Interaction of Neuronal Calcium Sensor-1 and ADP-ribosylation Factor 1 Allows Bidirectional Control of Phosphatidylinositol 4-Kinase β and trans-Golgi Network-Plasma Membrane Traffic

Abstract

We have identified a novel Ca 2؉ -dependent interaction between neuronal calcium sensor-1 (NCS-1) and the GTPase ARF1. Both of these proteins are localized to the Golgi complex, and both regulate phosphatidylinositol 4-kinase III␤ (PI(4)K␤). Spatial and temporal control of phosphatidylinositol 4-phosphate levels through activation of PI(4)K␤ is important for the recruitment of trafficking complexes to the trans-Golgi network (TGN) and vesicular traffic from this organelle. The NCS-1-ARF1 interaction and its specificity have been demonstrated through in vitro binding assays, in vitro enzyme assay, and through functional cellular assays. We show that NCS-1 can exert bidirectional effects to activate PI(4)K␤ on its own or inhibit the activation by ARF1. NCS-1 was shown to modulate the effects of expression of ARF mutants that disrupt Golgi morphology and to recruit GDPloaded ARF to the Golgi complex in a Ca 2؉ -dependent manner. We demonstrate antagonist effects of NCS-1 and ARF on constitutive and regulated exocytosis. The NCS-1-ARF1 interaction provides evidence for functional cross-talk between Ca 2؉ -dependent and ARF-dependent pathways in TGN to plasma membrane traffic.

Calneurons provide a calcium threshold for trans -Golgi network to plasma membrane trafficking

“…To next address the question of which functional consequences calneuron-1 binding might have for PI-4K␤'s enzymatic activity we performed in vitro kinase assays with bacterially-expressed proteins. Conflicting evidence exists whether myristoylated NCS-1 activates PI-4K␤ in vitro in a Ca 2ϩ -dependent manner and whether the interaction by itself is Ca 2ϩ -independent (11,19). We found that the basal activity of the kinase was unaltered in the presence of myristoylated NCS-1 when Ca 2ϩ was omitted from the buffer (Fig.…”

“…Moreover, its binding to PI-4K␤ seems to be of lower affinity as compared with the yeast proteins (11). It is therefore likely that the regulation of Pik1 and PI-4K␤ differs substantially with the latter being more susceptible to modulation via Ca 2ϩ transients at the Golgi (19).…”

Phospholipid Synthesis Participates in the Regulation of Diacylglycerol Required for Membrane Trafficking at the Golgi Complex

“…Recently, it has been shown that PITP␤ is needed for the vesicular transport at the Golgi (20). In this case, the authors suggested that PITP␤ is needed to bring PI to the Golgi for the synthesis of phosphatidylinositol 4-phosphate, which is involved in vesicular trafficking at the Golgi (21).…”

The Diversity of Calcium Sensor Proteins in the Regulation of Neuronal Function

“…NCS-1 can also interact with the mammalian Golgi enzyme PI4KIIIb and enhances its activity three-to 10-fold (Taverna et al 2002;Haynes et al 2005;de Barry et al 2006). The interaction with Golgi-associated PI4KIIIb suggests that it may regulate secretion through the modulation of phosphatidylinositol-dependent trafficking steps (Hendricks et al 1999;Zhao et al 2001;Haynes et al 2005). In support of this, NCS-1 has also been demonstrated to associate with another PI4KIIIb regulator ARF1, a small GTPase critical to multiple trafficking steps in mammalian cells Haynes et al 2007).…”