Coleman, James L. J. and Ngo, Tony and Smythe, Rhyll E. and Cleave, Andrew J. and Jones, Nicole M. and Graham, Robert M. and Smith, Nicola J. (2020) The N-terminus of GPR37L1 is proteolytically processed by matrix metalloproteases. Scientific Reports, 10 (1). ISSN 2045-2322
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Abstract
GPR37L1 is an orphan G protein-coupled receptor expressed exclusively in the brain and linked to seizures, neuroprotection and cardiovascular disease. Based upon the observation that fragments of the GPR37L1 N-terminus are found in human cerebrospinal fluid, we hypothesized that GPR37L1 was subject to post-translational modification. Heterologous expression of GPR37L1-eYFP in either HEK293 or U87 glioblastoma cells yielded two cell surface species of approximately equivalent abundance, the larger of which is N-glycosylated at Asn(105). The smaller species is produced by matrix metalloprotease/ADAM-mediated proteolysis (shown by the use of pharmacological inhibitors) and has a molecular weight identical to that of a mutant lacking the entire N-terminus, Delta122 GPR37L1. Serial truncation of the N-terminus prevented GPR37L1 expression except when the entire N-terminus was removed, narrowing the predicted site of N-terminal proteolysis to residues 105-122. Using yeast expressing different G protein chimeras, we found that wild type GPR37L1, but not Delta122 GPR37L1, coupled constitutively to Gpa1/Galphas and Gpa1/Galpha16 chimeras, in contrast to previous studies. We tested the peptides identified in cerebrospinal fluid as well as their putative newly-generated N-terminal 'tethered' counterparts in both wild type and Delta122 GPR37L1 Gpa1/Galphas strains but saw no effect, suggesting that GPR37L1 does not signal in a manner akin to the protease-activated receptor family. We also saw no evidence of receptor activation or regulation by the reported GPR37L1 ligand, prosaptide/TX14A. Finally, the proteolytically processed species predominated both in vivo and ex vivo in organotypic cerebellar slice preparations, suggesting that GPR37L1 is rapidly processed to a signaling-inactive form. Our data indicate that the function of GPR37L1 in vivo is tightly regulated by metalloprotease-dependent N-terminal cleavage.
| Item Type: | Article |
|---|---|
| Subjects: | R Medicine > R Medicine (General) |
| Depositing User: | Repository Administrator |
| Date Deposited: | 11 Oct 2021 03:40 |
| Last Modified: | 11 Oct 2021 03:40 |
| URI: | http://eprints.victorchang.edu.au/id/eprint/1136 |
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