Measuring steady-state and dynamic endoplasmic reticulum and Golgi Zn2+ with genetically encoded sensors

Zn2+ plays essential roles in biology, and cells have adopted exquisite mechanisms for regulating steady-state Zn2+ levels. Although much is known about total Zn2+ in cells, very little is known about its subcellular distribution. Yet defining the location of Zn2+ and how it changes with signaling events is essential for elucidating how cells regulate this essential ion. Here we create fluorescent sensors genetically targeted to the endoplasmic reticulum ( ER) and Golgi to monitor steady-state Zn2+ levels as well as flux of Zn2+ into and out of these organelles. These studies reveal that ER and Golgi contain a concentration of free Zn2+ that is 100 times lower than the cytosol. Both organelles take up Zn2+ when cytosolic levels are elevated, suggesting that the ER and Golgi can sequester elevated cytosolic Zn2+ and thus have the potential to play a role in influencing Zn2+ toxicity. ER Zn2+ homeostasis is perturbed by small molecule antagonists of Ca2+ homeostasis and ER Zn2+ is released upon elevation of cytosolic Ca2+ pointing to potential exchange of these two ions across the ER. This study provides direct evidence that Ca2+ signaling can influence Zn2+ homeostasis and vice versa, that Zn2+ dynamics may modulate Ca2+ signaling.