Zach Zimmerman, Spotlight Week 2
Paper: Identification of the Major Prostaglandin Glycerol Ester Hydrolase in Human Cancer Cells
Enzyme in a hay stack (if hay were hydrolytic…)
Endocannabinoids are lipid molecules implicated in a number of important physiological processes across many body systems. They are metabolized by several enzymes and converted into prostaglandin glyecerol esters (PG-Gs), which are implicated in myriad more processes including neuroinflammation, neural functioning, and high pain sensitivity. Understanding these pathways fully has been challenging historically because of the tendency of PG-Gs to be easily hydrolyzed to prostaglandins (PGs) in vivo. Experiments performed in vitro have shown at least 5 enzymes that will hydrolyze the PG-Gs, 2 of which show activity in vivo as well. A number of these enzymes also accept a broad range of substrates. All of these hydrolytic enzymes are members of the serine hydrolase superfamily, which classifies its members based on the use of an activated serine nucleophile for hydrolysis. Manna et. al. sought to shed light on the notion that a number of low-specific enzymes haphazardly hydrolyze PG-Gs in vivo. They theorized that a previously unidentified serine hydrolase was responsible for the hydrolysis of PGE2G (a specific PG-G) into the prostaglandin PGE2, a reaction that was shown to occur frequently in cancer cells, and attempted to find it.
By feeding human breast cancer cells PGE2G and analyzing the cell culture by LC-MS/MS, the authors show that PGE2G hydrolysis occurs in cancer cells. They demonstrate serine hydrolase activity by selectively inhibiting serine hydrolases and observing eradication of hydrolysis. An interrogation of serine hydrolase proteomics work done by Nomura et. al. against data from five cancer cell line experiments implicates a single protein as the source of PGE2G hydrolysis: lysophospholipase A2 (LYPLA2). siRNA experiments that reduce LYPLA2 gene expression show that without natural levels of LYPLA2, PGE2G hydrolysis is significantly reduced (Figure above, β-actin controlled). cDNA experiments then show PGE2G hydrolytic activity introduced in HEK293 cells. Substrate selectivity of LYPLA2 reveals itself as repeat experiments on the isoform lysophospholipase A1 fail to produce similar activity levels on PGE2G. In the presence of serum protein, LYPLA2 preferentially acts on PGE2G compared to 5 other substrates. By inhibiting LYPLA2 in vivo and observing increases in PG-G levels, the authors establish the importance of LYPLA2 in cells.
The sheer number of hydrolytic enzymes in cells makes it difficult to study the substrates of hydrolysis reactions. Manna et. al. use clever literature analysis to identify LYPLA2 as an important enzyme in cancer cells and then interrogate its role in the cell with numerous techniques. While the identification is exciting, the impact of the study should not be overstated. With so many enzymes performing similar functions and added complications of substrate isomerization, more work will need to affirm the unique relationship between LYPLA2 and PGE2G. If that happens, the true relevance of LYPLA2 as an enzyme in cancerous prostaglandin biosynthesis will emerge.
For Michael Chase: