Assaying nonspecific phospholipase C activity
Language English Country United States Media print
Document type Journal Article, Research Support, Non-U.S. Gov't
- MeSH
- Chromatography, Thin Layer MeSH
- Diglycerides metabolism MeSH
- Enzyme Assays methods MeSH
- Fluorescent Dyes metabolism MeSH
- Type C Phospholipases metabolism MeSH
- Reference Standards MeSH
- Plants enzymology MeSH
- Substrate Specificity MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
- Names of Substances
- 1,2-diacylglycerol MeSH Browser
- Diglycerides MeSH
- Fluorescent Dyes MeSH
- Type C Phospholipases MeSH
Plant nonspecific phospholipase C (NPC) is a recently described enzyme which plays a role in membrane rearrangement during phosphate starvation. It is also involved in responses of plants to brassinolide, abscisic acid (ABA), elicitors, and salt. The NPC activity is decreased in cells treated with aluminum. In the case of salt stress, the molecular mechanism of NPC action is based on accumulation of diacylglycerol (DAG) by hydrolysis of phospholipids and conversion of DAG, the product of NPC activity, to phosphatidic acid (PA) that participates in ABA signaling pathways. Here we describe a step-by-step protocol, which can be used to determine in situ or in vitro NPC activity. Determination is based on quantification of fluorescently labeled DAG as a product of cleavage of the fluorescently labeled substrate lipid, phosphatidylcholine. High-performance thin-layer chromatography is used for separation of fluorescent DAG. The spot is visualized with a laser scanner and the relative amounts of fluorescent DAG are quantified using imaging software.
References provided by Crossref.org
Non-specific phospholipase C4 mediates response to aluminum toxicity in Arabidopsis thaliana
