Enrichment of the inositol 1,4,5-trisphosphate receptor/Ca²⁺ channels in secretory granules and essential roles of chromogranins

Abstract

The high capacity, low affinity Ca²⁺ storage protein chromogranins are marker proteins of secretory granules that contain the most Ca²⁺ in secretory cells. Along with the abundantly expressed chromogranins, the IP₃R/Ca²⁺ channels, the major intracellular Ca²⁺ channels, are also expressed in secretory granules the most. Chromogranins not only induce formation of secretory granules but also are suggested to produce the small IP₃-sensitive nucleoplasmic Ca²⁺ store vesicles in the nucleus. Chromogranins A (CGA) and B (CGB) also directly bind the IP₃Rs and activate the IP₃R/Ca²⁺ channels at the intragranular pH 5.5. But at a near physiological pH 7.5 only CGB interacts with the IP₃Rs due to stronger interaction of CGB for the IP₃Rs, which is several orders of magnitude stronger than that of CGA, and activates the IP₃R/Ca²⁺ channels. Therefore, the CGB-IP₃R coupling is proposed to play key roles in the IP₃-mediated Ca²⁺ signaling mechanisms in the cytoplasm through both secretory granules and the ER, and in the nucleus through the small IP₃-sensitive nucleoplasmic Ca²⁺ store vesicles. Chromogranin B is further suggested to participate in transcription control and to target secretory granule components, including the IP₃Rs, to newly formed secretory granules. Defects in secretory granule-related functions are directly linked to major human diseases such as Alzheimer's disease, secretory cell cancers, cystic fibrosis, acute pancreatitis, and cardiac hypertrophy. Therefore, realization of secretory granules as the major intracellular Ca²⁺ storage and control organelle in secretory cells promises to open new horizon in understanding the Ca²⁺ storage, signaling, and control mechanisms throughout the biokingdom.

Introduction

Contrary to the conventional thought that secretory granules merely store a variety of hormones and bioactive molecules and release to the extracellular space, recent studies unraveled far more complex and sophisticated structural and functional characteristics of secretory granules, which demonstrated their crucial roles in the storage and control of intracellular concentrations of Ca²⁺ [1]. There are several intracellular organelles that participate in the storage and control of Ca²⁺ in the cell, i.e. the endoplasmic reticulum (ER) [2], lysosomes [3], endosomes [4], [5], mitochondria [6], [7], the Golgi [8], secretory granules [1], and the nucleus [9], [10].

Of these organelles, secretory granules contain by far the largest amount of cellular Ca²⁺ and the highest concentration of total (bound + free) Ca²⁺, ranging ∼15–40 mM or higher [5], [11], [12], [13], in a myriad of secretory cells across the biokingdom. Befitting the role of major intracellular Ca²⁺ storage and control organelle, secretory granules of virtually all secretory cells are loaded with large amounts of acidic Ca²⁺ storage proteins [14], [15], [16], [17], [18], [19] in an intragranular pH of ∼5.5, conditions that enable secretory granules to store such extraordinary amounts of Ca²⁺. In secretory granules of mammals chromogranins and secretogranins are the representative acidic Ca²⁺ storage proteins although mucins are also abundant in some cells such as airway epithelial and mast cells [20], [21], [22]. Further, chromogranin-like proteins and polyanionic polymers are widely localized in secretory granules of various organisms, ranging from species as primitive as phytoplankton to high vertebrates [23], [24], [25].

In addition, secretory granules contain a variety of ion channels, the IP₃R/Ca²⁺ channel being the most prominent one [26], [27], [28], and function as the major IP₃-sensitive intracellular Ca²⁺ stores in secretory cells [1], [29]. Besides the IP₃R/Ca²⁺ channels, ryanodine receptor (RyR)/Ca²⁺ channels [30], [31], Cl⁻ channels, H⁺-ATPase pump, Na⁺/Ca²⁺, Ca²⁺/H⁺, K⁺/H⁺, and Na⁺/H⁺ exchangers are known to exist [5], [20], [32], [33], [34], [35], [36]. In particular, Ca²⁺-activated apamin-sensitive K⁺ channels (ASKCa) are proposed to be essential in the function of secretory granules as the IP₃-sensitive Ca²⁺ store and as ion oscillators in the cytoplasm [20], [21], [22], [24]. The IP₃-induced Ca²⁺ release from secretory granules is thought to activate the ASKCa channels to transport K⁺ into the granules. The K⁺ transported into the granules exchanges with the matrix bound Ca²⁺, thus increasing the once low free Ca²⁺ concentrations inside the granules. These exchanges, combined with the IP₃-induced Ca²⁺ release and the Ca²⁺ transport via the granule membrane Ca²⁺-ATPase, are suggested to be primarily responsible for the oscillatory changes of Ca²⁺ concentrations in and out of the granules [20], [21], [22], [24]. Therefore, inhibition of the IP₃-induced Ca²⁺ release from secretory granules has been shown to inhibit the oscillatory changes of Ca²⁺ concentrations [21], [22], [24], [37].

In light of the fact that the main function of secretory cells in organisms is to store and release many essential biomolecules at the time and place needed inside the organisms, it is of extraordinary consequence that secretory granules the very organelle that stores hormones and other key biomolecules are the ones that also carry the signal (Ca²⁺) for secretion themselves. Furthermore secretory granules are also equipped with membrane proteins that participate in Ca²⁺ sensing as well as in fusion pore formation. Synaptotagmin and syntaxin that play pivotal roles in fusion pore formation and exocytosis are richly expressed on the membranes of secretory granules [38], [39], [40] and interact with the IP₃R/Ca²⁺ channels, which in turn are tightly bound to the intragranular chromogranins A (CGA) and B (CGB) [40]. The coupling of intragranular chromogranins with the membrane protein IP₃R/Ca²⁺ channels not only endows the secretory granule IP₃R/Ca²⁺ channels with high IP₃ sensitivity but also enables direct and highly sensitive communication between the intragranular contents and the extragranular environment that contains different concentrations of Ca²⁺, IP₃, and modulatory molecules.