Seizure‐protective Receptors GPR37L1 and GPR37 Regulate Calcium‐binding Protein S100A5 Expression and Secretion

Abstract

ObjectiveThe objective of this study is to investigate the mechanisms through which S100A5 expression and secretion is regulated by the receptors, GPR37L1 and GPR37.MethodsProteomic data obtained via mass spectrometry of mouse brain lysates was performed through the Emory Proteomics Core and follow‐up studies were performed studying cultured cells.Summary of resultsA mutation in the G protein‐coupled receptor GPR37L1 was recently found in a large family to be associated with a progressive form of epilepsy in which seizures start around puberty and increase in severity with age, ultimately resulting in death in the late teens to early twenties. Studies from our lab have revealed that knockout mice lacking GPR37L1 (and/or the closely related receptor GPR37) exhibit greatly increased susceptibility to seizures. To shed light on the potential mechanistic underpinnings of this phenotype, we performed proteomic analyses of whole mouse brain tissue from wild‐type vs. GPR37L1/GPR37 double knockout mice to identify proteins regulated by the absence versus presence of these receptors. The most dramatically downregulated protein in the knockout versus wild‐type brain tissue was S100A5, a brain‐specific calcium‐binding protein. The reduction of S100A5 expression in the knockout brain tissue was confirmed via Western blot. Furthermore, when S100A5 is co‐expressed with either GPR37L1 or GPR37 in HEK293T cells, a robust increase in secretion of S100A5 was observed. This secretion was receptor‐dependent, as over‐expression of S100A5 alone did not result in detectable S100A5 in the media. To dissect the G protein signaling pathway that mediates this secretion, inhibitors of the different G protein signaling pathways were used to treat cells that co‐expressed S100A5 with either receptor. These studies suggested that GPR37 and GPR37L1 signal through Gs to mediate S100A5 secretion, as treatment with H89, a protein kinase A (PKA) inhibitor, sharply reduces S100A5 secretion from transfected HEK293T cells. These findings may be of general interest because other members of the S100A protein family are known to be secreted and have neuroprotective actions within the brain, but little is known about the mechanisms controlling their secretion.ConclusionsS100A5 expression and secretion are both regulated by GPR37 and GPR37L1. Our studies reveal that these receptors signal in a PKA‐dependent manner to regulate S100A5 secretion.Support or Funding InformationPharmacological Sciences T32 Predoctoral Training Grant (4T32GM008602‐20), Pharmacological Sciences T32 Predoctoral Training Grant (4T32GM008602‐21), NIH Grants R21‐NS91986 and R01‐NS088413This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.