GPR158, an Orphan Member of G Protein-Coupled Receptor Family C: Glucocorticoid-Stimulated Expression and Novel Nuclear Role

Abstract

Members of the large G protein-coupled receptor (GPCR) clan are implicated in many physiological and disease processes, making them important therapeutic drug targets. In the present study, we follow up on results of a pilot study suggesting a functional relationship between glucocorticoid (GC)-induced ocular hypertension and GPR158, one of three orphan members of the GPCR Family C. GC treatment increases levels of GPR158 mRNA and protein through transcriptional mechanisms, in cultured trabecular meshwork (TBM) cells derived from the eye's aqueous outflow pathway. Like treatment with GCs, transient overexpression of GPR158 stimulates cell proliferation, while siRNA knockdown of endogenous GPR158 has the opposite effect. Both endogenous and overexpressed GPR158 show an unusual subcellular localization pattern, being found almost entirely in the nucleus. However, when cells are treated with inhibitors of clathrin-mediated endocytosis, GPR158 is shifted to the plasma membrane. Mutation of a bipartite nuclear localization signal (NLS) in the 8th helix also shifts GPR158 out of the nucleus, but in this case the protein is found in vesicles localized in the cytoplasm. These results suggest that newly synthesized GPR158 first traffics to the plasma membrane, where it rapidly undergoes endocytosis and translocation to the nucleus. Significantly, mutation of the NLS abrogates GPR158-mediated enhancement of cell proliferation, indicating a functional requirement for nuclear localization. GPR158 overexpression upregulates levels of the cell cycle regulator cyclin D1, but mutation of the NLS reverses this. Overexpression of GPR158 enhances the barrier function of a TBM cell monolayer, which is associated with an increase in the levels of tight junction proteins ZO-1 and occludin, similar to reported studies on GC treatment. Regulated paracellular permeability controls aqueous outflow facility in vivo. Since GCs stimulate GPR158 expression, the result is consistent with a role for elevation of GPR158 expression in GC-induced ocular hypertension.

Orphan Receptor GPR158 Is an Allosteric Modulator of RGS7 Catalytic Activity with an Essential Role in Dictating Its Expression and Localization in the Brain

“…It is also possible that GPR158 might have an additional G protein-independent mode in regulating cellular signaling. For example, a C-terminal fragment of GPR158 was shown to contain a nuclear localization sequence and play an important role in directing GPR158 in the nucleus to regulate cellular proliferation (77). However, regardless of whether or not GPR158/179 serve as GPCRs and/or affect signaling via an alternative mechanism, it is clear that they can impact cellular homeostasis by affecting the activity, localization, and expression of RGS7.…”

Integrative Genomic Approach Identifies Multiple Genes Involved in Cardiac Collagen Deposition

“…Gpr158, a relatively unstudied orphan member of class C of the large G-protein coupled receptor family, was recently implicated in the formation of tight junctions in ocular trabecular meshwork cells. 26 To our knowledge, no connection between Gpr158 and either fibrosis or cardiac function has been described to date. …”

Intranuclear Signaling Cascades Triggered by Nuclear GPCRs

“…It is expressed in a variety of tissues, but it is expressed at highest levels in the brain. In cultured trabecular meshwork cells, glucocorticoid (GC) treatment and the transient overexpression of GPR158 stimulates levels of GPR158 mRNA and protein through transcriptional mechanisms [36]. Both endogenous and overexpressed GPR158 show an unusual subcellular localization pattern, being found almost entirely in the nucleus.…”

“…Both endogenous and overexpressed GPR158 show an unusual subcellular localization pattern, being found almost entirely in the nucleus. The transient overexpression of GPR158 stimulates cell proliferation, up-regulates levels of the cell cycle regulator cyclin D1 and increases the levels of tight junction proteins ZO-1 (Table 1) [36]. Nuclear localization of endogenous and overexpressed GPR158 is prevented by inhibitors of clathrin-mediated endocytosis.…”

“…This suggests that GPR158 traffics to the plasma membrane, where it rapidly undergoes endocytosis and translocation to the nucleus. A bipartite NLS has been identified in the eighth helix, at the proximal end of the C-terminal cytoplasmic tail of the receptor (Table 2) [36].…”