Phospholipase C (PLC) is a class of membrane-associatedenzymes that cleavephospholipids just before thephosphate group (see figure). It is most commonly taken to be synonymous with the human forms of this enzyme, which play an important role ineukaryoticcellphysiology, in particularsignal transduction pathways. Phospholipase C's role in signal transduction is its cleavage ofphosphatidylinositol 4,5-bisphosphate (PIP2) intodiacyl glycerol (DAG) andinositol 1,4,5-trisphosphate (IP3), which serve assecond messengers. Activators of each PLC vary, but typically includeheterotrimeric G protein subunits, proteintyrosine kinases,small G proteins, Ca2+, and phospholipids.[1]
There are thirteen kinds of mammalian phospholipase C that are classified into six isotypes (β, γ, δ, ε, ζ, η) according to structure. Each PLC has unique and overlapping controls over expression and subcellular distribution. However, PLC is not limited to mammals, and is present in bacteria and Chromadorea as well.
Phospholipase C | |||||||||
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Identifiers | |||||||||
EC no. | 3.1.4.3 | ||||||||
CAS no. | 9001-86-9 | ||||||||
Databases | |||||||||
IntEnz | IntEnz view | ||||||||
BRENDA | BRENDA entry | ||||||||
ExPASy | NiceZyme view | ||||||||
KEGG | KEGG entry | ||||||||
MetaCyc | metabolic pathway | ||||||||
PRIAM | profile | ||||||||
PDB structures | RCSB PDBPDBePDBsum | ||||||||
Gene Ontology | AmiGO /QuickGO | ||||||||
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The extensive number of functions exerted by the PLC reaction requires that it be strictly regulated and able to respond to multiple extra- and intracellular inputs with appropriate kinetics. This need has guided the evolution of six isotypes of PLC in animals, each with a distinct mode of regulation. The pre-mRNA of PLC can also be subject to differential splicing such that a mammal may have up to 30 PLC enzymes.[2]
Most of the bacterial variants of phospholipase C are characterized into one of four groups of structurally related proteins. The toxic phospholipases C are capable of interacting with eukaryotic cell membranes and hydrolyzing phosphatidylcholine and sphingomyelin, leading to cell lysis.[3]
The class ofChromadorea also utilizes the enzyme phospholipase C to regulate the releases of calcium. The enzyme releasesinositol 1,4,5-trisphosphate (IP3) that denotes a signaling pathway involved in activating ovulation, the propelling of the oocyte into the spermatheca. This gene is involved in various activities like controlling GTPase, breaking down certain molecules, and binding to small GTPase. It helps in fighting bacteria and regulating protein movement in cells. It's found in the excretory system, intestines, nerves, and reproductive organs. The expression of the enzyme in the spermatheca is controlled by the transcription factorsFOS-1 and JUN-1.[4]
In mammals, PLCs share a conserved core structure and differ in other domains specific to each family. The core enzyme includes a splittriosephosphate isomerase (TIM) barrel,pleckstrin homology (PH) domain, four tandemEF hand domains, and aC2 domain.[1] The TIM barrel contains the active site, all catalytic residues, and a Ca2+ binding site. It has an autoinhibitory insert that interrupts its activity called an X-Y linker. The X-Y linker has been shown to occlude the active site, and with its removal, PLC is activated.[5]
The genes encodingalpha-toxin (Clostridium perfringens),Bacillus cereusPLC (BC-PLC), and PLCs fromClostridium bifermentans andListeria monocytogenes have been isolated and nucleotides sequenced. The sequences have significant homology, approximately 250 residues, from the N-terminus. Alpha-toxin has an additional 120 residues in the C-terminus. The C-terminus of the alpha-toxin has been reported as a "C2-like" domain, referencing the C2 domain found in eukaryotes that are involved in signal transduction and present in mammalianphosphoinositide phospholipase C.[6]
The primary catalyzed reaction of PLC occurs on an insoluble substrate at a lipid-water interface. The residues in the active site are conserved in all PLC isotypes. In animals, PLC selectively catalyzes the hydrolysis of the phospholipidphosphatidylinositol 4,5-bisphosphate (PIP2) on the glycerol side of the phosphodiester bond. There is the formation of a weakly enzyme-bound intermediate, inositol 1,2-cyclic phosphodiester, and release ofdiacylglycerol (DAG). The intermediate is then hydrolyzed toinositol 1,4,5-trisphosphate (IP3).[7] Thus the two end products are DAG and IP3. The acid/base catalysis requires two conserved histidine residues and a Ca2+ ion is needed for PIP2 hydrolysis. It has been observed that the active-site Ca2+ coordinates with four acidic residues and if any of the residues are mutated then a greater Ca2+ concentration is needed for catalysis.[8]
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Phosphoinositide-specific phospholipase C (PLC) is a key player in cell signaling processes. When cells encounter signals like hormones or growth factors, PLC breaks down a molecule called PIP2 to produce new signaling molecules. PIP2 is a type of molecule found in cell membranes. When cells receive certain signals from outside, an enzyme called PLC breaks down PIP2 into smaller molecules, which then send messages within the cell. Various types of PLC are activated differently, contributing to cells' ability to respond to their surroundings.
Receptors that activate this pathway are mainlyG protein-coupled receptors coupled to theGαq subunit, including:
Other, minor, activators than Gαq are:
PLC cleaves thephospholipidphosphatidylinositol 4,5-bisphosphate (PIP2) intodiacyl glycerol (DAG) andinositol 1,4,5-trisphosphate (IP3). Thus PLC has a profound impact on the depletion of PIP2, which acts as a membrane anchor or allosteric regulator and an agonist for manylipid-gated ion channels.[21][22] PIP2 also acts as the substrate for synthesis of the rarer lipidphosphatidylinositol 3,4,5-trisphosphate (PIP3), which is responsible for signaling in multiple reactions.[23] Therefore, PIP2 depletion by the PLC reaction is critical to the regulation of local PIP3 concentrations both in the plasma membrane and the nuclear membrane.
The two products of the PLC catalyzed reaction, DAG and IP3, are important second messengers that control diverse cellular processes and are substrates for synthesis of other important signaling molecules. When PIP2 is cleaved, DAG remains bound to the membrane, and IP3 is released as a soluble structure into thecytosol. IP3 then diffuses through the cytosol to bind toIP3 receptors, particularlycalcium channels in thesmooth endoplasmic reticulum (ER). This causes the cytosolic concentration of calcium to increase, causing a cascade of intracellular changes and activity.[24] In addition, calcium and DAG together work to activateprotein kinase C, which goes on to phosphorylate other molecules, leading to altered cellular activity.[24] End-effects include taste, tumor promotion, as well as vesicle exocytosis,superoxide production fromNADPH oxidase, andJNK activation.[24][25]
Both DAG and IP3 are substrates for the synthesis of regulatory molecules. DAG is the substrate for the synthesis ofphosphatidic acid, a regulatory molecule. IP3 is the rate-limiting substrate for the synthesis of inositol polyphosphates, which stimulate multiple protein kinases, transcription, and mRNA processing.[26] Regulation of PLC activity is thus vital to the coordination and regulation of other enzymes of pathways that are central to the control of cellular physiology.
Additionally, phospholipase C plays an important role in the inflammation pathway. The binding of agonists such asthrombin,epinephrine, orcollagen, toplatelet surface receptors can trigger the activation of phospholipase C to catalyze the release ofarachidonic acid from two major membrane phospholipids,phosphatidylinositol andphosphatidylcholine. Arachidonic acid can then go on into the cyclooxygenase pathway (producingprostoglandins (PGE1, PGE2, PGF2),prostacyclins (PGI2), orthromboxanes (TXA2)), and the lipoxygenase pathway (producingleukotrienes (LTB4, LTC4, LTD4, LTE4)).[27]
The bacterial variantClostridium perfringens type A produces alpha-toxin. The toxin has phospholipase C activity, and causeshemolysis, lethality, and dermonecrosis. At high concentrations, alpha-toxin induces massive degradation ofphosphatidylcholine andsphingomyelin, producing diacylglycerol andceramide, respectively. These molecules then participate in signal transduction pathways.[6] It has been reported that the toxin activates the arachidonic acid cascade in isolated rat aorta.[28] The toxin-induced contraction was related to generation of thromboxane A2 from arachidonic acid. Thus it is likely the bacterial PLC mimics the actions of endogenous PLC in eukaryotic cell membranes.