Optical Control of Metabotropic Glutamate Receptors

G-protein coupled receptors (GPCRs), the largest family of membrane signaling proteins, respond to neurotransmitters, hormones and small environmental molecules. The neuronal function of many GPCRs has been difficult to resolve because of an inability to gate them with subtype-specificity, spatial precision, speed and reversibility. To address this, we developed an approach for opto-chemical engineering native GPCRs. We applied this to the metabotropic glutamate receptors (mGluRs) to generate light-agonized and light-antagonized “LimGluRs”. The light-agonized “LimGluR2”, on which we focused, is fast, bistable, and supports multiple rounds of on/off switching. Light gates two of the primary neuronal functions of mGluR2: suppression of excitability and inhibition of neurotransmitter release. The light-antagonized “LimGluR2block” can be used to manipulate negative feedback of synaptically released glutamate on transmitter release. We generalize the optical control to two additional family members: mGluR3 and 6. The system works in rodent brain slice and in zebrafish in vivo, where we find that mGluR2 modulates the threshold for escape behavior. These light-gated mGluRs pave the way for determining the roles of mGluRs in synaptic plasticity, memory and disease

Directed synthesis of all four pure stereoisomers of the N,C-Coupled naphthylisoquinoline alkaloid ancistrocladinium A

The first preparation of the N,C-coupled naphthylisoquinoline alkaloid ancistrocladinium A and its likewise naturally occurring minor atropisomer, in an atropisomerically pure form, is described. The synthesis succeeded by resolution of the already rotationally hindered, and thus atropo-diastereomeric acetamide precursors, which were then, without major loss of stereochemical information, cyclized to the respective target molecules. The strategy was applied to the first synthesis of the regioisomeric product ancistrocladinium D, likewise in a stereochemically pure form

Directed Synthesis of All Four Pure Stereoisomers of the <i>N</i>,<i>C</i>‑Coupled Naphthylisoquinoline Alkaloid Ancistrocladinium A

The first preparation of the <i>N</i>,<i>C</i>-coupled naphthylisoquinoline alkaloid ancistrocladinium A and its likewise naturally occurring minor atropisomer, in an atropisomerically pure form, is described. The synthesis succeeded by resolution of the already rotationally hindered, and thus atropo-diastereomeric acetamide precursors, which were then, without major loss of stereochemical information, cyclized to the respective target molecules. The strategy was applied to the first synthesis of the regioisomeric product ancistrocladinium D, likewise in a stereochemically pure form

A Mechanical Switch Couples T Cell Receptor Triggering to the Cytoplasmic Juxtamembrane Regions of CD3ζζ

The eight-subunit T cell receptor (TCR)-CD3 complex is the primary determinant for T cell fate decisions. Yet how it relays ligand-specific information across the cell membrane for conversion to chemical signals remains unresolved. We hypothesized that TCR engagement triggers a change in the spatial relationship between the associated CD3ζζ subunits at the junction where they emerge from the membrane into the cytoplasm. Using three in situ proximity assays based on ID-PRIME, FRET, and EPOR activity, we determined that the cytosolic juxtamembrane regions of the CD3ζζ subunits are spread apart upon assembly into the TCR-CD3 complex. TCR engagement then triggered their apposition. This mechanical switch resides upstream of the CD3ζζ intracellular motifs that initiate chemical signaling, as well as the polybasic stretches that regulate signal potentiation. These findings provide a framework from which to examine triggering events for activating immune receptors and other complex molecular machines.National Institutes of Health (U.S.) (5R01CA186568