Rat Inositol 1,4,5-Trisphosphate 3-Kinase C Is Enzymatically Specialized for Basal Cellular Inositol Trisphosphate Phosphorylation and Shuttles Actively between Nucleus and Cytoplasm

Abstract

The calcium-liberating second messenger inositol 1,4,5-trisphosphate (Ins(1,4,5)P 3 ) is converted to inositol 1,3,4,5-tetrakisphosphate (Ins(1,3,4,5)P 4 ) by Ins(1,4,5)P 3 3-kinases (IP3Ks) that add a fourth phosphate group to the 3-position of the inositol ring. Two isoforms of IP3Ks (named A and B) from different vertebrate species have been well studied. Recently the cloning and examination of a human full-length cDNA encoding a novel isoform, termed human IP3K-C (HsIP3K-C), has been reported. In the present study we report the cloning of a full-length cDNA encoding a rat homologue of HsIP3K-C with a unique mRNA expression pattern, which differs remarkably from the tissue distribution of HsIP3K-C. Of the rat tissues examined, rat IP3K-C (RnIP3K-C) is mainly present in heart, brain, and testis and shows the strongest expression in an epidermal tissue, namely tongue epithelium. RnIP3K-C has a calculated molecular mass of ϳ74.5 kDa and shows an overall identity of ϳ75% with HsIP3K-C. A bacterially expressed, enzymatically active and Ca 2؉ -calmodulin-regulated fragment of this isoform displays remarkable enzymatic properties like a very low K m for Ins(1,4,5)P 3 (ϳ0.2 M), substrate inhibition by high concentrations of Ins(1,4,5)P 3 , allosteric product activation by Ins(1,3,4,5)P 4 in absence of Ca 2؉ -calmodulin (K a(app) 0.52 M), and the ability to efficiently phosphorylate a second InsP 3 substrate, inositol 2,4,5-trisphosphate, to inositol 2,4,5,6-tetrakisphosphate in the presence of Ins(1,3,4,5)P 4 . Furthermore, the RnIP3K-C fused with a fluorescent protein tag is actively transported into and out of the nucleus when transiently expressed in mammalian cells. A leucinerich nuclear export signal and an uncharacterized nuclear import activity are localized in the N-terminal domain of the protein and determine its nucleocytoplasmic shuttling. These findings point to a particular role of RnIP3K-C in nuclear inositol trisphosphate phosphorylation and cellular growth.For the conversion of the calcium-liberating second messenger inositol 1,4,5-trisphosphate (Ins(1,4,5)P 3 )

Regulation of innate immunity by inositol 1,3,4,5-tetrakisphosphate

“…A and B isoforms were found to be localized in both cytoskeleton and cytosolic, indicating a connection between cytoskeleton structure and InsP3K function. 5,[55][56][57] Consistent with this, similar with F-actin, InsP3KB is highly localized in the leading edge of polarized neutrophils. 16 In addition, since all three isoforms contain a calmodulin binding motif, the InsP3KB activity in neutrophils might also be regulated by calcium.…”

“…The accelerated proliferation of granulocyte monocyte progenitors may be a result of enhanced PtdIns(3,4,5)P3/Akt signaling, which has been recently implicated in myelopoiesis. 13 This signaling pathway was previously thought to be dependent solely upon concentrations of PtdIns (3,4,5) P3 in the plasma membrane. 14,15 Recently, we demonstrated that two inositol phosphates, InsP7 and Ins(1,3,4,5)P4, compete for Akt-PH domain binding with PtdIns(3,4,5)P3 both in vitro and in vivo, providing another level of regulation for Akt membrane translocation and activation.…”

“…Ins (1,3,4,5)P4, the product of InsP3KB, can exerts its function via binding to other cellular targets. Several proteins, including GAP1 IP4BP (also known as Rasa3), α-centaurin, and GAP1 m , also specifically interact with Ins(1,3,4,5)P4, 18,19 suggesting that the functions of these proteins might also be regulated by Ins (1,3,4,5) 21 Whether a similar mechanism also exists in hematopoietic progenitors was not investigated. InsP3K has also been reported to be a potential modulator of calcium mobilization, since it can decrease the level of Ins(1,4,5)P3, which mediates calcium release from internal store, by converting it to Ins(1,3,4,5)P4.…”

“…3 Ins(1,4,5)P3 can be converted to inositol 1,3,4,5-tetrakisphosphate (Ins(1,3,4,5)P4) by a family of inositol trisphosphate kinases (InsP3K). In mammalian cells, there are three isoforms of InsP3K, designated A, B and C. [4][5][6] They share a wellconserved carboxy-terminal catalytic domain, an inositol binding motif, and a calmodulin binding domain. The gene encoding InsP3KA is expressed exclusively in specific neuronal subpopulations in the central nervous system, in testis, and in some hematopoietic progenitor cell populations.…”

“…The genes encoding isoform B and C are expressed fairly ubiquitously. [5][6][7][8] The physiological functions of InsP3K in hematopoietic cells were recently studied using InsP3K knockout mice. It appeared that InsP3KB isoform contributes to the majority of InsP3 kinase activity in T and B cells.…”