The β- and γ-isoforms of type I PIP5K regulate distinct stages of Ca2+ signaling in mast cells

Abstract

Crosslinking of IgE receptors by antigen initiates Ca2+ mobilization in mast cells by activating phospholipase-Cγ-mediated hydrolysis of phosphatidylinositol-4,5-bisphosphate [PtdIns(4,5)P2]. The resulting inositol 1,4,5-trisphosphate-mediated Ca2+ release from the endoplasmic reticulum (ER) activates store-operated Ca2+ entry, which is necessary for exocytotic release of inflammatory mediators. To investigate roles for PtdIns(4,5)P2-synthesizing isozymes of the type I phosphatidylinositol 4-phosphate 5-kinase family (PIP5K-I) in mast cell signaling, we compared the ectopic expression of wild-type and catalytically inactive PIP5K-Iβ in RBL-2H3 mast cells. Surprisingly, both antigen and thapsigargin-stimulated Ca2+ influx were reduced by overexpression of active PIP5K-Iβ, whereas antigen-stimulated Ca2+ release from ER stores was unaffected. Consistent with these results, Ca2+ entry stimulated by antigen or thapsigargin was enhanced by expression of a plasma-membrane-associated inositol polyphosphate 5′-phosphatase, whereas antigen-stimulated Ca2+ release from stores was reduced. To investigate the role of PIP5K-Iγ in antigen-stimulated Ca2+ mobilization, we used bone-marrow-derived mast cells from PIP5K-Iγ–/– mice. Antigen-stimulated Ca2+ release from ER stores was substantially reduced in the absence of PIP5K-Iγ, but thapsigargin-mediated Ca2+ entry was unaffected. In summary, PIP5K-Iγ positively regulates antigen-stimulated Ca2+ release from ER stores, whereas PIP5K-Iβ negatively regulates store-operated Ca2+ entry, suggesting that these different PIP5K-I isoforms synthesize functionally distinct pools of PtdIns(4,5)P2 at the plasma membrane.

Lovastatin-Induced Phosphatidylinositol-4-Phosphate 5-Kinase Diffusion from Microvilli Stimulates ROMK Channels

“…The disassociation of PI(4)P5K I g from prominin-1 is not caused by a possible diffusion of prominin-1, because Western blots showed that the majority of prominin-1 was still located in microvilli (Figure 7B). Because PI(4)P5K I g is a major kinase that phosphorylates PI(4)P at five positions to produce PI(4,5)P 2 , 32 these data indicate that lovastatin may regulate ROMK1 channels by inducing PI(4)P5K I g diffusion into planar regions to initiate PI (4,5)P 2 synthesis near the channel.…”

Stimulated association of STIM1 and Orai1 is regulated by the balance of PtdIns(4,5)P2 between distinct membrane pools

“…We previously demonstrated that different isoforms of type I phosphatidylinositol 4-phosphate 5-kinase (PIP5KI), namely PIP5KI and PIP5KI, synthesize functionally distinguishable pools of PtdIns(4,5)P 2 that have distinct roles in SOCE and inositol trisphosphate (Ins(1,4,5)P 3 ) generation, respectively (Vasudevan et al, 2009). In the present study we characterize a membrane structural basis for distinctive pools of PtdIns(4,5)P 2 .…”

Comparative phenotypic analysis of the two major splice isoforms of phosphatidylinositol phosphate kinase type Iγ in vivo

“…PLC cleaves PtdIns(4,5)P 2 into DAG and IP 3 ; the latter binds to receptors on the endoplasmic reticulum and stimulates the release of Ca 2+ (Berridge, 2009). Knockdown or genetic ablation of PIPKIc results in diminished intracellular [Ca 2+ ] changes in HeLa cells, mast cells and mesenchymal stem cells (Wang et al, 2004;Vasudevan et al, 2009;Yu et al, 2011), implicating PIPKIc as the source of this pool of PtdIns(4,5)P 2 . Cationic amino acid transporter (CAT-2) knockout mice provide indirect in vivo evidence for PIPKIc involvement in Ca 2+ signaling in smooth muscle (Chen et al, 2007).…”

“…A role for PIPKIc in agonist-induced Ca 2+ signaling is supported by knockdown and knockout studies in several cell types (Wang et al, 2004;Vasudevan et al, 2009;Yu et al, 2011). To examine whether there are PIPKIc isoform-specific differences in Ca 2+ signaling in our mice, we used the ratiometric Fura-2 Ca 2+ indicator dye to monitor ATP-induced intracellular Ca 2+ release in WT and PIPKIc 2/2 fibroblasts.…”

“…Interestingly, a single allele of PIPKIc is sufficient for life (Volpicelli-Daley et al, 2010). Cells derived from the perinatal lethal PIPKIc mice have additionally been used to establish roles for PIPKIc in vesicle priming in chromaffin cells (Gong et al, 2005), FccR-mediated phagocytosis (Mao et al, 2009), Ca 2+ signaling in mast cells (Vasudevan et al, 2009) and adhesion and infiltration of neutrophils (Xu et al, 2010), whereas megakaryocytes derived from the embryonic lethal PIPKIc mouse have implicated PIPKIc in mediating attachment of the actin cytoskeleton to the plasma membrane in this cell type (Wang et al, 2008b).…”