ERK-Mediated Decoding of Calcium Oscillations in Pancreatic Beta Cells

Abstract

Calcium oscillations are thought to underlie pulsatility of insulin secretion from pancreatic beta-cells, which is necessary for maintaining glucose homeostasis. However, it is still unclear how this calcium signal is ‘decoded’ by other signaling molecules such as protein kinases to modulate functional responses in a context-dependent manner. In this study, we investigated crosstalk between calcium and ERK signaling pathways using FRET-based ERK biosensor, EKAR, and RFP-based calcium biosensor, RCaMP, in live MIN6 pancreatic beta-cells.We observed that different physiological stimuli elicit different dynamics of ERK activity potentially affecting cell function. Amino acid stimulation resulted in simultaneous transient increase in calcium and ERK activity. Membrane-depolarizing agents, however, resulted in calcium oscillations leading to sustained elevation of ERK activity. Further, we observed that artificial imposition of a train of calcium pulses mimicking “low-frequency oscillations” lead to simultaneous low-frequency ERK activity pulses. These results, together with model simulations, suggest that endogenous frequency of calcium oscillations in pancreatic beta-cells may be optimized for sustained ERK activation, potentially leading to efficient insulin gene transcription.Calcium oscillations were also found to lead to sustained activation of Ras, an upstream signaling molecule in the ERK cascade, indicating that “processing” of oscillatory calcium signal was likely mediated by calcium-sensitive RasGEFs and RasGAPs. We show here that calcium-sensitive RasGRF1, RasGRF2 and p120RasGAP are expressed in MIN6 cells which could thus dictate the temporal properties of the signal that is communicated to ERK. Further, we noticed that inhibition of ERK activity resulted in loss of calcium oscillations indicating feedback regulation of input calcium signal by ERK. The presence of this putative feedback loop together with calcium-dependent modulation of ERK activity highlights the intricate cross-regulation of the calcium and ERK signaling pathways in pancreatic beta-cells for precise modulation of functional responses.