bims-pideca Biomed News
on Class IA PI3K signalling in development and cancer
Issue of 2021‒08‒01
twenty papers selected by
Ralitsa Radostinova Madsen
University College London Cancer Institute
  1. A new layer of phosphoinositide-mediated allosteric regulation uncovered for SHIP2.
  2. Hepatic mTORC1 signaling activates ATF4 as part of its metabolic response to feeding and insulin.
  3. Ras/ERK and PI3K/AKT signaling differentially regulate oncogenic ERG mediated transcription in prostate cells.
  4. PIK3R3, part of the regulatory domain of PI3K, is upregulated in sarcoma stem-like cells and promotes invasion, migration, and chemotherapy resistance.
  5. Gαq activation modulates autophagy by promoting mTORC1 signaling.
  6. PIP4Ks impact on PI3K, FOXP3, and UHRF1 signaling and modulate human regulatory T cell proliferation and immunosuppressive activity.
  7. Targeting mTOR signaling overcomes acquired resistance to combined BRAF and MEK inhibition in BRAF-mutant melanoma.
  8. Sequential oncogenic mutations influence cell competition.
  9. Defective insulin-stimulated GLUT4 translocation in brown adipocytes induces systemic glucose homeostasis dysregulation independent of thermogenesis in female mice.
  10. Insulin signaling regulates longevity through protein phosphorylation in Caenorhabditis elegans.
  11. Targeting B-cell receptor and PI3K signaling in diffuse large B-cell lymphoma.
  12. Brain-specific inhibition of mTORC1 eliminates side effects resulting from mTORC1 blockade in the periphery and reduces alcohol intake in mice.
  13. The emerging role of the fetal insulin receptor (IR-A) in hormone-refractory breast cancer.
  14. Low-level mosaicism in tuberous sclerosis complex in four unrelated patients: Comparison of clinical characteristics and diagnostic pathways.
  15. Tissue clearing to examine tumour complexity in three dimensions.
  16. Stearic acid blunts growth-factor signaling via oleoylation of GNAI proteins.
  17. A single-cell type transcriptomics map of human tissues.
  18. Molecular control of cell density-mediated exit to quiescence.
  19. Counteracting age-related VEGF signaling insufficiency promotes healthy aging and extends life span.
  20. AKT3-mediated IWS1 phosphorylation promotes the proliferation of EGFR-mutant lung adenocarcinomas through cell cycle-regulated U2AF2 RNA splicing.
  1. FASEB J. 2021 08;35(8): e21815
    A new layer of phosphoinositide-mediated allosteric regulation uncovered for SHIP2.
    Johanne Le Coq, Pilar López Navajas, Bárbara Rodrigo Martin, Carlos Alfonso, Daniel Lietha.
      The Src homology 2 containing inositol 5-phosphatase 2 (SHIP2) is a large multidomain enzyme that catalyzes the dephosphorylation of the phospholipid phosphatidylinositol 3,4,5-triphosphate (PI(3,4,5)P3 ) to form PI(3,4)P2 . PI(3,4,5)P3 is a key lipid second messenger controlling the recruitment of signaling proteins to the plasma membrane, thereby regulating a plethora of cellular events, including proliferation, growth, apoptosis, and cytoskeletal rearrangements. SHIP2, alongside PI3K and PTEN, regulates PI(3,4,5)P3 levels at the plasma membrane and has been heavily implicated in serious diseases such as cancer and type 2 diabetes; however, many aspects of its regulation mechanism remain elusive. We recently reported an activating effect of the SHIP2 C2 domain and here we describe an additional layer of regulation via the pleckstrin homology-related (PHR) domain. We show a phosphoinositide-induced transition to a high activity state of the enzyme that increases phosphatase activity up to 10-15 fold. We further show that PI(3,4)P2 directly interacts with the PHR domain to trigger this allosteric activation. Modeling of the PHR-phosphatase-C2 region of SHIP2 on the membrane suggests no major inter-domain interactions with the PHR domain, but close contacts between the two linkers offer a possible path of allosteric communication. Together, our data show that the PHR domain acts as an allosteric module regulating the catalytic activity of SHIP2 in response to specific phosphoinositide levels in the cell membrane.
    Keywords:  SHIP2; allosteric regulation; enzyme kinetics; lipid phosphatase; phosphinositides
    DOI:  https://doi.org/10.1096/fj.202100561R
  2. Mol Metab. 2021 Jul 22. pii: S2212-8778(21)00156-3. [Epub ahead of print] 101309
    Hepatic mTORC1 signaling activates ATF4 as part of its metabolic response to feeding and insulin.
    Vanessa Byles, Yann Cormerais, Krystle Kalafut, Victor Barrera, James E Hughes Hallett, Shannan Ho Sui, John M Asara, Christopher M Adams, Gerta Hoxhaj, Issam Ben-Sahra, Brendan D Manning.
      OBJECTIVE: The mechanistic target of rapamycin complex 1 (mTORC1) is dynamically regulated by fasting and feeding cycles in the liver to promote protein and lipid synthesis while suppressing autophagy. However, beyond these functions, the metabolic response of the liver to feeding and insulin signaling orchestrated by mTORC1 remains poorly defined. Here, we determine whether ATF4, a stress responsive transcription factor recently found to be independently regulated by mTORC1 signaling in proliferating cells, is responsive to hepatic mTORC1 signaling to alter hepatocyte metabolism.METHODS: ATF4 protein levels and expression of canonical gene targets were analyzed in the liver following fasting and physiological feeding in the presence or absence of the mTORC1 inhibitor rapamycin. Primary hepatocytes from wild-type or liver-specific Atf4 knockout (LAtf4KO) mice were used to characterize the effects of insulin-stimulated mTORC1-ATF4 function on hepatocyte gene expression and metabolism. Both unbiased steady-state metabolomics and stable-isotope tracing methods were employed to define mTORC1 and ATF4-dependent metabolic changes. RNA-sequencing was used to determine global changes in feeding-induced transcripts in the livers of wild-type versus LAtf4KO mice.
    RESULTS: We demonstrate that ATF4 and its metabolic gene targets are stimulated by mTORC1 signaling in the liver in response to feeding and in a hepatocyte-intrinsic manner by insulin. While we demonstrate that de novo purine and pyrimidine synthesis is stimulated by insulin through mTORC1 signaling in primary hepatocytes, this regulation was independent of ATF4. Metabolomics and metabolite tracing studies revealed that insulin-mTORC1-ATF4 signaling stimulates pathways of non-essential amino acid synthesis in primary hepatocytes, including those of alanine, aspartate, methionine, and cysteine, but not serine.
    CONCLUSION: The results demonstrate that ATF4 is a novel metabolic effector of mTORC1 in liver, extending the molecular consequences of feeding and insulin-induced mTORC1 signaling in this key metabolic tissue to the control of amino acid metabolism.
    Keywords:  ATF4; feeding; insulin; liver; mTORC1; methionine metabolism
    DOI:  https://doi.org/10.1016/j.molmet.2021.101309
  3. PLoS Genet. 2021 Jul 27. 17(7): e1009708
    Ras/ERK and PI3K/AKT signaling differentially regulate oncogenic ERG mediated transcription in prostate cells.
    Brady G Strittmatter, Travis J Jerde, Peter C Hollenhorst.
      The TMPRSS2/ERG gene rearrangement occurs in 50% of prostate tumors and results in expression of the transcription factor ERG, which is normally silent in prostate cells. ERG expression promotes prostate tumor formation and luminal epithelial cell fates when combined with PI3K/AKT pathway activation, however the mechanism of synergy is not known. In contrast to luminal fates, expression of ERG alone in immortalized normal prostate epithelial cells promotes cell migration and epithelial to mesenchymal transition (EMT). Migration requires ERG serine 96 phosphorylation via endogenous Ras/ERK signaling. We found that a phosphomimetic mutant, S96E ERG, drove tumor formation and clonogenic survival without activated AKT. S96 was only phosphorylated on nuclear ERG, and differential recruitment of ERK to a subset of ERG-bound chromatin associated with ERG-activated, but not ERG-repressed genes. S96E did not alter ERG genomic binding, but caused a loss of ERG-mediated repression, EZH2 binding and H3K27 methylation. In contrast, AKT activation altered the ERG cistrome and promoted expression of luminal cell fate genes. These data suggest that, depending on AKT status, ERG can promote either luminal or EMT transcription programs, but ERG can promote tumorigenesis independent of these cell fates and tumorigenesis requires only the transcriptional activation function.
    DOI:  https://doi.org/10.1371/journal.pgen.1009708
  4. Cell Death Dis. 2021 Jul 29. 12(8): 749
    PIK3R3, part of the regulatory domain of PI3K, is upregulated in sarcoma stem-like cells and promotes invasion, migration, and chemotherapy resistance.
    Changhwan Yoon, Jun Lu, Sandra W Ryeom, M Celeste Simon, Sam S Yoon.
      To identify drivers of sarcoma cancer stem-like cells (CSCs), we compared gene expression using RNA sequencing between HT1080 fibrosarcoma and SK-LMS-1 leiomyosarcoma spheroids (which are enriched for CSCs) compared with the parent populations. The most overexpressed survival signaling-related gene in spheroids was phosphoinositide-3-kinase regulatory subunit 3 (PIK3R3), a regulatory subunit of PI3K, which functions in tumorigenesis and metastasis. In a human sarcoma microarray, PIK3R3 was also overexpressed by 4.1-fold compared with normal tissues. PIK3R3 inhibition using shRNA in the HT1080, SK-LMS-1, and DDLS8817 dedifferentiated liposarcoma in spheroids and in CD133+ cells (a CSC marker) reduced expression of CD133 and the stem cell factor Nanog and blocked spheroid formation by 61-71%. Mechanistic studies showed that in spheroid cells, PIK3R3 activated AKT and ERK signaling. Inhibition of PIK3R3, AKT, or ERK using shRNA or inhibitors decreased expression of Nanog, spheroid formation by 68-73%, and anchorage-independent growth by 76-91%. PIK3R3 or ERK1/2 inhibition similarly blocked sarcoma spheroid cell migration, invasion, secretion of MMP-2, xenograft invasion into adjacent normal tissue, and chemotherapy resistance. Together, these results show that signaling through the PIK3R3/ERK/Nanog axis promotes sarcoma CSC phenotypes such as migration, invasion, and chemotherapy resistance, and identify PIK3R3 as a potential therapeutic target in sarcoma.
    DOI:  https://doi.org/10.1038/s41419-021-04036-5
  5. Nat Commun. 2021 07 27. 12(1): 4540
    Gαq activation modulates autophagy by promoting mTORC1 signaling.
    Sofía Cabezudo, Maria Sanz-Flores, Alvaro Caballero, Inmaculada Tasset, Elena Rebollo, Antonio Diaz, Anna M Aragay, Ana María Cuervo, Federico Mayor, Catalina Ribas.
      The mTORC1 node plays a major role in autophagy modulation. We report a role of the ubiquitous Gαq subunit, a known transducer of plasma membrane G protein-coupled receptors signaling, as a core modulator of mTORC1 and autophagy. Cells lacking Gαq/11 display higher basal autophagy, enhanced autophagy induction upon different types of nutrient stress along with a decreased mTORC1 activation status. They are also unable to reactivate mTORC1 and thus inactivate ongoing autophagy upon nutrient recovery. Conversely, stimulation of Gαq/11 promotes sustained mTORC1 pathway activation and reversion of autophagy promoted by serum or amino acids removal. Gαq is present in autophagic compartments and lysosomes and is part of the mTORC1 multi-molecular complex, contributing to its assembly and activation via its nutrient status-sensitive interaction with p62, which displays features of a Gαq effector. Gαq emerges as a central regulator of the autophagy machinery required to maintain cellular homeostasis upon nutrient fluctuations.
    DOI:  https://doi.org/10.1038/s41467-021-24811-4
  6. Proc Natl Acad Sci U S A. 2021 Aug 03. pii: e2010053118. [Epub ahead of print]118(31):
    PIP4Ks impact on PI3K, FOXP3, and UHRF1 signaling and modulate human regulatory T cell proliferation and immunosuppressive activity.
    Alessandro Poli, Shidqiyyah Abdul-Hamid, Antonio Enrico Zaurito, Francesca Campagnoli, Valeria Bevilacqua, Bhavwanti Sheth, Roberta Fiume, Massimiliano Pagani, Sergio Abrignani, Nullin Divecha.
      Regulatory T cells (Tregs) play fundamental roles in maintaining peripheral tolerance to prevent autoimmunity and limit legitimate immune responses, a feature hijacked in tumor microenvironments in which the recruitment of Tregs often extinguishes immune surveillance through suppression of T-effector cell signaling and tumor cell killing. The pharmacological tuning of Treg activity without impacting on T conventional (Tconv) cell activity would likely be beneficial in the treatment of various human pathologies. PIP4K2A, 2B, and 2C constitute a family of lipid kinases that phosphorylate PtdIns5P to PtdIns(4,5)P 2 They are involved in stress signaling, act as synthetic lethal targets in p53-null tumors, and in mice, the loss of PIP4K2C leads to late onset hyperinflammation. Accordingly, a human single nucleotide polymorphism (SNP) near the PIP4K2C gene is linked with susceptibility to autoimmune diseases. How PIP4Ks impact on human T cell signaling is not known. Using ex vivo human primary T cells, we found that PIP4K activity is required for Treg cell signaling and immunosuppressive activity. Genetic and pharmacological inhibition of PIP4K in Tregs reduces signaling through the PI3K, mTORC1/S6, and MAPK pathways, impairs cell proliferation, and increases activation-induced cell death while sparing Tconv. PIP4K and PI3K signaling regulate the expression of the Treg master transcriptional activator FOXP3 and the epigenetic signaling protein Ubiquitin-like containing PHD and RING finger domains 1 (UHRF1). Our studies suggest that the pharmacological inhibition of PIP4K can reprogram human Treg identity while leaving Tconv cell signaling and T-helper differentiation to largely intact potentially enhancing overall immunological activity.
    Keywords:  Phosphatidylinositol 5-phosphate 4-kinase; T-regulatory cells; UHRF1; immunosuppression; phosphoinositide kinases
    DOI:  https://doi.org/10.1073/pnas.2010053118
  7. Oncogene. 2021 Jul 24.
    Targeting mTOR signaling overcomes acquired resistance to combined BRAF and MEK inhibition in BRAF-mutant melanoma.
    Beike Wang, Wei Zhang, Gao Zhang, Lawrence Kwong, Hezhe Lu, Jiufeng Tan, Norah Sadek, Min Xiao, Jie Zhang, Marilyne Labrie, Sergio Randell, Aurelie Beroard, Eric Sugarman, Vito W Rebecca, Zhi Wei, Yiling Lu, Gordon B Mills, Jeffrey Field, Jessie Villanueva, Xiaowei Xu, Meenhard Herlyn, Wei Guo.
      Targeting MAPK pathway using a combination of BRAF and MEK inhibitors is an efficient strategy to treat melanoma harboring BRAF-mutation. The development of acquired resistance is inevitable due to the signaling pathway rewiring. Combining western blotting, immunohistochemistry, and reverse phase protein array (RPPA), we aim to understanding the role of the mTORC1 signaling pathway, a center node of intracellular signaling network, in mediating drug resistance of BRAF-mutant melanoma to the combination of BRAF inhibitor (BRAFi) and MEK inhibitor (MEKi) therapy. The mTORC1 signaling pathway is initially suppressed by BRAFi and MEKi combination in melanoma but rebounds overtime after tumors acquire resistance to the combination therapy (CR) as assayed in cultured cells and PDX models. In vitro experiments showed that a subset of CR melanoma cells was sensitive to mTORC1 inhibition. The mTOR inhibitors, rapamycin and NVP-BEZ235, induced cell cycle arrest and apoptosis in CR cell lines. As a proof-of-principle, we demonstrated that rapamycin and NVP-BEZ235 treatment reduced tumor growth in CR xenograft models. Mechanistically, AKT or ERK contributes to the activation of mTORC1 in CR cells, depending on PTEN status of these cells. Our study reveals that mTOR activation is essential for drug resistance of melanoma to MAPK inhibitors, and provides insight into the rewiring of the signaling networks in CR melanoma.
    DOI:  https://doi.org/10.1038/s41388-021-01911-5
  8. Curr Biol. 2021 Jul 16. pii: S0960-9822(21)00893-9. [Epub ahead of print]
    Sequential oncogenic mutations influence cell competition.
    Koki Kohashi, Yusuke Mori, Rika Narumi, Kei Kozawa, Tomoko Kamasaki, Susumu Ishikawa, Mihoko Kajita, Rei Kobayashi, Yoichiro Tamori, Yasuyuki Fujita.
      At the initial stage of carcinogenesis, newly emerging transformed cells are often eliminated from epithelial layers via cell competition with the surrounding normal cells. For instance, when surrounded by normal cells, oncoprotein RasV12-transformed cells are extruded into the apical lumen of epithelia. During cancer development, multiple oncogenic mutations accumulate within epithelial tissues. However, it remains elusive whether and how cell competition is also involved in this process. In this study, using a mammalian cell culture model system, we have investigated what happens upon the consecutive mutations of Ras and tumor suppressor protein Scribble. When Ras mutation occurs under the Scribble-knockdown background, apical extrusion of Scribble/Ras double-mutant cells is strongly diminished. In addition, at the boundary with Scribble/Ras cells, Scribble-knockdown cells frequently undergo apoptosis and are actively engulfed by the neighboring Scribble/Ras cells. The comparable apoptosis and engulfment phenotypes are also observed in Drosophila epithelial tissues between Scribble/Ras double-mutant and Scribble single-mutant cells. Furthermore, mitochondrial membrane potential is enhanced in Scribble/Ras cells, causing the increased mitochondrial reactive oxygen species (ROS). Suppression of mitochondrial membrane potential or ROS production diminishes apoptosis and engulfment of the surrounding Scribble-knockdown cells, indicating that mitochondrial metabolism plays a key role in the competitive interaction between double- and single-mutant cells. Moreover, mTOR (mechanistic target of rapamycin kinase) acts downstream of these processes. These results imply that sequential oncogenic mutations can profoundly influence cell competition, a transition from loser to winner. Further studies would open new avenues for cell competition-based cancer treatment, thereby blocking clonal expansion of more malignant populations within tumors.
    Keywords:  ROS; RasV12; Scribble; apoptosis; cell competition; engulfment; entosis; mTOR; mitochondria; sequential mutations
    DOI:  https://doi.org/10.1016/j.cub.2021.06.064
  9. Mol Metab. 2021 Jul 21. pii: S2212-8778(21)00152-6. [Epub ahead of print] 101305
    Defective insulin-stimulated GLUT4 translocation in brown adipocytes induces systemic glucose homeostasis dysregulation independent of thermogenesis in female mice.
    Belén Picatoste, Lucie Yammine, Rosemary A Leahey, David Soares, Emma F Johnson, Paul Cohen, Timothy E McGraw.
      OBJECTIVE: Recent studies indicate that brown adipose tissue, in addition to its role in thermogenesis, has a role in the regulation of whole-body metabolism. Here we characterize the metabolic effects of deleting Rab10, a protein key for insulin stimulation of glucose uptake into white adipocytes, solely from brown adipocytes.METHODS: We used a murine brown adipocyte cell line and stromal vascular fraction-derived in vitro differentiated brown adipocytes to study the role of Rab10 in insulin-stimulated GLUT4 translocation to the plasma membrane and insulin-stimulated glucose uptake. We generated a brown adipocyte specific Rab10 knockout for in vivo studies of metabolism and thermoregulation.
    RESULTS: We demonstrate that deletion of Rab10 from brown adipocytes results in a two-fold reduction of insulin-stimulated glucose transport by reducing translocation of the GLUT4 glucose transporter to the plasma membrane, an effect linked to whole-body glucose intolerance and insulin-resistance in female mice. This effect on metabolism is independent of the thermogenic function of brown adipocytes, thereby revealing a metabolism-specific role for brown adipocytes in female mice. The reduced glucose uptake induced by Rab10 deletion disrupts ChREBP regulation of de novo lipogenesis (DNL) genes, providing a potential link between DNL in brown adipocytes and whole-body metabolic regulation in female mice. On the other hand, deletion of Rab10 from male mice does not induce systemic insulin resistance, although ChREBP regulation is disrupted.
    CONCLUSION: Our studies of Rab10 reveal a role for insulin-regulated glucose transport into brown adipocytes in whole-body metabolic homeostasis of female mice. Importantly, the contribution of brown adipocytes to whole-body metabolic regulation is independent of its role in thermogenesis. It is not clear whether the whole-body metabolic sexual dimorphism is because female mice are permissive to the effects of Rab10 deletion from brown adipocytes or male mice are resistant to the effect.
    Keywords:  Brown adipose tissue; GLUT4; Rab10; diabetes; glucose homeostasis
    DOI:  https://doi.org/10.1016/j.molmet.2021.101305
  10. Nat Commun. 2021 07 27. 12(1): 4568
    Insulin signaling regulates longevity through protein phosphorylation in Caenorhabditis elegans.
    Wen-Jun Li, Chen-Wei Wang, Li Tao, Yong-Hong Yan, Mei-Jun Zhang, Ze-Xian Liu, Yu-Xin Li, Han-Qing Zhao, Xue-Mei Li, Xian-Dong He, Yu Xue, Meng-Qiu Dong.
      Insulin/IGF-1 Signaling (IIS) is known to constrain longevity by inhibiting the transcription factor FOXO. How phosphorylation mediated by IIS kinases regulates lifespan beyond FOXO remains unclear. Here, we profile IIS-dependent phosphorylation changes in a large-scale quantitative phosphoproteomic analysis of wild-type and three IIS mutant Caenorhabditis elegans strains. We quantify more than 15,000 phosphosites and find that 476 of these are differentially phosphorylated in the long-lived daf-2/insulin receptor mutant. We develop a machine learning-based method to prioritize 25 potential lifespan-related phosphosites. We perform validations to show that AKT-1 pT492 inhibits DAF-16/FOXO and compensates the loss of daf-2 function, that EIF-2α pS49 potently inhibits protein synthesis and daf-2 longevity, and that reduced phosphorylation of multiple germline proteins apparently transmits reduced DAF-2 signaling to the soma. In addition, an analysis of kinases with enriched substrates detects that casein kinase 2 (CK2) subunits negatively regulate lifespan. Our study reveals detailed functional insights into longevity.
    DOI:  https://doi.org/10.1038/s41467-021-24816-z
  11. Blood. 2021 Jul 28. pii: blood.2020006784. [Epub ahead of print]
    Targeting B-cell receptor and PI3K signaling in diffuse large B-cell lymphoma.
    Wendan Xu, Philipp Berning, Georg Lenz.
      Diffuse large B-cell lymphoma (DLBCL) is a heterogeneous diagnostic category comprising distinct molecular subtypes characterized by diverse genetic aberrations that dictate patient outcome. As roughly one-third of DLBCL patients are not cured by current standard chemo-immunotherapy a better understanding of the molecular pathogenesis is warranted to improve outcome. B-cell receptor (BCR) signaling is crucial for the development, growth and survival of both normal and a substantial fraction of malignant B-cells. Various analyses revealed genetic alterations of central components of the BCR or its downstream signaling effectors in some subtypes of DLBCL. Thus, BCR signaling and the downstream NF-κB and PI3K cascades have been proposed as potential targets for the treatment of DLBCL patients. As one of the main effectors of BCR activation, PI3K mediated signals play a crucial role in the pathogenesis and survival of DLBCL. In this review, we summarize our current understanding of BCR signaling with a special focus on the PI3K pathway in DLBCL and how to utilize this knowledge therapeutically.
    DOI:  https://doi.org/10.1182/blood.2020006784
  12. Nat Commun. 2021 07 27. 12(1): 4407
    Brain-specific inhibition of mTORC1 eliminates side effects resulting from mTORC1 blockade in the periphery and reduces alcohol intake in mice.
    Yann Ehinger, Ziyang Zhang, Khanhky Phamluong, Drishti Soneja, Kevan M Shokat, Dorit Ron.
      Alcohol Use Disorder (AUD) affects a large portion of the population. Unfortunately, efficacious medications to treat the disease are limited. Studies in rodents suggest that mTORC1 plays a crucial role in mechanisms underlying phenotypes such as heavy alcohol intake, habit, and relapse. Thus, mTORC1 inhibitors, which are used in the clinic, are promising therapeutic agents to treat AUD. However, chronic inhibition of mTORC1 in the periphery produces undesirable side effects, which limit their potential use for the treatment of AUD. To overcome these limitations, we designed a binary drug strategy in which male mice were treated with the mTORC1 inhibitor RapaLink-1 together with a small molecule (RapaBlock) to protect mTORC1 activity in the periphery. We show that whereas RapaLink-1 administration blocked mTORC1 activation in the liver, RapaBlock abolished the inhibitory action of Rapalink-1. RapaBlock also prevented the adverse side effects produced by chronic inhibition of mTORC1. Importantly, co-administration of RapaLink-1 and RapaBlock inhibited alcohol-dependent mTORC1 activation in the nucleus accumbens and attenuated alcohol seeking and drinking.
    DOI:  https://doi.org/10.1038/s41467-021-24567-x
  13. Endocrinology. 2021 Jul 25. pii: bqab147. [Epub ahead of print]
    The emerging role of the fetal insulin receptor (IR-A) in hormone-refractory breast cancer.
    Tanvi Mathur, Douglas Yee.
      The type 1 insulin-like growth factor receptor (IGF-1R) is a transmembrane tyrosine kinase receptor and a mediator of the biologic effects of insulin-like growth factor-I and -II (IGF-I/II). Inhibitors of IGF-1R signaling were tested in clinical cancer trials aiming to assess the utility of this receptor as a therapeutic target; essentially all IGF-1R inhibitors failed to provide an additional benefit compared to standard-of-care therapy. In this review, we will evaluate the role the insulin receptor (IR) plays in mediating IGF signaling and subsequent metabolic and mitogenic effects as one possible reason for these failures. IR is expressed as two isoforms, with the fetal isoform IR-A derived from alternative splicing and loss of exon 11, the adult isoform (IR-B) includes this exon. Cancer frequently re-express fetal proteins and this appears to be the case in cancer with a re-expression of the fetal isoform and an increased IR-A: IR-B ratio. The biological effects of IR isoform signaling are complex and not completely understood although it has been suggested that IR-A could stimulate mitogenic signaling pathways, play a role in cancer cell stemness, and mediated tolerance to cancer therapies. From a clinical perspective, the IR-A overexpression in cancer may explain why targeting IGF-1R alone was not successful. However, given the predominance of IR-A expression in cancer, it may also be possible to develop isoform specific inhibitors and avoid the metabolic consequences of inhibiting IR-B. If such inhibitors could be developed, then IR-A expression could serve as a predictive biomarker and co-targeting IR-A and IGF-1R could provide a novel, more effective therapy method.
    Keywords:  breast cancer; cancer cell stemness; insulin; insulin receptor; insulin-like growth factors; type I insulin-like growth factor
    DOI:  https://doi.org/10.1210/endocr/bqab147
  14. Am J Med Genet A. 2021 Jul 30.
    Low-level mosaicism in tuberous sclerosis complex in four unrelated patients: Comparison of clinical characteristics and diagnostic pathways.
    Héctor Hugo Manzanilla-Romero, Denisa Weis, Simon Schnaiter, Sabine Rudnik-Schöneborn.
      Tuberous sclerosis complex (TSC) is an autosomal dominant neurocutaneous syndrome caused by either TSC1 or TSC2 gene mutations. About 15% of TSC patients remain without genetic diagnosis by conventional analysis despite clinical evidence. It is important to identify somatic mosaics, as therapeutic options are now available in patients with TSC1 or TSC2 mutations. Here, we describe the clinical and genetic characteristics of four male TSC patients with low-level mosaicism. Patients presented at ages between 9 months and 32 years. Clinical manifestations varied considerably and included brain lesions in all four patients, cardiac rhabdomyomas in two young patients, skin involvement in two patients, and retinal hamartomas and renal angiomyolipomas in three patients. One patient presented with epileptic seizures and psychomotor delay. Low levels of mosaicism for TSC1 or TSC2 mutation were found in different tissue samples employing next generation sequencing and multiple ligation-dependent probe amplification. The five disease-associated variants, including one second-hit mutation, include three truncating mutations and one deletion in TSC2, and one truncating mutation in TSC1. Sanger sequencing, allele-specific oligonucleotide PCR (ASO-PCR), and droplet digital PCR were used to confirm and quantify the disclosed mutations. Genetic identification of low-level mosaicism for TSC remains challenging but is important for optimal surveillance and management.
    Keywords:  genetic diagnosis; next generation sequencing; somatic mosaicism; tuberous sclerosis complex
    DOI:  https://doi.org/10.1002/ajmg.a.62433
  15. Nat Rev Cancer. 2021 Jul 30.
    Tissue clearing to examine tumour complexity in three dimensions.
    Jorge Almagro, Hendrik A Messal, May Zaw Thin, Jacco van Rheenen, Axel Behrens.
      The visualization of whole organs and organisms through tissue clearing and fluorescence volumetric imaging has revolutionized the way we look at biological samples. Its application to solid tumours is changing our perception of tumour architecture, revealing signalling networks and cell interactions critical in tumour progression, and provides a powerful new strategy for cancer diagnostics. This Review introduces the latest advances in tissue clearing and three-dimensional imaging, examines the challenges in clearing epithelia - the tissue of origin of most malignancies - and discusses the insights that tissue clearing has brought to cancer research, as well as the prospective applications to experimental and clinical oncology.
    DOI:  https://doi.org/10.1038/s41568-021-00382-w
  16. Nat Commun. 2021 07 28. 12(1): 4590
    Stearic acid blunts growth-factor signaling via oleoylation of GNAI proteins.
    Hana Nůsková, Marina V Serebryakova, Anna Ferrer-Caelles, Timo Sachsenheimer, Christian Lüchtenborg, Aubry K Miller, Britta Brügger, Larisa V Kordyukova, Aurelio A Teleman.
      Covalent attachment of C16:0 to proteins (palmitoylation) regulates protein function. Proteins are also S-acylated by other fatty acids including C18:0. Whether protein acylation with different fatty acids has different functional outcomes is not well studied. We show here that C18:0 (stearate) and C18:1 (oleate) compete with C16:0 to S-acylate Cys3 of GNAI proteins. C18:0 becomes desaturated so that C18:0 and C18:1 both cause S-oleoylation of GNAI. Exposure of cells to C16:0 or C18:0 shifts GNAI acylation towards palmitoylation or oleoylation, respectively. Oleoylation causes GNAI proteins to shift out of cell membrane detergent-resistant fractions where they potentiate EGFR signaling. Consequently, exposure of cells to C18:0 reduces recruitment of Gab1 to EGFR and reduces AKT activation. This provides a molecular mechanism for the anti-tumor effects of C18:0, uncovers a mechanistic link how metabolites affect cell signaling, and provides evidence that the identity of the fatty acid acylating a protein can have functional consequences.
    DOI:  https://doi.org/10.1038/s41467-021-24844-9
  17. Sci Adv. 2021 Jul;pii: eabh2169. [Epub ahead of print]7(31):
    A single-cell type transcriptomics map of human tissues.
    Max Karlsson, Cheng Zhang, Loren Méar, Wen Zhong, Andreas Digre, Borbala Katona, Evelina Sjöstedt, Lynn Butler, Jacob Odeberg, Philip Dusart, Fredrik Edfors, Per Oksvold, Kalle von Feilitzen, Martin Zwahlen, Muhammad Arif, Ozlem Altay, Xiangyu Li, Mehmet Ozcan, Adil Mardonoglu, Linn Fagerberg, Jan Mulder, Yonglun Luo, Fredrik Ponten, Mathias Uhlén, Cecilia Lindskog.
      Advances in molecular profiling have opened up the possibility to map the expression of genes in cells, tissues, and organs in the human body. Here, we combined single-cell transcriptomics analysis with spatial antibody-based protein profiling to create a high-resolution single-cell type map of human tissues. An open access atlas has been launched to allow researchers to explore the expression of human protein-coding genes in 192 individual cell type clusters. An expression specificity classification was performed to determine the number of genes elevated in each cell type, allowing comparisons with bulk transcriptomics data. The analysis highlights distinct expression clusters corresponding to cell types sharing similar functions, both within the same organs and between organs.
    DOI:  https://doi.org/10.1126/sciadv.abh2169
  18. Cell Rep. 2021 Jul 27. pii: S2211-1247(21)00853-6. [Epub ahead of print]36(4): 109436
    Molecular control of cell density-mediated exit to quiescence.
    Yilin Fan, Tobias Meyer.
      Contact inhibition of cell proliferation regulates tissue size and prevents uncontrolled cell expansion. When cell density increases, contact inhibition can force proliferating cells into quiescence. Here we show that the variable memory of local cell density experienced by a mother cell controls the levels of the cyclin-dependent kinase (CDK) activator cyclin D1 and inhibitor p27 in newborn daughters, which direct cells to proliferation or quiescence. Much of this regulation can be explained by rapid suppression of ERK activity by high cell density in mothers, which leads to lower cyclin D1 and higher p27 levels in daughters. Strikingly, cell density and mitogen signals compete by shifting the ratio of cyclin D1/p27 levels below or above a single sharp threshold that controls the proliferation decision. Thus, the history of competing cell density and mitogen signals experienced by mothers is funneled into a precise activator-inhibitor balance that decides the fate of daughter cells.
    Keywords:  CDK; ERK; cell cycle; cell density; cell proliferation; contact inhibition; quiescence; ultrasensitivity
    DOI:  https://doi.org/10.1016/j.celrep.2021.109436
  19. Science. 2021 07 30. pii: eabc8479. [Epub ahead of print]373(6554):
    Counteracting age-related VEGF signaling insufficiency promotes healthy aging and extends life span.
    M Grunewald, S Kumar, H Sharife, E Volinsky, A Gileles-Hillel, T Licht, A Permyakova, L Hinden, S Azar, Y Friedmann, P Kupetz, R Tzuberi, A Anisimov, K Alitalo, M Horwitz, S Leebhoff, O Z Khoma, R Hlushchuk, V Djonov, R Abramovitch, J Tam, E Keshet.
      Aging is an established risk factor for vascular diseases, but vascular aging itself may contribute to the progressive deterioration of organ function. Here, we show in aged mice that vascular endothelial growth factor (VEGF) signaling insufficiency, which is caused by increased production of decoy receptors, may drive physiological aging across multiple organ systems. Increasing VEGF signaling prevented age-associated capillary loss, improved organ perfusion and function, and extended life span. Healthier aging was evidenced by favorable metabolism and body composition and amelioration of aging-associated pathologies including hepatic steatosis, sarcopenia, osteoporosis, "inflammaging" (age-related multiorgan chronic inflammation), and increased tumor burden. These results indicate that VEGF signaling insufficiency affects organ aging in mice and suggest that modulating this pathway may result in increased mammalian life span and improved overall health.
    DOI:  https://doi.org/10.1126/science.abc8479
  20. Nat Commun. 2021 Jul 30. 12(1): 4624
    AKT3-mediated IWS1 phosphorylation promotes the proliferation of EGFR-mutant lung adenocarcinomas through cell cycle-regulated U2AF2 RNA splicing.
    Georgios I Laliotis, Evangelia Chavdoula, Maria D Paraskevopoulou, Abdul Kaba, Alessandro La Ferlita, Satishkumar Singh, Vollter Anastas, Keith A Nair, Arturo Orlacchio, Vasiliki Taraslia, Ioannis Vlachos, Marina Capece, Artemis Hatzigeorgiou, Dario Palmieri, Christos Tsatsanis, Salvatore Alaimo, Lalit Sehgal, David P Carbone, Vincenzo Coppola, Philip N Tsichlis.
      AKT-phosphorylated IWS1 regulates alternative RNA splicing via a pathway that is active in lung cancer. RNA-seq studies in lung adenocarcinoma cells lacking phosphorylated IWS1, identified a exon 2-deficient U2AF2 splice variant. Here, we show that exon 2 inclusion in the U2AF2 mRNA is a cell cycle-dependent process that is regulated by LEDGF/SRSF1 splicing complexes, whose assembly is controlled by the IWS1 phosphorylation-dependent deposition of histone H3K36me3 marks in the body of target genes. The exon 2-deficient U2AF2 mRNA encodes a Serine-Arginine-Rich (RS) domain-deficient U2AF65, which is defective in CDCA5 pre-mRNA processing. This results in downregulation of the CDCA5-encoded protein Sororin, a phosphorylation target and regulator of ERK, G2/M arrest and impaired cell proliferation and tumor growth. Analysis of human lung adenocarcinomas, confirmed activation of the pathway in EGFR-mutant tumors and showed that pathway activity correlates with tumor stage, histologic grade, metastasis, relapse after treatment, and poor prognosis.
    DOI:  https://doi.org/10.1038/s41467-021-24795-1
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