Plasma Membrane In Response Research Articles

PI 3-kinase and disease

Phosphoinositide 3-Kinase (PI3K) is a central enzyme in a signaling pathway that mediates cellular responses to growth factors. This enzyme phosphorylates the 3 position of phosphatidylinositol-4,5-bisphosphate to produce phosphatidylinositol-3,4,5-trisphosphate (PIP3) at the plasma membrane. A number of signaling proteins, including the Ser/Thr protein kinases, AKT and PDK1, contain pleckstrin homology domains that bind specifically to PIP3. Thus, the generation of PIP3 at the plasma membrane in response to activation of PI3K by growth factors results in the initiation of downstream Ser/Thr phosphorylation cascades that control a variety of cellular responses. The signaling pathway downstream of PI3K is highly conserved from worms and flies to humans and genetic analysis of the pathway has revealed a conserved role in regulating glucose metabolism and cell growth. Based on deletion of genes encoding the catalytic or regulatory subunits of PI3K in the mouse, PI3K mediates insulin dependent regulation of glucose metabolism, and defects in activation of this pathway result in insulin resistance. In contrast, mutational events that lead to hyperactivation of the PI3K pathway result in cancers. Activating mutations in PIK3CA, encoding the p110alpha catalytic subunit of PI3K or inactivating mutations in PTEN, a phosphoinositide 3-phosphatases that reverses the effects of PI3K, are among the most common events in solid tumors. We have generated mouse models in which a mutated form of the PIK3CA gene is expressed in a tissue specific and reversibly inducible manner. These mice develop cancers that are dependent on continuous expression of the mutant PIK3CA gene. The PIK3CA driven tumors are FDG-PET positive and turning off PI 3-Kinase with PI3K inhibitors that are in human clinical trials results in an acute decline in FDG-PET signal that precedes tumor shrinkage. These results suggest that the ability of PI3K to stimulate high rates of glucose uptake and metabolism may be critical for the survival of PIK3CA mutant tumors. The role of PI3K inhibitors for treating cancers in mouse models and in human trials will be discussed.

ZnT4 provides zinc to zinc-dependent proteins in the trans-Golgi network critical for cell function and Zn export in mammary epithelial cells

Zinc (Zn) transporter 4 (ZnT4) plays a key role in mammary gland Zn metabolism. A mutation in ZnT4 (SLC30A4) that targets the protein for degradation is responsible for the "lethal milk" (lm/lm) mouse phenotype. ZnT4 protein is only detected in the secreting mammary gland, and lm/lm mice have ∼35% less Zn in milk, decreased mammary gland size, and decreased milk secretion. However, the precise contribution of ZnT4 is unknown. We used cultured mouse mammary epithelial cells (HC11) and determined that ZnT4 was localized to the trans-Golgi network (TGN) and cell membrane and transported Zn from the cytoplasm. ZnT4-mediated Zn import into the TGN directly contributed to labile Zn accumulation as ZnT4 overexpression increased FluoZin3 fluorescence. Moreover, ZnT4 provided Zn for metallation of galactosyltransferase, a Zn-dependent protein localized within the TGN that is critical for milk secretion, and carbonic anhydrase VI, a Zn-dependent protein secreted from the TGN into milk. We further noted that ZnT4 relocalized to the cell membrane in response to Zn. Together these studies demonstrated that ZnT4 transports Zn into the TGN, which is critical for key secretory functions of the mammary cell.

Characterization of Differential Protein Tethering at the Plasma Membrane in Response to Epidermal Growth Factor Signaling

Physical tethering of membrane proteins to the cortical actin cytoskeleton provides functional organization to the plasma membrane and contributes to diverse cellular processes including cell signaling, vesicular trafficking, endocytosis, and migration. For these processes to occur, membrane protein tethering must be dynamically regulated in response to environmental cues. In this study, we describe a novel biochemical scheme for isolating the complement of plasma membrane proteins that are physically tethered to the actin cytoskeleton. We utilized this method in combination with tandem liquid chromatography/mass spectrometry (LC-MS/MS) to demonstrate that cytoskeletal tethering of membrane proteins is acutely regulated by epidermal growth factor (EGF) in normal human kidney (HK2) cells. Our results indicate that several proteins known to be involved in EGF signaling, as well as other proteins not traditionally associated with this pathway, are tethered to the cytoskeleton in dynamic fashion. Further analysis of one hit from our proteomic survey, the receptor phosphotyrosine phosphatase PTPRS, revealed a correlation between cytoskeletal tethering and endosomal trafficking in response to EGF. This finding parallels previous indications that PTPRS is involved in the desensitization of EGFR and provides a potential mechanism to coordinate localization of these two membrane proteins in the same compartment upon EGFR activation.

Abstract 361: Cyclophilin A Mediates Angiotensin II--Stimulated NADPH Oxidase Activation in Vascular Smooth Muscle Cells

Objective: Cyclophilin A (CyPA) is a ubiquitously expressed cytosolic protein that possesses PPIase activity and scaffold function. CyPA regulates Angiotensin II (Ang II) induced reactive oxygen species (ROS) production in vascular smooth muscle cells. However, the mechanism of this CyPA regulation remains unclear. We hypothesized that CyPA regulates plasma membrane translocation of NADPH oxidase cytosolic subunit, p47phox, which is required for NADPH oxidase structural organization and activity. Methods and results: Immunofluorescence studies in rat aortic smooth muscle cells revealed that CyPA translocated from the cytosol to the plasma membrane in response to Ang II in a time dependent manner with a peak at 10min (46.4±5.4 fold increase). Mouse Aortic Smooth Muscle Cells (MASM) were isolated from mice lacking CyPA (CyPA-/-) and wild type controls (WT), treated with Ang II (100nM) and immunofluorescence analysis was performed. Ang II induced p47phox plasma membrane translocation at 10min in WT mice. However, p47 phox translocation was significantly inhibited in CyPA -/- MASM. CyPA and p47phox colocalized at the plasma membrane in response to Ang II. Further analysis using subcellular fractionation studies confirmed that Ang II induced p47phox plasma membrane translocation was inhibited in CyPA -/- MASM compared to WT (1.2±2.7 vs 4.3±3.4 fold increase). Coimmunoprecipitation analyses confirmed that Ang II increased CyPA association with p47phox in a time dependent manner (2.5±3.4 fold increase at 10min). Finally, pretreatment with the PPIase activity inhibitor, cyclosporine A (1uM), could not inhibit CyPA association with p47phox and CyPA mediated p47phox translocation to the plasma membrane. Conclusion: These data suggest that Ang II promotes an association between CyPA and p47phox that enhances plasma membrane translocation of p47phox. This is proposed to increase the NADPH oxidase activity thereby increasing cellular ROS production. This process is independent of the PPIase activity of CyPA. Therefore, inhibition of the CyPA and p47phox association could be a future therapeutic target for Ang II induced ROS regulated cardiovascular diseases such as atherosclerosis and abdominal aortic aneurysm formation.

IP3 receptor processing by the ERAD pathway

Inositol 1,4,5‐trisphosphate (IP3) receptors are large ER membrane proteins which form tetrameric channels that govern the release of Ca2+ stored within the ER lumen of vertebrate cells. They are named for their ability to bind to and be opened by the second messenger IP3, which is generated at the plasma membrane in response to cell surface receptor activation. Thus, IP3 receptors are pivotal in signaling pathways that couple extracellular hormones, neurotransmitters and growth factors to increases in cytoplasmic Ca2+ concentration and the regulation of Ca2+‐dependent events (e.g. secretion, fertilization, apoptosis and gene expression). Surprisingly, when IP3 receptors are activated they are rapidly destroyed via the ER‐associated degradation (ERAD) pathway, a ubiquitination‐and proteasome‐dependent mechanism that clears the ER of aberrant proteins. I will review recent studies aimed at identifying which ERAD pathway components mediate IP3 receptor ERAD, focusing on a novel complex composed of the ER membrane proteins erlin 1 and 2 which binds to IP3 receptors immediately after they are activated, and the E3 ligase RNF170, which participates in IP3 receptor ubiquitination.

AS160: a new Na,K‐ATPase partner that regulates the trafficking of the sodium pump in response to energy depletion and renal ischemia

The Na,K‐ATPase appears to be located in two different cellular pools. The major sodium pump pool is located at the plasma membrane, while the other pool resides in cytoplasmic vesicular compartments which can translocate to the plasma membrane in response to physiological stimuli. We have identified AS160 as a new Na,K‐ATPase partner. We also demonstrated that the knockdown of AS160 increases the level of cellular Na,K‐ATPase activity, suggesting that AS160 modulates the activity of the enzyme. In the present study we characterized aspects of the physiological role of the interaction between AS160 and Na,K‐ATPase. We find that AS160 is required for the intracellular accumulation of the Na, K‐ATPase that occurs in response to energy depletion in cultured epithelial cells. In intact animals ischemic kidney injury has been shown to induce internalization of the Na,K‐ATPase from the plasma membranes of renal epithelial cells. We assessed the distribution of the Na,K‐ATPase following renal ischemia and reperfusion in kidneys from wild type and AS160 knockout mice. We observed a substantial decrease in the injury‐induced cytoplasmic accumulation of the Na,K‐ATPase in the renal epithelial cells of the AS160 knockout mice as compared to those of the wild type controls. We propose that AS160 plays a key role in regulating the redistribution of the Na,K‐ATPase that occurs in response to energy depletion and renal ischemia.

Sodium Channel Blockers as Therapeutic Target for Treating Epilepsy: Recent Updates

The voltage-gated sodium channels (VGSCs) are a family of membrane proteins forming a pore, through which they selectively conduct sodium ions inward and outward cell's plasma membranes in response to variations of membrane potentials, playing a fundamental role in controlling cellular excitability. Growing evidences suggest that abnormal VGSCs are involved in the pathophysiology of both acquired and inherited epilepsy. Approximately two dozen drugs are currently marketed for the treatment of epilepsy and most of them act as sodium channel blockers, preventing the return of the channels to the active state by stabilizing the inactive form. Despite the many drugs on the market, 30% of patients continue to experience seizures even in the presence of optimal doses of AEDs, while others continue to suffer from medication induced side effects. Thus, there is a great need to continue the search for new AEDs that are not only more effective, but also have a better side effects profile. For this reason, many efforts have been made in the recent years to identify new sodium channel blockers for the treatment of epilepsy. These studies have led to different classes of compounds, characterized by a great structural diversity. The aim of this review is to provide an introduction on the structure and function of the sodium channels, followed by a brief historical perspective on the sodium channel blockers in use as anticonvulsant drugs. Moreover, it will focus on the medicinal chemistry of the sodium channel blockers recently published (2008-2011) and the drug design/molecular modeling studies related to the receptor.

Imaging P2X4 Receptor Lateral Mobility in Microglia

ATP-gated ionotropic P2X4 receptors are up-regulated in activated microglia and are critical for the development of neuropathic pain, a microglia-associated disorder. However, the nature of how plasma membrane P2X4 receptors are regulated in microglia is not fully understood. We used single-molecule imaging to track quantum dot-labeled P2X4 receptors to explore P2X4 receptor mobility in the processes of resting and activated microglia. We find that plasma membrane P2X4 receptor lateral mobility in resting microglial processes is largely random, consisting of mobile and slowly mobile receptors. Moreover, lateral mobility is P2X subunit- and cell-specific, increased in an ATP activation and calcium-dependent manner, and enhanced in activated microglia by the p38 MAPK pathway that selectively regulates slowly mobile receptors. Thus, our data indicate that P2X4 receptors are dynamically regulated mobile ATP sensors, sampling more of the plasma membrane in response to ATP and during the activated state of microglia that is associated with nervous system dysfunction.

Protein Implicated in Nonsyndromic Mental Retardation Regulates Protein Kinase A (PKA) Activity

Mutation of the coiled-coil and C2 domain-containing 1A (CC2D1A) gene, which encodes a C2 domain and DM14 domain-containing protein, has been linked to severe autosomal recessive nonsyndromic mental retardation. Using a mouse model that produces a truncated form of CC2D1A that lacks the C2 domain and three of the four DM14 domains, we show that CC2D1A is important for neuronal differentiation and brain development. CC2D1A mutant neurons are hypersensitive to stress and have a reduced capacity to form dendrites and synapses in culture. At the biochemical level, CC2D1A transduces signals to the cyclic adenosine 3',5'-monophosphate (cAMP)-protein kinase A (PKA) pathway during neuronal cell differentiation. PKA activity is compromised, and the translocation of its catalytic subunit to the nucleus is also defective in CC2D1A mutant cells. Consistently, phosphorylation of the PKA target cAMP-responsive element-binding protein, at serine 133, is nearly abolished in CC2D1A mutant cells. The defects in cAMP/PKA signaling were observed in fibroblast, macrophage, and neuronal primary cells derived from the CC2D1A KO mice. CC2D1A associates with the cAMP-PKA complex following forskolin treatment and accumulates in vesicles or on the plasma membrane in wild-type cells, suggesting that CC2D1A may recruit the PKA complex to the membrane to facilitate signal transduction. Together, our data show that CC2D1A is an important regulator of the cAMP/PKA signaling pathway, which may be the underlying cause for impaired mental function in nonsyndromic mental retardation patients with CC2D1A mutation.

A novel murine model for non-alcoholic steatohepatitis developed by combination of a high-fat diet and oxidized low-density lipoprotein

Non-alcoholic steatohepatitis (NASH) is the hepatic manifestation of metabolic syndrome that is characterized by steatosis, inflammation, and fibrosis, and may progress to cirrhosis and carcinoma. To investigate its pathogenic processes, we established a novel murine model for NASH by combination of a high-fat diet (HFD) and oxidized low-density lipoprotein (oxLDL). Mice that received HFD for 23 weeks showed hepatic steatosis, slight fibrosis, and a high level of lipid peroxidation compared with a regular diet (RD)-fed mice. Hepatic injury and elevated tumor necrosis factor (TNF)-α mRNA expression were also detected in these mice. Moreover, oxLDL administration to HFD-fed mice during weeks 21–23 not only aggravated hepatic steatosis, fibrosis, and lipid metabolism, but also resulted in intense inflammation, including severe hepatic injury and inflammatory cell infiltration, which are the typical histological features of NASH. Inflammation was accompanied by increased gene expression of TNF-α and interleukin (IL)-6. Additionally, the livers of RD-fed animals treated with oxLDL during weeks 21–23 were characterized by foamy macrophages and inflammatory cell infiltration along with an elevated IL-6 mRNA level. These results suggest that an increased oxidative state, including HFD-induced intracellular lipid peroxidation and its extracellular source from oxLDL, is the actual trigger for hepatic inflammation in which liver injury is mediated by TNF-α and inflammatory cell accumulation is dependent on IL-6. HFD and oxLDL also induced insulin resistance in mice; additionally, oxLDL downregulated insulin secretion. In this model, CD36 overexpression was observed in the hepatocytes of HFD-fed mice and those treated with HFD and oxLDL, and in the hepatic macrophages of RD-fed mice immediately after oxLDL treatment. In vitro experiments indicated a rapid and transient elevation of CD36 on macrophage plasma membrane in response to oxLDL. Our findings demonstrate that CD36 expressed on hepatocytes and hepatic macrophages mediates the pathophysiology of NASH.

Plasticity of plasma membrane compartmentalization during plant immune responses

Plasma membranes require high levels of plasticity to modulate the perception and transduction of extracellular and intracellular signals. Dynamic lateral assembly of protein complexes combined with an independent compositional lipid patterning in both membrane leaflets provide cells the opportunity to decorate this interface with specific proteins in an organized but dynamic manner. Such ability to dynamically reorganize the protein content of the plasma membrane is essential for the regulation of processes such as polarity of transport, development, and microbial infection. While the plant cell wall represents the first physical and mostly unspecific barrier for invading microbes, the plasma membrane is at the forefront of microbial recognition and initiation of defense responses. Accumulating evidence indicating dynamic compartmentalization of plasma membranes in response to environmental cues has increased the interest in the compositional heterogeneity of this bilayer. Here, we elucidate the recruitment of specific proteins into defined membrane structures that ensure functional compartmentalization of the bilayer during infection processes.

Inversin, Wnt signaling and primary cilia

Mutations of the ankyrin-repeat protein Inversin, a member of a diverse family of more than 12 proteins, cause nephronophthisis (NPH), an autosomal recessive cystic kidney disease associated with extra-renal manifestations such as retinitis pigmentosa, cerebellar aplasia and situs inversus. Most NPH gene products (NPHPs) localize to the cilium, and appear to control the transport of cargo protein to the cilium by forming functional networks. Inversin interacts with NPHP1 and NPHP3, and shares with NPHP4 the ability to antagonize Dishevelled-stimulated canonical Wnt signaling, potentially through recruitment of the Anaphase Promoting Complex (APC/C). However, Dishevelled antagonism may be confined towards the basal body, thereby polarizing motile cilia on the cells of the ventral node and respiratory tract. Inversin is essential for recruiting Dishevelled to the plasma membrane in response to activated Frizzled, a crucial step in planar cell polarity signaling. During vertebrate pronephros development, the Inversin-mediated translocation of Dishevelled appears to orchestrate the migration of cells and differentiation of segments that correspond to the mammalian loop of Henle. Thus, defective tubule migration and elongation may contribute to concentration defects and cause cyst formation in patients with NPH.

Carboxyl-terminal modulator protein induces apoptosis by regulating mitochondrial function in lung cancer cells

Serine/threonine protein kinase B (PKB/Akt) is involved in cell survival and growth. Carboxyl-terminal modulator protein (CTMP), a novel Akt binding partner, prevents Akt activation at the plasma membrane in response to various stimuli, and thus possesses a tumor suppressor-like function. In a previous study, we have demonstrated that CTMP inhibits tumor progression by facilitating apoptosis in a mouse lung cancer model. However, the precise mechanism of CTMP-induced apoptosis remains to be elucidated. The present study was performed to examine the role of CTMP in mitochondrial-mediated apoptosis and regulation of mitochondrial function in human lung carcinoma cells. Our results showed that CTMP altered mitochondrial morphology and caused the release of cytochrome c by inhibiting OPA1 expression. Additionally, CTMP facilitated mitochondrial-mediated apoptosis by inhibiting heat-shock protein 27 and preventing cytochrome c interaction with Apaf-1. Our data suggest that CTMP may therefore play a critical role in mitochondrial-mediated apoptosis in lung cancer cells.

Active Trans-Plasma Membrane Water Cycling in Yeast Is Revealed by NMR

Plasma membrane water transport is a crucial cellular phenomenon. Net water movement in response to an osmotic gradient changes cell volume. Steady-state exchange of water molecules, with no net flux or volume change, occurs by passive diffusion through the phospholipid bilayer and passage through membrane proteins. The hypothesis is tested that plasma membrane water exchange also correlates with ATP-driven membrane transport activity in yeast (Saccharomyces cerevisiae). Longitudinal 1H2O NMR relaxation time constant (T1) values were measured in yeast suspensions containing extracellular relaxation reagent. Two-site-exchange analysis quantified the reversible exchange kinetics as the mean intracellular water lifetime (τi), where τi−1 is the pseudo-first-order rate constant for water efflux. To modulate cellular ATP, yeast suspensions were bubbled with 95%O2/5%CO2 (O2) or 95%N2/5%CO2 (N2). ATP was high during O2, and τi−1 was 3.1 s−1 at 25°C. After changing to N2, ATP decreased and τi−1 was 1.8 s−1. The principal active yeast ion transport protein is the plasma membrane H+-ATPase. Studies using the H+-ATPase inhibitor ebselen or a yeast genetic strain with reduced H+-ATPase found reduced τi−1, notwithstanding high ATP. Steady-state water exchange correlates with H+-ATPase activity. At volume steady state, water is cycling across the plasma membrane in response to metabolic transport activity.

ATP7A Gene Addition to the Choroid Plexus Results in Long-term Rescue of the Lethal Copper Transport Defect in a Menkes Disease Mouse Model

Menkes disease is a lethal infantile neurodegenerative disorder of copper metabolism caused by mutations in a P-type ATPase, ATP7A. Currently available treatment (daily subcutaneous copper injections) is not entirely effective in the majority of affected individuals. The mottled-brindled (mo-br) mouse recapitulates the Menkes phenotype, including abnormal copper transport to the brain owing to mutation in the murine homolog, Atp7a, and dies by 14 days of age. We documented that mo-br mice on C57BL/6 background were not rescued by peripheral copper administration, and used this model to evaluate brain-directed therapies. Neonatal mo-br mice received lateral ventricle injections of either adeno-associated virus serotype 5 (AAV5) harboring a reduced-size human ATP7A (rsATP7A) complementary DNA (cDNA), copper chloride, or both. AAV5-rsATP7A showed selective transduction of choroid plexus epithelia and AAV5-rsATP7A plus copper combination treatment rescued mo-br mice; 86% survived to weaning (21 days), median survival increased to 43 days, 37% lived beyond 100 days, and 22% survived to the study end point (300 days). This synergistic treatment effect correlated with increased brain copper levels, enhanced activity of dopamine-β-hydroxylase, a copper-dependent enzyme, and correction of brain pathology. Our findings provide the first definitive evidence that gene therapy may have clinical utility in the treatment of Menkes disease.

Neuronal P2X2 Receptors Are Mobile ATP Sensors That Explore the Plasma Membrane When Activated

ATP-gated ionotropic P2X2 receptors are widely expressed in neurons. Although the electrophysiological properties of P2X2 receptors have been extensively studied, little is known about the plasma membrane lateral mobility of P2X2 receptors or whether receptor mobility is regulated by ATP. Here we used single-molecule imaging with simultaneous whole-cell voltage-clamp recordings to track quantum dot-labeled P2X2 receptors in the dendrites of rat hippocampal neurons to explore P2X2 receptor mobility and its regulation. We find that plasma membrane P2X2 receptor lateral mobility in dendrites is heterogeneous but mostly Brownian in nature, consisting of mobile and slowly mobile receptor pools. Moreover, lateral mobility is P2X2 subunit and cell specific, is increased in an activation-dependent manner, and is regulated by cytosolic VILIP1, a calcium binding protein. Our data provide the first direct measures of P2X receptor mobility and show that P2X2 receptors are mobile ATP sensors, sampling more of the dendritic plasma membrane in response to ATP.

The S100A10-Annexin A2 Complex Provides a Novel Asymmetric Platform for Membrane Repair

Membrane repair is mediated by multiprotein complexes, such as that formed between the dimeric EF-hand protein S100A10, the calcium- and phospholipid-binding protein annexin A2, the enlargeosome protein AHNAK, and members of the transmembrane ferlin family. Although interactions between these proteins have been shown, little is known about their structural arrangement and mechanisms of formation. In this work, we used a non-covalent complex between S100A10 and the N terminus of annexin A2 (residues 1-15) and a designed hybrid protein (A10A2), where S100A10 is linked in tandem to the N-terminal region of annexin A2, to explore the binding region, stoichiometry, and affinity with a synthetic peptide from the C terminus of AHNAK. Using multiple biophysical methods, we identified a novel asymmetric arrangement between a single AHNAK peptide and the A10A2 dimer. The AHNAK peptide was shown to require the annexin A2 N terminus, indicating that the AHNAK binding site comprises regions on both S100A10 and annexin proteins. NMR spectroscopy was used to show that the AHNAK binding surface comprised residues from helix IV in S100A10 and the C-terminal portion from the annexin A2 peptide. This novel surface maps to the exposed side of helices IV and IV' of the S100 dimeric structure, a region not identified in any previous S100 target protein structures. The results provide the first structural details of the ternary S100A10 protein complex required for membrane repair.

Phosphatidylserine Binding Is Essential for Plasma Membrane Recruitment and Signaling Function of 3-Phosphoinositide-dependent Kinase-1

3-Phosphoinositide-dependent kinase-1 (PDK1) is a ubiquitously expressed serine/threonine kinase that functions downstream of phosphoinositide 3-kinase. Although binding of 3'-phosphoinositides, phosphatidylinositol 3,4,5-trisphosphate and phosphatidylinositol 3,4-bisphosphate, to the pleckstrin homology (PH) domain of PDK1 is known to be essential for its interaction with and activation of downstream kinases, the mechanism by which PDK1 is recruited to the plasma membrane remains controversial. Our surface plasmon resonance analysis of the PDK1 PH domain and selected mutants shows that the PH domain specifically binds phosphatidylserine using a site that is separate from the canonical phosphoinositide-binding site. Further cell studies show that this specific phosphatidylserine binding is important for the plasma membrane localization and signaling function of PDK1.

The role of proton transporters in epithelial Wnt signaling pathways

The planar cell polarity (PCP) pathway polarizes epithelia in the plane of a tissue. It regulates form and function of tissues and manifests itself by the polarized formation of cellular appendages such as epidermal hairs and cilia. Defects in the pathway are often associated with organ malformation and disease. In the kidney, the molecular events leading to cyst formation in polycystic kidney disease involve the PCP pathway. PCP is, however, best understood in Drosophila where genetic screens have identified a group of PCP core proteins including the Wnt receptor Frizzled (Fz), Dishevelled (Dsh), and Flamingo (Fmi). These proteins can localize to opposite parts of the plasma membrane in response to a poorly understood symmetry breaking event. Recent evidence suggests that proton transporters may play a role in regulating the asymmetric localization of PCP core proteins. Several papers have reported that the (pro)renin receptor, which is an associated subunit of the proton pumping V-ATPase, is required for PCP, but also for canonical Wnt signaling. Here, we discuss the implications of these findings for diverse developmental settings.

Mda-9/Syntenin Protein Positively Regulates the Activation of Akt Protein by Facilitating Integrin-linked Kinase Adaptor Function during Adhesion to Type I Collagen

The integrin-linked kinase (ILK)-PINCH1-α-parvin (IPP) complex functions as a signaling platform for integrins that modulates various cellular processes. ILK functions as a central adaptor for the assembly of IPP complex. We report here that mda-9/syntenin, a positive regulator of cancer metastasis, regulates the activation of Akt (also known as protein kinase B) by facilitating ILK adaptor function during adhesion to type I collagen (COL-I) in human breast cancer cells. COL-I stimulation induced the phosphorylation and plasma membrane translocation of Akt. Inhibition of mda-9/syntenin or expression of mutant ILK (E359K) significantly blocked the translocation of both ILK and Akt to the plasma membrane. mda-9/syntenin associated with ILK, and this association was increased at the plasma membrane by COL-I stimulation. Knockdown of mda-9/syntenin impaired COL-I-induced association of ILK with Akt and plasma membrane targeting of ILK-Akt complex. These results demonstrated that mda-9/syntenin regulates the activation of Akt by controlling the plasma membrane targeting of Akt via a mechanism that facilitates the association of Akt with ILK at the plasma membrane during adhesion to COL-I. On a striking note, inhibition of mda-9/syntenin impaired COL-I-induced plasma membrane translocation of the IPP complex and assembly of integrin β1-IPP signaling complexes. Thus, our study defines the role of mda-9/syntenin in ILK adaptor function and describes a new mechanism of mda-9/syntenin for regulation of cell migration.