bims-pideca Biomed News
on Class IA PI3K signalling in development and cancer
Issue of 2020‒09‒06
thirteen papers selected by
Ralitsa Radostinova Madsen
University College London Cancer Institute

  1. Cancer Res. 2020 Aug 31. pii: canres.0505.2020. [Epub ahead of print]
      KDM5B (lysine[K]-specific demethylase 5B) is frequently upregulated in various human cancers including prostate cancer (PCa). KDM5B controls H3K4me3/2 levels and regulates gene transcription and cell differentiation, yet the contributions of KDM5B to PCa tumorigenesis remains unknown. In this study, we investigated the functional role of KDM5B in epigenetic dysregulation and PCa progression in cultured cells and in mouse models of prostate epithelium-specific mutant Pten/Kdm5b. Kdm5b deficiency resulted in a significant delay in the onset of PCa in Pten-null mice, while Kdm5b loss alone caused no morphological abnormalities in mouse prostates. At 6 months of age, the prostate weight of Pten/Kdm5b mice was reduced by up to 70% compared to that of Pten mice. Pathological analysis revealed Pten/Kdm5b mice displayed mild morphological changes with hyperplasia in prostates, whereas age-matched Pten littermates developed high grade-prostatic intraepithelial neoplasia (HG-PIN) and PCa. Mechanistically, KDM5B governed phosphatidylinositol 3-kinase (PI3K)/AKT signaling in PCa in vitro and in vivo. KDM5B directly bound the PIK3CA promoter and KDM5B knockout resulted in a significant reduction of P110α and PIP3 levels and subsequent decrease in proliferation of human PCa cells. Conversely, KDM5B overexpression resulted in increased PI3K/AKT signaling. Loss of Kdm5b abrogated the hyperactivation of AKT signaling by decreasing P110α/P85 levels in Pten/Kdm5b mice. Taken together, our findings reveal that KDM5B acts as a key regulator of PI3K/AKT signaling; they also support the concept that targeting KDM5B is a novel and effective therapeutic strategy against PCa.
  2. Proc Natl Acad Sci U S A. 2020 Sep 02. pii: 202008980. [Epub ahead of print]
      Low-glucose and -insulin conditions, associated with ketogenic diets, can reduce the activity of the mechanistic target of rapamycin complex 1 (mTORC1) signaling pathway, potentially leading to a range of positive medical and health-related effects. Here, we determined whether mTORC1 signaling is also a target for decanoic acid, a key component of the medium-chain triglyceride (MCT) ketogenic diet. Using a tractable model system, Dictyostelium, we show that decanoic acid can decrease mTORC1 activity, under conditions of constant glucose and in the absence of insulin, measured by phosphorylation of eukaryotic translation initiation factor 4E-binding protein 1 (4E-BP1). We determine that this effect of decanoic acid is dependent on a ubiquitin regulatory X domain-containing protein, mediating inhibition of a conserved Dictyostelium AAA ATPase, p97, a homolog of the human transitional endoplasmic reticulum ATPase (VCP/p97) protein. We then demonstrate that decanoic acid decreases mTORC1 activity in the absence of insulin and under high-glucose conditions in ex vivo rat hippocampus and in tuberous sclerosis complex (TSC) patient-derived astrocytes. Our data therefore indicate that dietary decanoic acid may provide a new therapeutic approach to down-regulate mTORC1 signaling.
    Keywords:  Dictyostelium discoideum; decanoic acid; epilepsy; mTOR; tuberous sclerosis complex
  3. FEBS Open Bio. 2020 Sep 02.
      PI(3,4,5)P3 is required for AKT activation. The level of PI(3,4,5)P3 is constantly regulated through balanced synthesis by PI3 kinase and degradation by phosphoinositide phosphatases PTEN and SHIP2, known as negative regulators of AKT. Here, I show that SHIP2 inhibition in cervical cancer cell lines alters H2 O2 -mediated AKT and MAPK/ERK pathway activation. In addition, SHIP2 inhibition enhances reactive oxygen species generation. Interestingly, I found that SHIP2 inhibition and H2 O2 treatment enhance lipid and protein phosphatase activity of PTEN. Pharmacological targeting or RNAi-mediated knockdown of PTEN rescue ERK and AKT activation. Using a series of pharmacological and biochemical approaches, I provide evidence that crosstalk between SHIP2 and PTEN occurs upon an increase in oxidative stress to modulate the activity of MAPK and PI3/ATK pathways.
    Keywords:  PI3K/Akt pathway; PTEN; ROS; SHIP2; SHIP2 inhibitor
  4. J Biol Chem. 2020 Sep 02. pii: jbc.RA120.013121. [Epub ahead of print]
      Branched-chain α-keto acids (BCKAs) are catabolites of branched-chain amino acids (BCAAs). Intracellular BCKAs is cleared by branched-chain ketoacid dehydrogenase (BCKDH), which is sensitive to inhibitory phosphorylation by BCKD kinase (BCKDK). Accumulation of BCKAs is an indicator of defective BCAA catabolism and has been correlated with glucose intolerance and cardiac dysfunction. However, it is unclear whether BCKAs directly alter insulin signaling and function in the skeletal and cardiac muscle cell. Furthermore, the role of excess fatty acids (FA) in perturbing BCAA catabolism and BCKA availability merits investigation. By using immunoblot and UPLC MS/MS to analyze the hearts of fasted mice, we observed decreased BCAA catabolizing enzyme expression and increased circulating BCKAs, but not BCAAs. In mice subjected to diet-induced obesity (DIO), we observed similar increases in circulating BCKAs with concomitant changes in BCAA catabolizing enzyme expression only in the skeletal muscle. Effects of DIO were recapitulated by simulating lipotoxicity in skeletal muscle cells treated with saturated FA, palmitate. Exposure of muscle cells to high concentrations of BCKAs resulted in inhibition of insulin-induced AKT phosphorylation, decreased glucose uptake and mitochondrial oxygen consumption. Altering intracellular clearance of BCKAs by genetic modulation of BCKDK and BCKDHA expression showed similar effects on AKT phosphorylation. BCKAs increased protein translation and mTORC1 activation. Pretreating cells with mTORC1 inhibitor rapamycin restored BCKAs effect on insulin-induced AKT phosphorylation. This study provides evidence for FA mediated regulation of BCAA catabolizing enzymes, BCKA content and highlights the biological role of BCKAs in regulating muscle insulin signaling and function.
    Keywords:  BCKA; Insulin signaling; amino acid; cardiomyocyte; cardiomyocytes; insulin resistance; protein translation; skeletal muscle; skeletal muscle metabolism; translation
  5. Dev Cell. 2020 Sep 01. pii: S1534-5807(20)30599-2. [Epub ahead of print]
      The Ras/PI3K/extracellular signal-regulated kinases (ERK) signaling network plays fundamental roles in cell growth, survival, and migration and is frequently activated in cancer. Here, we show that the activities of the signaling network propagate as coordinated waves, biased by growth factor, which drive actin-based protrusions in human epithelial cells. The network exhibits hallmarks of biochemical excitability: the annihilation of oppositely directed waves, all-or-none responsiveness, and refractoriness. Abrupt perturbations to Ras, PI(4,5)P2, PI(3,4)P2, ERK, and TORC2 alter the threshold, observations that define positive and negative feedback loops within the network. Oncogenic transformation dramatically increases the wave activity, the frequency of ERK pulses, and the sensitivity to EGF stimuli. Wave activity was progressively enhanced across a series of increasingly metastatic breast cancer cell lines. The view that oncogenic transformation is a shift to a lower threshold of excitable Ras/PI3K/ERK network, caused by various combinations of genetic insults, can facilitate the assessment of cancer severity and effectiveness of interventions.
    Keywords:  ERK; PI(3,4)P2; PI(4,5)P2; PI3K; Ras; excitability; oncogenic transformation; threshold; wave
  6. J Biol Chem. 2020 Aug 31. pii: jbc.RA120.013634. [Epub ahead of print]
      Muscle atrophy is regulated by the balance between protein degradation and synthesis. Foxo1, a transcription factor, helps to determine this balance by activating pro-atrophic gene transcription when present in muscle fiber nuclei. Foxo1 nuclear efflux is promoted by Akt-mediated Foxo1 phosphorylation, eliminating Foxo1's atrophy promoting effect. Akt activation can be promoted by IGF1 or Insulin via a pathway including IGF1 or Insulin, PI3K, and Akt. We used confocal fluorescence time-lapse imaging of Foxo1-GFP in adult isolated living muscle fibers maintained in culture to explore the effects of IGF1 and insulin on Foxo1-GFP nuclear efflux with and without pharmacological inhibitors. We observed that while Akt inhibitor blocks the IGF1 or insulin induced effect on Foxo1 nuclear efflux, PI3K inhibitors, which we show to be effective in these fibers, do not. We also found that inhibition of the protein kinases Ack1 or ATM, contributes to the suppression of Foxo1 nuclear efflux after IGF1. These results indicate a novel pathway that has been unexplored in the IGF1 or insulin induced regulation of Foxo1 and present information useful for both therapeutic interventions for muscle atrophy, as well as further investigative areas into insulin insensitivity and type 2 diabetes.
    Keywords:  Akt PKB; FOXO; insulin; insulin-like growth factor (IGF); nuclear translocation; protein phosphorylation; skeletal muscle
  7. Nat Cell Biol. 2020 Sep;22(9): 1091-1102
      Organs and cells must adapt to shear stress induced by biological fluids, but how fluid flow contributes to the execution of specific cell programs is poorly understood. Here we show that shear stress favours mitochondrial biogenesis and metabolic reprogramming to ensure energy production and cellular adaptation in kidney epithelial cells. Shear stress stimulates lipophagy, contributing to the production of fatty acids that provide mitochondrial substrates to generate ATP through β-oxidation. This flow-induced process is dependent on the primary cilia located on the apical side of epithelial cells. The interplay between fluid flow and lipid metabolism was confirmed in vivo using a unilateral ureteral obstruction mouse model. Finally, primary cilium-dependent lipophagy and mitochondrial biogenesis are required to support energy-consuming cellular processes such as glucose reabsorption, gluconeogenesis and cytoskeletal remodelling. Our findings demonstrate how primary cilia and autophagy are involved in the translation of mechanical forces into metabolic adaptation.
  8. Nat Cancer. 2020 Apr;1(4): 382-393
      Alpelisib is a selective inhibitor of PI3Kα, shown to improve outcomes for PIK3CA mutant, hormone receptor positive (HR+) metastatic breast cancers (MBC) when combined with antiestrogen therapy. To uncover mechanisms of resistance, we conducted a detailed, longitudinal analysis of tumor and plasma circulating tumor DNA among such patients from a phase I/II trial combining alpelisib with an aromatase inhibitor (AI) (NCT01870505). The trial's primary objective was to establish safety with maculopapular rash emerging as the most common grade 3 adverse event (33%). Among 44 evaluable patients, the observed clinical benefit rate was 52%. Correlating genetic alterations with outcome, we identified loss-of-function PTEN mutations in 25% of patients with resistance. ESR1 activating mutations also expanded in number and allele fraction during treatment and were associated with resistance. These data indicate that genomic alterations that mediate resistance to alpelisib or antiestrogen may promote disease progression and highlight PTEN loss as a recurrent mechanism of resistance to PI3Kα inhibition.
  9. Oncogene. 2020 Sep 02.
      Angiogenesis inhibitors, such as the receptor tyrosine kinase (RTK) inhibitor sunitinib, target vascular endothelial growth factor (VEGF) signaling in cancers. However, only a fraction of patients respond, and most ultimately develop resistance to current angiogenesis inhibitor therapies. Activity of alternative pro-angiogenic growth factors, acting via RTK or G-protein coupled receptors (GPCR), may mediate VEGF inhibitor resistance. The phosphoinositide 3-kinase (PI3K)β isoform is uniquely coupled to both RTK and GPCRs. We investigated the role of endothelial cell (EC) PI3Kβ in tumor angiogenesis. Pro-angiogenic GPCR ligands were expressed by patient-derived renal cell carcinomas (PD-RCC), and selective inactivation of PI3Kβ reduced PD-RCC-stimulated EC spheroid sprouting. EC-specific PI3Kβ knockout (ΕC-βKO) in mice potentiated the sunitinib-induced reduction in subcutaneous growth of LLC1 and B16F10, and lung metastasis of B16F10 tumors. Compared to single-agent sunitinib treatment, tumors in sunitinib-treated ΕC-βKO mice showed a marked decrease in microvessel density, and reduced new vessel formation. The fraction of perfused mature tumor microvessels was increased in ΕC-βKO mice suggesting immature microvessels were most sensitive to combined sunitinib and PI3Kβ inactivation. Taken together, EC PI3Kβ inactivation with sunitinib inhibition reduces microvessel turnover and decreases heterogeneity of the tumor microenvironment, hence PI3Kβ inhibition may be a useful adjuvant antiangiogenesis therapy with sunitinib.
  10. Cell Syst. 2020 Aug 25. pii: S2405-4712(20)30287-8. [Epub ahead of print]
      A common approach to benchmarking of single-cell transcriptomics tools is to generate synthetic datasets that statistically resemble experimental data. However, most existing single-cell simulators do not incorporate transcription factor-gene regulatory interactions that underlie expression dynamics. Here, we present SERGIO, a simulator of single-cell gene expression data that models the stochastic nature of transcription as well as regulation of genes by multiple transcription factors according to a user-provided gene regulatory network. SERGIO can simulate any number of cell types in steady state or cells differentiating to multiple fates. We show that datasets generated by SERGIO are statistically comparable to experimental data generated by Illumina HiSeq2000, Drop-seq, Illumina 10X chromium, and Smart-seq. We use SERGIO to benchmark several single-cell analysis tools, including GRN inference methods, and identify Tcf7, Gata3, and Bcl11b as key drivers of T cell differentiation by performing in silico knockout experiments. SERGIO is freely available for download here:
    Keywords:  RNA velocity; benchmarking single-cell analysis tools; differentiation trajectories; gene regulatory networks; simulations; single-cell RNA-seq
  11. Cell Metab. 2020 Aug 26. pii: S1550-4131(20)30414-9. [Epub ahead of print]
      Nonalcoholic fatty liver disease is strongly associated with hepatic insulin resistance (HIR); however, the key lipid species and molecular mechanisms linking these conditions are widely debated. We developed a subcellular fractionation method to quantify diacylglycerol (DAG) stereoisomers and ceramides in the endoplasmic reticulum (ER), mitochondria, plasma membrane (PM), lipid droplets, and cytosol. Acute knockdown (KD) of diacylglycerol acyltransferase-2 in liver induced HIR in rats. This was due to PM sn-1,2-DAG accumulation, which promoted PKCϵ activation and insulin receptor kinase (IRK)-T1160 phosphorylation, resulting in decreased IRK-Y1162 phosphorylation. Liver PM sn-1,2-DAG content and IRK-T1160 phosphorylation were also higher in humans with HIR. In rats, liver-specific PKCϵ KD ameliorated high-fat diet-induced HIR by lowering IRK-T1160 phosphorylation, while liver-specific overexpression of constitutively active PKCϵ-induced HIR by promoting IRK-T1160 phosphorylation. These data identify PM sn-1,2-DAGs as the key pool of lipids that activate PKCϵ and that hepatic PKCϵ is both necessary and sufficient in mediating HIR.
    Keywords:  ceramides; dicylglycerols; hepatic glucose production; hepatic glycogen synthesis; hepatic insulin resistance; insulin receptor phosphorylation; liquid chromatography-tandem mass spectrometry; nonalcoholic fatty liver disease; protein kinase C-epsilon; type 2 diabetes
  12. Diabetes. 2020 Aug 31. pii: db200439. [Epub ahead of print]
      Skeletal muscle insulin resistance is a prominent early feature in the pathogenesis of type 2 diabetes (T2D). In attempt to overcome this defect, we generated mice overexpressing insulin receptors (IR) specifically in skeletal muscle (IRMOE). On normal chow, IRMOE mice have similar body weight as controls, but an increase in lean mass and glycolytic muscle fibers and reduced fat mass. IRMOE mice also show higher basal phosphorylation of IR, IRS-1 and Akt in muscle and improved glucose tolerance compared to controls. When challenged with high fat diet (HFD), IRMOE mice are protected from diet-induced obesity. This is associated with reduced inflammation in fat and liver, improved glucose tolerance and improved systemic insulin sensitivity. Surprisingly, however, in both chow and HFD-fed mice, insulin stimulated Akt phosphorylation is significantly reduced in muscle of IRMOE mice, indicating post-receptor insulin resistance. RNA sequencing reveals downregulation of several post-receptor signaling proteins that contribute to this resistance. Thus, enhancing early insulin signaling in muscle by overexpression of the insulin receptor protects mice from diet-induced obesity and its effects on glucose metabolism. However, chronic overstimulation of this pathway leads to post-receptor desensitization, indicating the critical balance between normal signaling and hyperstimulation of the insulin signaling pathway.
  13. Elife. 2020 Sep 02. pii: e58737. [Epub ahead of print]9
      Outer radial glial (oRG) cells are a population of neural stem cells prevalent in the developing human cortex that contribute to its cellular diversity and evolutionary expansion. The mammalian Target of Rapamycin (mTOR) signaling pathway is active in human oRG cells. Mutations in mTOR pathway genes are linked to a variety of neurodevelopmental disorders and malformations of cortical development. We find that dysregulation of mTOR signaling specifically affects oRG cells, but not other progenitor types, by changing the actin cytoskeleton through the activity of the Rho-GTPase, CDC42. These effects change oRG cellular morphology, migration, and mitotic behavior, but do not affect proliferation or cell fate. Thus, mTOR signaling can regulate the architecture of the developing human cortex by maintaining the cytoskeletal organization of oRG cells and the radial glia scaffold. Our study provides insight into how mTOR dysregulation may contribute to neurodevelopmental disease.
    Keywords:  human; human cortex; neuroscience; organoids; outer radial glia; regenerative medicine; stem cells