Elucidating the molecular logic of a metabotropic glutamate receptor heterodimer

Metabotropic glutamate (mGlu) receptor protomers can heterodimerize, leading to different pharmacology compared to their homodimeric counterparts. Here, we use complemented donor-acceptor resonance energy transfer (CODA-RET) technology that distinguishes signaling from defined mGlu heterodimers or homodimers, together with targeted mutagenesis of receptor protomers and computational docking, to elucidate the mechanism of activation and differential pharmacology in mGlu_2/4 heteromers. We demonstrate that positive allosteric modulators (PAMs) that bind an upper allosteric pocket in the mGlu₄ transmembrane domain are active at both mGlu_4/4 homomers and mGlu_2/4 heteromers, while those that bind a lower allosteric pocket within the same domain are efficacious in homomers but not heteromers. We further demonstrate that both protomers of mGlu_2/4 heteromers are cis-activated by their orthosteric agonists, signaling independently with no trans-activation detected. Intriguingly, however, upper pocket mGlu₄ PAMs enable trans-activation in mGlu_2/4 heteromers from mGlu₄ to the mGlu₂ protomer and also enhance cis-activation of the mGlu₂ protomer. While mGlu₂ PAMs enhanced mGlu₂ cis-activation in the heterodimer, we were unable to detect trans-activation in the opposite direction from mGlu₂ to the mGlu₄ protomer, suggesting an asymmetry of signaling. These insights into the molecular logic of this receptor heteromer are critical to building toward precision targeted therapies for multiple neuropsychiatric disorders.