Evolutionary optimization of allosteric activation by Cl- and Cl- conduction in vesicular glutamate transporters

Abstract

Vesicular glutamate transporters harness proton gradients to load glutamate into synaptic vesicles, while mediating luminal chloride efflux through a channel-like conductance. We combined heterologous expression in mammalian cells, whole-cell patch-clamp recordings and mathematical modeling to functionally characterize the Drosophila melanogaster vesicular glutamate transporter DVGLUT and to compare it to a rat counterpart, rVGLUT1. As in mammalian VGLUTs, DVGLUT glutamate transport is coupled to proton exchange, in a 1:1 stoichiometry. In both, luminal Cl- is necessary as allosteric activator, however, the allosteric affinity is higher in fly than in rat transporters. DVGLUT anion channels exhibit lower unitary currents, but higher anion channel open probabilities, resulting in larger Cl- currents for the fly transporter in the presence of cytoplasmic glutamate. The higher allosteric affinity together with the enhanced anion channel activity may serve as evolutionary adaptation of VGLUTs to lower ion concentrations in Drosophila, illustrating the impact of these particular features for synaptic vesicle filling.