Differential calcium handling by the cis and trans regions of the Golgi apparatus

High Ca²⁺ content in the Golgi apparatus (Go) is essential for protein processing and sorting. In addition, the Go can shape the cytosolic Ca²⁺ signals by releasing or sequestering Ca²⁺. We generated two new aequorin-based Ca²⁺ probes to specifically measure Ca²⁺ in the cis/cis-to-medial-Go (cGo) or the trans-Go (tGo). Ca²⁺ homoeostasis in these compartments and in the endoplasmic reticulum (ER) has been studied and compared. Moreover, the relative size of each subcompartment was estimated from aequorin consumption. We found that the cGo accumulates Ca²⁺ to high concentrations (150–300 μM) through the sarco plasmic/endoplasmic reticulum Ca²⁺-ATPase (SERCA). The tGo, in turn, is divided into two subcompartments: tGo1 and tGo2. The subcompartment tGo1 contains 20% of the aequorin and has a high internal [Ca²⁺]; Ca²⁺ is accumulated in this subcompartment via the secretory pathway Ca²⁺-ATPase 1 (SPCA-1) at a very high affinity (K₅₀=30 nM). The subcompartment tGo2 contains 80% of aequorin, has a lower [Ca²⁺] and no SPCA-1 activity; Ca²⁺ uptake happens through SERCA and is slower than in tGo1. The two tGo subcompartments, tGo1 and tGo2, are diffusionally isolated. Inositol trisphosphate mobilizes Ca²⁺ from the cGo and tGo2, but not from tGo1, whereas caffeine releases Ca²⁺ from all the Golgi regions, and nicotinic acid dinucleotide phosphate and cADP ribose from none.