Abstract
Brain-specific angiogenesis inhibitor-1 (BAI1) is an adhesion G protein-coupled receptor that has been studied primarily for its anti-angiogenic and anti-tumorigenic properties. We found that overexpression of BAI1 results in activation of the Rho pathway via a Gα(12/13)-dependent mechanism, with truncation of the BAI1 N terminus resulting in a dramatic enhancement in receptor signaling. This constitutive activity of the truncated BAI1 mutant also resulted in enhanced downstream phosphorylation of ERK as well as increased receptor association with β-arrestin2 and increased ubiquitination of the receptor. To gain insights into the regulation of BAI1 signaling, we screened the C terminus of BAI1 against a proteomic array of PDZ domains to identify novel interacting partners. These screens revealed that the BAI1 C terminus interacts with a variety of PDZ domains from synaptic proteins, including MAGI-3. Removal of the BAI1 PDZ-binding motif resulted in attenuation of receptor signaling to Rho but had no effect on ERK activation. Conversely, co-expression with MAGI-3 was found to potentiate signaling to ERK by constitutively active BAI1 in a manner that was dependent on the PDZ-binding motif of the receptor. Biochemical fractionation studies revealed that BAI1 is highly enriched in post-synaptic density fractions, a finding consistent with our observations that BAI1 can interact with PDZ proteins known to be concentrated in the post-synaptic density. These findings demonstrate that BAI1 is a synaptic receptor that can activate both the Rho and ERK pathways, with the N-terminal and C-terminal regions of the receptor playing key roles in the regulation of BAI1 signaling activity.
Highlights
Brain-specific angiogenesis inhibitor-1 (BAI1) is an adhesion receptor; little is known about its signaling or localization
BAI1 has been studied for its anti-angiogenic and anti-tumorigenic properties in glial cells (5, 13, 17) as well as its ability to facilitate the engulfment of apoptotic cells (10) and induce myogenesis (19), but little is known about the signaling pathways by which BAI1 can mediate its various physiological actions
These findings place BAI1 on the short list of adhesion G protein-coupled receptors (GPCRs) for which coupling to G proteins has been documented: GPR56 (22, 27) and CD97 (28) have been shown to activate Rho via G␣12/13, latrophilin-1 has been shown to couple to G␣q and G␣o (37, 38), GPR133 (39) and GPR114 (40) have been shown to increase cAMP production through coupling to G␣s, and GPR97 has been shown to couple to G␣o (40)
Summary
BAI1 is an adhesion receptor; little is known about its signaling or localization. Results: BAI1 activates Rho in a G protein-dependent manner, binds to synaptic scaffold proteins, and is highly enriched in the postsynaptic density. Co-expression with MAGI-3 was found to potentiate signaling to ERK by constitutively active BAI1 in a manner that was dependent on the PDZ-binding motif of the receptor. Biochemical fractionation studies revealed that BAI1 is highly enriched in post-synaptic density fractions, a finding consistent with our observations that BAI1 can interact with PDZ proteins known to be concentrated in the post-synaptic density These findings demonstrate that BAI1 is a synaptic receptor that can activate both the Rho and ERK pathways, with the N-terminal and C-terminal regions of the receptor playing key roles in the regulation of BAI1 signaling activity. The BAI1 C terminus has been reported to bind to two PDZ domain-containing scaffold proteins (20, 21), but nothing is known at present about the functional significance of these interactions. These studies led to a series of novel insights about the signaling, regulation, and subcellular localization of BAI1 and new ideas about the potential physiological importance of BAI1 in vivo
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