Signaling networks are spatiotemporally organized in order to sense diverse inputs, process information, and carry out specific cellular tasks. In pancreatic β cells, Ca²⁺, cyclic adenosine monophosphate (cAMP), and Protein Kinase A (PKA) exist in an oscillatory circuit characterized by a high degree of feedback. Here, we describe a mode of regulation within this circuit involving a spatial dependence of the relative phase between cAMP, PKA, and Ca²⁺. We show that nanodomain clustering of Ca²⁺-sensitive adenylyl cyclases drives oscillations of local cAMP levels to be precisely in-phase with Ca²⁺ oscillations, whereas Ca²⁺-sensitive phosphodiesterases maintain out-of-phase oscillations outside of the nanodomain. Disruption of this precise phase relationship perturbs Ca²⁺ oscillations, suggesting the relative phase within an oscillatory circuit can encode specific functional information. This work unveils a novel mechanism of cAMP compartmentation utilized for localized tuning of an oscillatory circuit and has broad implications for the spatiotemporal regulation of signaling networks.

MIN6 pancreatic beta cells expressing FRET-based biosensors were imaging using an epifluorescent microscope. FRET ratio was calculated using sensitized emission channel / donor excited channel (PKA sensor) or vice versa for cAMP (inverse FRET sensor). Single-cell time traces were collected and analyzed with MATLAB to study compartmentalized oscillatory dynamics.

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