Encyclopedia of Signaling Molecules

2018 Edition
| Editors: Sangdun Choi

Phosphatidylinositol 4-Kinase (PI4K2B)

Reference work entry
DOI: https://doi.org/10.1007/978-3-319-67199-4_101524

Synonyms

Historical Background

Phosphatidylinositol kinase activity was initially defined by Mitchell and colleagues as one that could transfer the γ-phosphoryl group of 32P-labeled ATP to phosphatidylinositol in membrane fractions derived from tissues (Michell et al. 1967). Two decades later, studies involving the use of cell and tissue extracts revealed that phosphatidylinositol kinase activity could be separated into three distinct types based on sensitivity to inhibitors and migration in a sucrose gradient (Whitman et al. 1987). These were described as the type I, II, and III phosphatidylinositol kinases. The type I enzyme was later found to phosphorylate the D3 position of the myo-inositol moiety of phosphatidylinositol and is now referred to as phosphatidylinositol 3-kinase (PI3K). The type II and III enzymes phosphorylate the D4 position but exhibit different biochemical characteristics such as sensitivities to inhibitors and migration in a sucrose gradient. The type III enzymes are sensitive to the PI3K inhibitor, wortmannin, albeit at higher concentrations. The type II enzymes on the other hand are insensitive to wortmannin but are inhibited by micromolar (μM) concentrations of adenosine and have a low KM for ATP. For several years, these were the only distinguishing features for studying the different isoforms (Balla and Balla 2006). This resulted in some difficulties in probing the functions of individual activities, as sometimes results were obscured by contamination with trace amounts of the highly active type II enzyme factions. Advancements in genomics and molecular cloning in the past two decades made unambiguous characterization of the different isoforms a possibility. Recently, crystal structures of catalytic sites of the type II enzymes were solved, revealing certain structural features that could be used in the development of potent pharmacological agents (Baumlova et al. 2014; Klima et al. 2015; Zhou et al. 2014). Molecular cloning has also led to a greatly improved understanding of the cellular roles of each isoform and has drawn greater attention to their involvement in the regulation of various cellular processes. The focus of this article is to give a description of the PI4KIIβ isoform, whose cellular roles are gradually being identified.

Structural and Molecular Features of PI4KIIβ

PI4KIIβ is a 55 kDa protein that possesses an overall fold similar to that of PI4KIIα due to sequence similarity. However, the two isoforms exhibit differences in their N-termini that extend over approximately 1-90 amino acid residues (Fig. 1). Like PI4KIIα, PI4KIIβ is insensitive to wortmannin, inhibited by micromolar (μM) concentrations of adenosine, and has a low KM for ATP. Crystal structure of the kinase domain revealed the presence of conserved molecular architecture similar to that of PI4KIIα and significantly different from the type III PI4Ks. Like PI4KIIα, the kinase domain of PI4KIIβ comprises an N-lobe and C-lobe that flank an ATP-binding grove. The N-lobe is made up of two α-helices that surround four antiparallel β-sheets while the C-lobe comprises two α-helices that surround two antiparallel β-sheets. The C-lobe also comprises two other distal α-helices that form a scaffold for the catalytically active part of the kinase and may provide allosteric regulation (Fig. 2). Within the N-Lobe is a cysteine-rich motif (CCPCC) that is palmitoylated and required for membrane tethering: a structural feature that modulates enzyme activity. The C-lobe possesses a hydrophobic binding pocket predicted to facilitate membrane association and binding of the lipid substrate (Klima et al. 2015). The kinase domain also comprises a lateral hydrophobic pocket that differs from that of PI4KIIα in spatial orientation of tryptophan residues Trp366 and Trp357, which are proposed to facilitate membrane anchorage (Baumlova et al. 2014). The authors also studied the effects of inhibitors by obtaining crystals bound to the nucleoside analogue MD59. However, they only succeeded in obtaining these with PI4KIIα. This crystal revealed the presence of three pockets proximal to the inhibitor and could be exploited for drug design. Resolution of the crystal structures of these proteins has opened new opportunities for enzymology and drug development. However, subtle differences between the catalytic domains of PI4KIIα and PI4KIIβ as well as inability to crystallize the divergent N-termini of the two isoforms imply that the design of an inhibitor with the ability to discriminate between the two isoforms is still very challenging.
Phosphatidylinositol 4-Kinase (PI4K2B), Fig. 1

Main structural features of the type II PI4Ks. The two isoforms share similarity in their kinase domains. The cysteine-rich CCPCC motif is palmitoylated, facilitating stronger membrane association. Both isoforms also have identical molecular weights (~55 kDa) but have divergent amino termini. The N-terminal of PI4KIIα includes a proline-rich domain while that of PI4KIIβ is enriched with clusters of acidic amino acid R groups

Phosphatidylinositol 4-Kinase (PI4K2B), Fig. 2

Kinase domain of PI4KIIβ showing the peptide backbone. The N-lobe is depicted in yellow and the C-lobe shown in green flank an ATP-binding groove. This domain comprises seven α-helices which are numbered H1-H7 and six β-sheets numbered S1-S6 (reproduced with permission of the International Union of Crystallography)

Subcellular Localization and Cellular Roles of PI4KIIβ

PI4KIIβ associates with membranes via dynamic palmitoylation of multiple cysteine residues within it catalytic domain (Barylko et al. 2001; Barylko et al. 2009). Molecular studies reveal that palmitoylation is required for catalytic activity of the type II PI4Ks. At a steady state, approximately 30% of total cellular PI4KIIβ is palmitoylated and distributed between membrane and cytoplasmic pools. The cytoplasmic pool is inactive and stabilized via its interaction with heat shock protein 90 (Hsp90). Short exposure to the Hsp90 inhibitor, geldanamycin, relieves PI4KIIβ of this interaction and facilitates palmitoylation, membrane docking, and subsequent synthesis of PI4P (Jung et al. 2011). PI4KIIβ catalyses phosphorylation of the D-4 hydroxyl group on the myo-inositol moiety of phosphatidylinositol (PI) to form PI4-phosphate (PI4P) which serves as a precursor for the synthesis of phosphatidylinositol(4,5)-bisphosphate (PI(4,5)P2) in agonist-stimulated signaling pathways. Growth factor activation induces membrane recruitment of PI4KIIβ, particularly to membrane ruffles in a Rac GTPase dependent manner (Wei et al. 2002). Beyond the generation of PI4P for PI(4,5)P2 synthesis, only a few other cellular roles have been ascribed to PI4KIIβ.

PI4KIIβ localizes to the trans-Golgi network (TGN) where it interacts with clathrin adaptor, AP-1 (Wieffer et al. 2013). Loss of this isoform via RNA interference results in TGN fragmentation and mislocalization of Mannose 6-phosphate receptor (MPR), which is required for delivery of lysosomal hydrolases. Thus, PI4KIIβ may play a role in vesicular trafficking between the TGN and late endosomes/lysosomes. PI4KIIβ is also associated with the Dishevelled (Dvl) component of the Wnt receptor and regulates endosomal recycling of Frizzled (Fz) component in canonical Wnt signaling (Wieffer et al. 2013), which in turn plays a central role in development and metastasis (Moon 2005). PI4KIIβ also interacts with the tetraspanin, CD81, and regulates the formation of nonendosomal vesicles that sequester the actin-binding protein, actinin, thereby altering cytoskeletal assembly, consequently preventing the migration of tumor cell lines in vitro (Mazzocca et al. 2008).

Recruitment of PI4KIIβ to the plasma membrane following growth factor stimulation (Wei et al. 2002) is suggestive of a role in growth factor mediated signaling where PI4KIIβ supplies PI4P for second messenger synthesis. PI4KIIβ has also been implicated in the regulation of tyrosine phosphorylation in response to T-cell activation (Sinha et al. 2013) while its localization to nucleoplasmic storage vesicles may play a role in nuclear calcium mediated signaling (Yoo et al. 2014).

Role in Health and Disease

PI4KIIβ plays a role in recycling of the Wnt receptor Frizzled (Wieffer et al. 2013) and upregulation of Wnt signaling in different tumors suggest that PI4KIIβ may play a role in certain human cancers. PI4KIIβ may serve as a metastasis suppressor in hepatic cancers. As reported by Carloni and colleagues, depletion of this enzyme results in the generation of a cellular phenotype characterized by dysregulated actin cytoskeleton and migration (Mazzocca et al. 2008). However, the mechanism by which this isoform regulates actin polymerization and cell migration is poorly understood. PI4KIIβ may also serve as a positive marker for response to adjuvant therapy in colorectal cancers as its overexpression along with that of STIM2 is associated with better prognosis, while siRNA-mediated loss of expression results in increased proliferation of primary tumors in vitro (Aytes et al. 2008) (meeting abstract). In addition, increased loss of heterozygosity on chromosome 4 in regions spanning 4p15.2-4p14 locus in colorectal cancer suggests that PI4KIIβ (localized to 4p15.2) may be a tumor suppressor (Zheng et al. 2008).

PI4KIIβ may be required in the innate immune response as it has been implicated in the early phases or phagosome maturation in macrophages (Jeschke et al. 2015). It has also been implicated in other immune responses involving T-lymphocytes and may contribute to the antitumor response of CD4+ T lymphocytes (Griffioen et al. 2008; Sinha et al. 2013).

Summary

PI4KIIβ catalyses the synthesis of PI4P. Like its closely related isoform, PI4KIIα, it contributes to the generation of membrane pools of PI4P, particularly on TGN and endosomal membranes. The two isoforms share close amino acid homology in their kinase domains but have highly dissimilar amino termini. This divergence may determine membrane localization and protein interaction. It is also important to note that there are subtle differences within the highly conserved catalytic domains which may be exploited in drug or inhibitor design.

Beyond its synthesis of PI4P, PI4KIIβ also engages with certain proteins via secondary interactions and such interactions have been implicated in different cellular processes. It regulates membrane trafficking pathways between the TGN and late endosomes and may also play a role in the regulation of growth factor signaling as its membrane localization is influenced by growth factor activation. It has also been described in different aspects of T-lymphocyte signaling while its involvement in the early stages of phagosome maturation also suggests that it may play a part in innate immunity.

This enzyme has also been described in developmental processes in zebrafish and may also play a role in development in higher vertebrates (including humans). PI4KIIβ expression has also been implicated in certain cancer types particularly those of the gut and the liver and may serve as a tumor suppressor. Overall, other noncatalytic roles of PI4KIIβ are gradually being unraveled, and improved structural information can aid study designs, thereby aiding in understanding nonredundant cellular roles of this isoform.

References

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© Springer International Publishing AG 2018

Authors and Affiliations

  1. 1.Lipid and Membrane Biology Group, UCL Institute for Liver and Digestive Health, Division of MedicineUniversity College LondonLondonUK