Calmodulin-controlled Spatial Decoding of Oscillatory Ca 2 + Signals by 1 Calcineurin 2 3 4

36 Calcineurin is responsible for mediating a wide variety of cellular processes in response 37 to dynamic calcium (Ca) signals, yet the precise mechanisms involved in the spatiotemporal 38 control of calcineurin signaling are poorly understood. Here, we use genetically encoded 39 fluorescent biosensors to directly probe the role of cytosolic Ca oscillations in modulating 40 calcineurin activity dynamics in insulin-secreting MIN6 β-cells. We show that Ca oscillations 41 induce distinct temporal patterns of calcineurin activity in the cytosol and plasma membrane 42 versus at the ER and mitochondria in these cells. Furthermore, we found that these differential 43 calcineurin activity patterns are determined by variations in the subcellular distribution of 44 calmodulin (CaM), indicating that CaM plays an active role in shaping both the spatial and 45 temporal aspects of calcineurin signaling. Together, our findings provide new insights into the 46 mechanisms by which oscillatory signals are decoded to generate specific functional outputs 47 within different cellular compartments. 48