September 2012

Insights into the regulation of phosphatidate phosphatase activity and lipid homeostasis have come to light through studies with yeast

Lipid droplets inside yeast cells; photo courtesy of Sepp Kohlwein 
Lipid droplets inside yeast cells.
Photo courtesy of Sepp Kohlwein. 

The obesity epidemic and derivative metabolic diseases, such as diabetes, have stimulated much interest in controlling fat (triacylglycerol) metabolism. Of many enzymes involved in fat production, phosphatidate phosphatase, or PAP, has emerged as an important regulatory enzyme. PAP, which catalyzes the dephosphorylation of phosphatidate to produce diacylglycerol, functions at the penultimate step of triacylglycerol synthesis and controls the pathways by which membrane phospholipids are synthesized. Because the lipid precursors phosphatidate and diacylglycerol also play roles as signaling molecules, the importance of PAP activity is implicated in the control of diverse cellular processes, including transcription, activation of cell growth, membrane proliferation, secretion and vesicular trafficking.

The yeast Saccharomyces cerevisiae serves as an excellent eukaryotic model in elucidating the enzymological, kinetic and regulatory properties of PAP. A gene (PAH1) encoding PAP was first identified in S. cerevisiae, leading to the identification of homologous PAP-encoding genes in humans, mice, flies, worms and plants. The phenotypic characterization of yeast cells lacking or overexpressing the PAP gene has revealed the importance of the enzyme in lipid metabolism and cell physiology. The PAP-deficient cells, which contain reduced levels of diacylglycerol and increased levels of phosphatidate, exhibit a great reduction in triacylglycerol synthesis and a marked increase in phospholipid synthesis. The elevated level of phosphatidate also is associated with the induced expression of phospholipid synthesis genes and an abnormal expansion of the nuclear/endoplasmic reticulum membrane. The reduced level of diacylglycerol correlates with defects in the formation of the lipid droplet (fat storage organelle) and in vacuole homeostasis. Reduced ability to synthesize triacylglycerol renders the PAP mutant acutely sensitive to fatty acid-induced lipotoxicity. On the flip side, higher PAP activity is also deleterious to lipid homeostasis and cell physiology. For example, the overexpression of an unregulated phosphorylation-deficient form of PAP is lethal, and the toxic effect is attributed to the depletion of phosphatidate required for phospholipid synthesis and to the accumulation of diacylglycerol to a toxic level. Thus, a tight regulation of PAP activity is essential to balance the synthesis and turnover of lipids.

Recent studies with yeast PAP have shown that phosphorylation/dephosphorylation is a major mechanism by which the enzyme activity is controlled. PAP is a peripheral membrane enzyme, which translocates from the cytosol to the nuclear/endoplasmic reticulum membrane via interaction with the Nem1p-Spo7p phosphatase complex. The enzyme in the cytosol is highly phosphorylated, and the phosphorylated form is recruited to and dephosphorylated by the membrane-associated phosphatase complex. Upon its dephosphorylation, PAP directly associates with the membrane using its N-terminal amphipathic helix. Phosphorylation of PAP is mediated by multiple protein kinases that include cyclin-dependent kinases (e.g., Pho85p and Cdc28p), protein kinase A, protein kinase C and casein kinase II. Identification of phosphorylation target sites and the interdependencies of the various phosphorylations are being interrogated with the aim of understanding how PAP activity is fine-tuned to control lipid homeostasis.

 

Photo of Gil-Soo HanGil-Soo Han (gshan@rci.rutgers.edu) is an assistant research professor and member of the Center for Lipid Research at Rutgers University.






Photo of George M. CarmanGeorge M. Carman (carman@aesop.rutgers.edu) is a Rutgers Board of Governors professor and director of the Center for Lipid Research at Rutgers University.

 
 
 


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