Article

PI3K/Akt signaling requires spatial compartmentalization in plasma membrane microdomains

Department of Pharmacology, The Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA.
Proceedings of the National Academy of Sciences (Impact Factor: 9.81). 08/2011; 108(35):14509-14. DOI: 10.1073/pnas.1019386108
Source: PubMed

ABSTRACT Spatial compartmentalization of signaling pathway components generally defines the specificity and enhances the efficiency of signal transduction. The phosphatidylinositol 3-kinase (PI3K)/Akt pathway is known to be compartmentalized within plasma membrane microdomains; however, the underlying mechanisms and functional impact of this compartmentalization are not well understood. Here, we show that phosphoinositide-dependent kinase 1 is activated in membrane rafts in response to growth factors, whereas the negative regulator of the pathway, phosphatase and tensin homolog deleted on chromosome 10 (PTEN), is primarily localized in nonraft regions. Alteration of this compartmentalization, either by genetic targeting or ceramide-induced recruitment of PTEN to rafts, abolishes the activity of the entire pathway. These findings reveal critical steps in raft-mediated PI3K/Akt activation and demonstrate the essential role of membrane microdomain compartmentalization in enabling PI3K/Akt signaling. They further suggest that dysregulation of this compartmentalization may underlie pathological complications such as insulin resistance.

0 Followers
 · 
124 Views
  • Source
    • "PI3K activates PKC f (Le Good et al., 1998) and PKC f inhibition using a specific blocker or siRNA against PKC f reversed the increase in K Ca 1.1 levels in the gastric SMCs of CerS2-null mice, suggesting that PI3K activation plays a critical role in K Ca 1.1 upregulation. PI3K activation by phosphorylation might be caused by a change in lipid rafts, which are associated with several signal transduction molecules including Src family kinases and small G proteins (Yoshizaki et al., 2008) and act as important regulators of PI3K (Gao et al., 2011). In addition, K Ca 1.1 is localized to caveolae (Brainard et al., 2005), a subset of lipid rafts, and has a caveolin-binding motif (Suzuki et al., 2013). "
    [Show abstract] [Hide abstract]
    ABSTRACT: KC a 1.1 regulates smooth muscle contractility by modulating membrane potential, and age-associated changes in KC a 1.1 expression may contribute to the development of motility disorders of the gastrointestinal tract. Sphingolipids (SLs) are important structural components of cellular membranes whose altered composition may affect KC a 1.1 expression. Thus, in this study, we examined whether altered SL composition due to aging may affect the contractility of gastric smooth muscle (GSM). We studied changes in ceramide synthases (CerS) and SL levels in the GSM of mice of varying ages and compared them with those in young CerS2-null mice. The levels of C16- and C18-ceramides, sphinganine, sphingosine, and sphingosine 1-phosphate were increased, and levels of C22, C24:1 and C24 ceramides were decreased in the GSM of both aged wild-type and young CerS2-null mice. The altered SL composition upregulated KC a 1.1 and increased KC a 1.1 currents, while no change was observed in KC a 1.1 channel activity. The upregulation of KC a 1.1 impaired intracellular Ca(2+) mobilization and decreased phosphorylated myosin light chain levels, causing GSM contractile dysfunction. Additionally, phosphoinositide 3-kinase, protein kinase Cζ , c-Jun N-terminal kinases, and nuclear factor kappa-B were found to be involved in KC a 1.1 upregulation. Our findings suggest that age-associated changes in SL composition or CerS2 ablation upregulate KC a 1.1 via the phosphoinositide 3-kinase/protein kinase Cζ /c-Jun N-terminal kinases/nuclear factor kappa-B-mediated pathway and impair Ca(2+) mobilization, which thereby induces the contractile dysfunction of GSM. CerS2-null mice exhibited similar effects to aged wild-type mice; therefore, CerS2-null mouse models may be utilized for investigating the pathogenesis of aging-associated motility disorders. © 2015 The Authors. Aging Cell published by the Anatomical Society and John Wiley & Sons Ltd.
    Aging cell 08/2015; DOI:10.1111/acel.12388 · 5.94 Impact Factor
  • Source
    • "This protocol was adapted from [51], [52]. Briefly, confluent transfected HeLa cells were washed with ice-cold PBS, scraped, collected in 1 mL of lysis buffer (10 mM Tris, pH 7.4, 150 mM NaCl, 5 mM EDTA, 2 mM PMSF, 20 µg/ml leupeptin, and 20 µg/ml aprotinin, 1% Triton X100) and left on ice for 1 h. "
    [Show abstract] [Hide abstract]
    ABSTRACT: We developed genetically-encoded fluorescent sensors based on Förster Resonance Energy Transfer to monitor phosphatidic acid (PA) fluctuations in the plasma membrane using Spo20 as PA-binding motif. Basal PA levels and phospholipase D activity varied in different cell types. In addition, stimuli that activate PA phosphatases, leading to lower PA levels, increased lamellipodia and filopodia formation. Lower PA levels were observed in the leading edge than in the trailing edge of migrating HeLa cells. In MSC80 and OLN93 cells, which are stable cell lines derived from Schwann cells and oligodendrocytes, respectively, a higher ratio of diacylglycerol to PA levels was demonstrated in the membrane processes involved in myelination, compared to the cell body. We propose that the PA sensors reported here are valuable tools to unveil the role of PA in a variety of intracellular signaling pathways.
    PLoS ONE 07/2014; 9(7):e102526. DOI:10.1371/journal.pone.0102526 · 3.23 Impact Factor
  • Source
    • "Since we observed IRS1-independent inhibition of Akt phosphorylation in response to NPC1 inhibition, a defect is likely to lie at the level of phosphatidylinositol-3-kinase (PI3K) or at the upstream kinases of Akt, PDK1 and/or mTORC2. A number of components of this signalling nexus have been localised to cholesterol-rich lipid raft domains including phosphatidylinostol-2-phosphate (PIP2) [35]–[37] the substrate for PI3K and both PDK1 [38] and mTORC2 [39]. Further, PTEN, a negative regulator of Akt phosphorylation has been reported to be excluded from these domains [38]. "
    [Show abstract] [Hide abstract]
    ABSTRACT: The Niemann-Pick disease, type C1 (NPC1) gene encodes a transmembrane protein involved in cholesterol efflux from the lysosome. SNPs within NPC1 have been associated with obesity and type 2 diabetes, and mice heterozygous or null for NPC1 are insulin resistant. However, the molecular mechanism underpinning this association is currently undefined. This study aimed to investigate the effects of inhibiting NPC1 function on insulin action in adipocytes. Both pharmacological and genetic inhibition of NPC1 impaired insulin action. This impairment was evident at the level of insulin signalling and insulin-mediated glucose transport in the short term and decreased GLUT4 expression due to reduced liver X receptor (LXR) transcriptional activity in the long-term. These data show that cholesterol homeostasis through NPC1 plays a crucial role in maintaining insulin action at multiple levels in adipocytes.
    PLoS ONE 04/2014; 9(4):e95598. DOI:10.1371/journal.pone.0095598 · 3.23 Impact Factor
Show more

Preview

Download
1 Download
Available from