Abstract
Mammalian patatin-like phospholipase domain–containing proteins (PNPLAs) are lipid-metabolizing enzymes with essential roles in energy metabolism, skin barrier development, and brain function. A detailed annotation of enzymatic activities and structure–function relationships remains an important prerequisite to understand PNPLA functions in (patho-)physiology, for example, in disorders such as neutral lipid storage disease, non-alcoholic fatty liver disease, and neurodegenerative syndromes. In this study, we characterized the structural features controlling the subcellular localization and enzymatic activity of PNPLA7, a poorly annotated phospholipase linked to insulin signaling and energy metabolism. We show that PNPLA7 is an endoplasmic reticulum (ER) transmembrane protein that specifically promotes hydrolysis of lysophosphatidylcholine in mammalian cells. We found that transmembrane and regulatory domains in the PNPLA7 N-terminal region cooperate to regulate ER targeting but are dispensable for substrate hydrolysis. Enzymatic activity is instead mediated by the C-terminal domain, which maintains full catalytic competence even in the absence of N-terminal regions. Upon elevated fatty acid flux, the catalytic domain targets cellular lipid droplets and promotes interactions of PNPLA7 with these organelles in response to increased cAMP levels. We conclude that PNPLA7 acts as an ER-anchored lysophosphatidylcholine hydrolase that is composed of specific functional domains mediating catalytic activity, subcellular positioning, and interactions with cellular organelles. Our study provides critical structural insights into an evolutionarily conserved class of phospholipid-metabolizing enzymes.
Highlights
Mammalian patatin-like phospholipase domain– containing proteins (PNPLAs) are lipid-metabolizing enzymes with essential roles in energy metabolism, skin barrier development, and brain function
We assessed the role of PNPLA7 in cellular lipid metabolism and analyzed the structure–function relationships between domain architecture, subcellular distribution, and enzymatic activity of the protein
PNPLA7 is closely related to PNPLA6, a protein involved in axon maintenance and brain function, which has been previously shown to regulate hydrolysis of cellular LPC and PC [11, 14, 22]
Summary
Functional domains of PNPLA7 cause a neurotoxic syndrome termed OP-induced delayed neuropathy. In contrast to PNPLA6, little is known about the molecular and physiological function(s) of the closely related PNPLA7 ( termed neuropathy target esterase-related esterase) Both proteins share a highly conserved domain architecture that is assembled of the enzymatic patatin-like phospholipase domain and extensive “non-enzymatic” segments of poorly defined function including three putative cyclic nucleotide monophosphate (cNMP)-binding sites [1, 25]. PNPLA7 localizes to the ER and lipid droplets (LDs), which are cellular lipid storage organelles with pivotal functions in energy metabolism and lipid trafficking [25, 27] These observations closely link PNPLA7 to lipid and energy metabolism, it is presently unknown how fluctuations in PNPLA7 expression or subcellular distribution affect lipid homeostasis of cells or tissues [25]. Our study provides novel structural insights into an evolutionarily conserved class of phospholipid-metabolizing enzymes
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