Suyong Choi

IQGAP1 is a novel phosphatidylinositol 4,5 bisphosphate effector in regulation of directional cell migration

Phosphatidylinositol 4,5 bisphosphate (PIP2) is a key lipid messenger for regulation of cell migration. PIP2 modulates many effectors, but the specificity of PIP2 signalling can be defined by interactions of PIP2‐generating enzymes with PIP2 effectors. Here, we show that type Iγ phosphatidylinositol 4‐phosphate 5‐kinase (PIPKIγ) interacts with the cytoskeleton regulator, IQGAP1, and modulates IQGAP1 function in migration. We reveal that PIPKIγ is required for IQGAP1 recruitment to the leading edge membrane in response to integrin or growth factor receptor activation. Moreover, IQGAP1 is a PIP2 effector that directly binds PIP2 through a polybasic motif and PIP2 binding activates IQGAP1, facilitating actin polymerization. IQGAP1 mutants that lack PIPKIγ or PIP2 binding lose the ability to control directional cell migration. Collectively, these data reveal a synergy between PIPKIγ and IQGAP1 in the control of cell migration.

PIP kinases define PI4,5P2 signaling specificity by association with effectors ☆

Phosphatidylinositol 4,5-bisphosphate (PI4,5P₂) is an essential lipid messenger with roles in all eukaryotes and most aspects of human physiology. By controlling the targeting and activity of its effectors, PI4,5P₂modulates processes, such as cell migration, vesicular trafficking, cellular morphogenesis, signaling and gene expression. In cells, PI4,5P₂has a much higher concentration than other phosphoinositide species and its total content is largely unchanged in response to extracellular stimuli. The discovery of a vast array of PI4,5P₂ binding proteins is consistent with data showing that the majority of cellular PI4,5P₂is sequestered. This supports a mechanism where PI4,5P₂functions as a localized and highly specific messenger. Further support of this mechanism comes from the de novo synthesis of PI4,5P₂which is often linked with PIP kinase interaction with PI4,5P₂effectors and is a mechanism to define specificity of PI4,5P₂signaling. The association of PI4,5P₂-generating enzymes with PI4,5P₂effectors regulate effector function both temporally and spatially in cells. In this review, the PI4,5P₂effectors whose functions are tightly regulated by associations with PI4,5P₂-generating enzymes will be discussed. This article is part of a Special Issue entitled Phosphoinositides.

Emerging roles of PtdIns(4,5)P2 – beyond the plasma membrane

ABSTRACT Phosphoinositides are a collection of lipid messengers that regulate most subcellular processes. Amongst the seven phosphoinositide species, the roles for phosphatidylinositol 4,5-bisphosphate [PtdIns(4,5)P2] at the plasma membrane, such as in endocytosis, exocytosis, actin polymerization and focal adhesion assembly, have been extensively studied. Recent studies have argued for the existence of PtdIns(4,5)P2 at multiple intracellular compartments, including the nucleus, endosomes, lysosomes, autolysosomes, autophagic precursor membranes, ER, mitochondria and the Golgi complex. Although the generation, regulation and functions of PtdIns(4,5)P2 are less well-defined in most other intracellular compartments, accumulating evidence demonstrates crucial roles for PtdIns(4,5)P2 in endolysosomal trafficking, endosomal recycling, as well as autophagosomal pathways, which are the focus of this Commentary. We summarize and discuss how phosphatidylinositol phosphate kinases, PtdIns(4,5)P2 and PtdIns(4,5)P2-effectors regulate these intracellular protein and membrane trafficking events. Summary: In this Commentary, we review current evidence supporting intracellular PtdIns(4,5)P2 signaling, with a focus on PtdIns(4,5)P2 regulation of endolysosomal trafficking and protein recycling, as well as autophagic membrane trafficking.

Isoform 5 of PIPKIγ regulates the endosomal trafficking and degradation of E-cadherin

ABSTRACT Phosphatidylinositol phosphate kinases (PIPKs) have distinct cellular targeting, allowing for site-specific synthesis of phosphatidylinositol 4,5-bisphosphate [PI(4,5)P2] to activate specific signaling cascades required for cellular processes. Several C-terminal splice variants of PIPKI&ggr; (also known as PIP5K1C) exist, and have been implicated in a multitude of cellular roles. PI(4,5)P2 serves as a fundamental regulator of E-cadherin transport, and PI(4,5)P2-generating enzymes are important signaling relays in these pathways. We present evidence that the isoform 5 splice variant of PIPKI&ggr; (PIPKI&ggr;i5) associates with E-cadherin and promotes its lysosomal degradation. Additionally, we show that the endosomal trafficking proteins SNX5 and SNX6 associate with PIPKI&ggr;i5 and inhibit PIPKI&ggr;i5-mediated E-cadherin degradation. Following HGF stimulation, activated Src directly phosphorylates PIPKI&ggr;i5. Phosphorylation of the PIPKI&ggr;i5 C-terminus regulates its association with SNX5 and, consequently, E-cadherin degradation. Additionally, this PIPKI&ggr;i5-mediated pathway requires Rab7 to promote degradation of internalized E-cadherin. Taken together, the data indicate that PIPKI&ggr;i5 and SNX5 are crucial regulators of E-cadherin sorting and degradation. PIPKI&ggr;i5, SNX and phosphoinositide regulation of lysosomal sorting represent a novel area of PI(4,5)P2 signaling and research. PIPKI&ggr;i5 regulation of E-cadherin sorting for degradation might have broad implications in development and tissue maintenance, and enhanced PIPKI&ggr;i5 function might have pathogenic consequences due to downregulation of E-cadherin.

Phosphatidylinositol Phosphate 5-Kinase Iγi2 in Association with Src Controls Anchorage-independent Growth of Tumor Cells

Background: PIPKIγ isoforms play roles in cell migration, polarization, and membrane trafficking and are overexpressed in triple-negative breast cancers, indicating protumorigenic functions. Results: PIPKIγi2 associates with the C terminus of Src, and this interaction interdependently controls their functioning. Conclusion: PIPKIγi2 and Src synergistically control anchorage-independent tumor cell growth. Significance: This study shows unexpected mechanisms for a phosphatidylinositol 4,5-biphosphate-generating enzyme that synergizes with the proto-oncogene Src to regulate oncogenic signaling. A fundamental property of tumor cells is to defy anoikis, cell death caused by a lack of cell-matrix interaction, and grow in an anchorage-independent manner. How tumor cells organize signaling molecules at the plasma membrane to sustain oncogenic signals in the absence of cell-matrix interactions remains poorly understood. Here, we describe a role for phosphatidylinositol 4-phosphate 5-kinase (PIPK) Iγi2 in controlling anchorage-independent growth of tumor cells in coordination with the proto-oncogene Src. PIPKIγi2 regulated Src activation downstream of growth factor receptors and integrins. PIPKIγi2 directly interacted with the C-terminal tail of Src and regulated its subcellular localization in concert with talin, a cytoskeletal protein targeted to focal adhesions. Co-expression of PIPKIγi2 and Src synergistically induced the anchorage-independent growth of nonmalignant cells. This study uncovers a novel mechanism where a phosphoinositide-synthesizing enzyme, PIPKIγi2, functions with the proto-oncogene Src, to regulate oncogenic signaling.

PtdIns(4,5)P2 signaling regulates ATG14 and autophagy

Significance Autophagy is a conserved lysosomal degradation pathway, the deregulation of which is found in many human diseases, including cancers, neurodegeneration diseases, and aging. Understanding the molecular mechanisms of autophagy regulation is the basis for improving clinical therapeutics. Phosphatidylinositol 3-phosphate [PtdIns(3)P] is a key lipid messenger directly involved in autophagy initiation. Here we discovered that PtdIns(4,5)P2 plays an equally important role in autophagy. The autophagy-related protein 14 (ATG14) specifically interacts with PIPKIγi5, a PtdIns(4,5)P2 generating enzyme that controls ATG14 stability and protein–protein interactions. PtdIns(4,5)P2 directly binds ATG14 and regulates the ATG14–VPS34 (a class III phosphatidylinositol 3-kinase) complex assembly. Our study identifies a PtdIns(4,5)P2 signaling pathway that is directly involved in autophagosome membrane initiation and argues for a previously unappreciated role for endosomes in autophagy initiation. Autophagy is a regulated self-digestion pathway with fundamental roles in cell homeostasis and diseases. Autophagy is regulated by coordinated actions of a series of autophagy-related (ATG) proteins. The Barkor/ATG14(L)–VPS34 (a class III phosphatidylinositol 3-kinase) complex and its product phosphatidylinositol 3-phosphate [PtdIns(3)P] play key roles in autophagy initiation. ATG14 contains a C-terminal Barkor/ATG14(L) autophagosome-targeting sequence (BATS) domain that senses the curvature of PtdIns(3)P-containing membrane. The BATS domain also strongly binds PtdIns(4,5)P2, but the functional significance has been unclear. Here we show that ATG14 specifically interacts with type Iγ PIP kinase isoform 5 (PIPKIγi5), an enzyme that generates PtdIns(4,5)P2 in mammalian cells. Autophagosomes have associated PIPKIγi5 and PtdIns(4,5)P2 that are colocalized with late endosomes and the endoplasmic reticulum. PtdIns(4,5)P2 generation at these sites requires PIPKIγi5. Loss of PIPKIγi5 results in a loss of ATG14, UV irradiation resistance-associated gene, and Beclin 1 and a block of autophagy. PtdIns(4,5)P2 binding to the ATG14–BATS domain regulates ATG14 interaction with VPS34 and Beclin 1, and thus plays a key role in ATG14 complex assembly and autophagy initiation. This study identifies an unexpected role for PtdIns(4,5)P2 signaling in the regulation of ATG14 complex and autophagy.

Phosphatidylinositol Phosphate 5-Kinase Iγ and Phosphoinositide 3-Kinase/Akt Signaling Couple to Promote Oncogenic Growth.

Background: PIP2 generated by PIPKI family members regulates many cellular functions, and PIP2 is a PI3K substrate. Results: Loss of PIPKIγ or PIPKIγi2 impaired Akt activation. PIPKIγi2 and Src activate Akt and anchorage-independent growth. Conclusion: PI3K/Akt signaling is regulated by coupling with PIPKIγ. Significance: The coupling of PIPKIγ and PI3K indicates a mechanism for Akt activation that enhances oncogenic growth. The assembly of signaling complexes at the plasma membrane is required for the initiation and propagation of cellular signaling upon cell activation. The class I PI3K and the serine/threonine-specific protein kinase Akt signaling pathways (PI3K/Akt) are often activated in tumors. These pathways are initiated by the generation of phosphatidylinositol 3,4,5-triphosphate (PIP3) by PI3K-mediated phosphorylation of phosphatidylinositol 4,5-biphosphate (PIP2), synthesized by phosphatidylinositol 4-phosphate 5-kinase (PIPKI) enzymes. The mechanism of how tumor cells recruit and organize the PIP2-synthesizing enzymes with PI3K in the plasma membrane for activation of PI3K/Akt signaling is not defined. Here, we demonstrated a role for the phosphatidylinositol 4-phosphate 5-kinase Iγ (PIPKIγ) in PI3K/Akt signaling. PIPKIγ is overexpressed in triple-negative breast cancers. Loss of PIPKIγ or its focal adhesion-targeting variant, PIPKIγi2, impaired PI3K/Akt activation upon stimulation with growth factors or extracellular matrix proteins in different tumor cells. PIPKIγi2 assembles into a complex containing Src and PI3K; Src was required for the recruitment of PI3K enzyme into the complex. PIPKIγi2 interaction with Src and its lipid kinase activity were required for promoting PI3K/Akt signaling. These results define a mechanism by which PIPKIγi2 and PI3K are integrated into a complex regulated by Src, resulting in the spatial generation of PIP2, which is the substrate PI3K required for PIP3 generation and subsequent Akt activation. This study elucidates the mechanism by which PIP2-generating enzyme controls Akt activation upstream of a PI3K enzyme. This pathway may represent a signaling nexus required for the survival and growth of metastasizing and circulating tumor cells in vivo.

IQGAP1 is a phosphoinositide effector and kinase scaffold.

Phosphatidylinositol 4,5-bisphosphate (PI4,5P2) is a lipid messenger that regulates a wide variety of cellular functions. The majority of cellular PI4,5P2 is generated by isoforms of the type I phosphatidylinositol phosphate kinases (PIPKI) that are generated from three genes, and each PIPKI isoform has a unique distribution and function in cells. It has been shown that the signaling specificity of PI4,5P2 can be determined by a physical association of PIPKs with PI4,5P2 effectors. IQGAP1 is newly identified as an interactor of multiple isoforms of PIPKs. Considering the versatile roles of IQGAP1 in cellular signaling pathways, IQGAP1 may confer isoform-specific roles of PIPKs in distinct cellular locations. In this mini review, the emerging roles of PIPKs that are regulated by an association with IQGAP1 will be summarized. Focuses will be on cell migration, vesicle trafficking, cell signaling, and nuclear events.

And Akt‐ion! IQGAP1 in control of signaling pathways

ERK signaling and Akt signaling are inversely correlated in some cancers. Yet, the precise molecular mechanism for cross‐inhibition remains unclear. In this issue of The EMBO Journal, Pan et al ( ) show that when Akt is on, its phosphorylated cytoplasmic substrate FOXO1 turns off ERK activity by reshaping the Ras‐ERK scaffold IQGAP1.

PIPKIγ and talin couple phosphoinositide and adhesion signaling to control the epithelial to mesenchymal transition.

Epithelial cells acquire migratory/invasive and stemness traits upon conversion to the mesenchymal phenotype. The expression of E-cadherin is a key to this transition; yet precise understanding of the pathways involved in integrating E-cadherin loss to the gain of mesenchymal traits remains poorly understood. Here, we show that phosphoinositide-generating enzyme, PIPKIγ, expression is upregulated upon epithelial–mesenchymal transition (EMT) and together with the cytoskeletal protein talin assemble into a signaling complex upon E-cadherin loss. PIPKIγ and talin together control the adhesion and phosphoinositide signaling that regulates conversion to the mesenchymal phenotypes. PIPKIγ and talin regulate the stability of E-cadherin transcriptional repressors, snail and slug, induced by transforming growth factor-β1 or extracellular matrix protein. Loss of PIPKIγ or talin or their interaction impaired EMT and the acquisition of cell motility and stemness. This demonstrates a mechanism where a phosphoinositide-generating enzyme PIPKIγ couples with a cytoskeletal protein talin to control the acquisition of mesenchymal phenotypes.