for regulation through G-protein-coupled receptors and is generally regulated by a subunits of the Gq family of heterotrimeric G proteins, linked to a subset of such receptors. Additionally, their bg subunits have also been shown to regulate PLCb, although the efficacy of this activation is not comparable to that seen with Gqa. PLCb can also act as an activator of the GTPase activity of Gq, effectively behaving as a GTPase activating protein (GAP).
