Doc2b Ca2+-binding site mutants act as a gain of function at rest and loss of function during neuronal activity

Bourgeois-Jaarsma, Q.; Verhage, M.; Groffen, A. J.

doi:10.1101/536581

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bioRxiv · 10.1101/536581

Doc2b Ca2+-binding site mutants act as a gain of function at rest and loss of function during neuronal activity

Bourgeois-Jaarsma, Q.·Verhage, M.·Groffen, A. J.

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Abstract

Communication between neurons involves presynaptic neurotransmitter release which can be evoked by action potentials or occur spontaneously as a result of stochastic vesicle fusion. The Ca2+-binding double C2 proteins Doc2a and -b regulate both spontaneous and asynchronous evoked release, but the mechanism remains unclear. Here, we compared wildtype Doc2b with two Ca2+ binding site mutants named DN and 6A, respectively considered gain-and loss-of function mutants and carrying the substitutions D218,220N or D163,218,220,303,357,359A. We found that both mutants bound phospholipids at low free Ca2+ concentrations and were membrane-associated in neurons at rest, mimicking a Ca2+ activated state. Their overexpression in hippocampal primary neurons culture had similar effects on spontaneous and evoked release, inducing higher mEPSC frequencies and increased short-term depression. Together, these data suggest that the DN and 6A mutants both act as gain-of-function mutants at resting conditions but as loss-of-function during neuronal activity.

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Bourgeois-Jaarsma, Q., Verhage, M., Groffen, A. J.. 2019-01-31. Doc2b Ca2+-binding site mutants act as a gain of function at rest and loss of function during neuronal activity. https://doi.org/10.1101/536581

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