Publications by authors named "Anita Morén"

13 Publications

  • Page 1 of 1

The noncoding MIR100HG RNA enhances the autocrine function of transforming growth factor β signaling.

Oncogene 2021 May 4;40(21):3748-3765. Epub 2021 May 4.

Department of Medical Biochemistry and Microbiology, Science for Life Laboratory, Box 582, Biomedical Center, Uppsala University, Uppsala, Sweden.

Activation of the transforming growth factor β (TGFβ) pathway modulates the expression of genes involved in cell growth arrest, motility, and embryogenesis. An expression screen for long noncoding RNAs indicated that TGFβ induced mir-100-let-7a-2-mir-125b-1 cluster host gene (MIR100HG) expression in diverse cancer types, thus confirming an earlier demonstration of TGFβ-mediated transcriptional induction of MIR100HG in pancreatic adenocarcinoma. MIR100HG depletion attenuated TGFβ signaling, expression of TGFβ-target genes, and TGFβ-mediated cell cycle arrest. Moreover, MIR100HG silencing inhibited both normal and cancer cell motility and enhanced the cytotoxicity of cytostatic drugs. MIR100HG overexpression had an inverse impact on TGFβ signaling responses. Screening for downstream effectors of MIR100HG identified the ligand TGFβ1. MIR100HG and TGFB1 mRNA formed ribonucleoprotein complexes with the RNA-binding protein HuR, promoting TGFβ1 cytokine secretion. In addition, TGFβ regulated let-7a-2-3p, miR-125b-5p, and miR-125b-1-3p expression, all encoded by MIR100HG intron-3. Certain intron-3 miRNAs may be involved in TGFβ/SMAD-mediated responses (let-7a-2-3p) and others (miR-100, miR-125b) in resistance to cytotoxic drugs mediated by MIR100HG. In support of a model whereby TGFβ induces MIR100HG, which then enhances TGFβ1 secretion, analysis of human carcinomas showed that MIR100HG expression correlated with expression of TGFB1 and its downstream extracellular target TGFBI. Thus, MIR100HG controls the magnitude of TGFβ signaling via TGFβ1 autoinduction and secretion in carcinomas.
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http://dx.doi.org/10.1038/s41388-021-01803-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8154591PMC
May 2021

The TGFB2-AS1 lncRNA Regulates TGF-β Signaling by Modulating Corepressor Activity.

Cell Rep 2019 Sep;28(12):3182-3198.e11

Department of Medical Biochemistry and Microbiology, Science for Life Laboratory, and Ludwig Cancer Research Box 582, Biomedical Center, Uppsala University, 751 23 Uppsala, Sweden. Electronic address:

Molecular processes involving lncRNAs regulate cell function. By applying transcriptomics, we identify lncRNAs whose expression is regulated by transforming growth factor β (TGF-β). Upon silencing individual lncRNAs, we identify several that regulate TGF-β signaling. Among these lncRNAs, TGFB2-antisense RNA1 (TGFB2-AS1) is induced by TGF-β through Smad and protein kinase pathways and resides in the nucleus. Depleting TGFB2-AS1 enhances TGF-β/Smad-mediated transcription and expression of hallmark TGF-β-target genes. Increased dose of TGFB2-AS1 reduces expression of these genes, attenuates TGF-β-induced cell growth arrest, and alters BMP and Wnt pathway gene profiles. Mechanistically, TGFB2-AS1, mainly via its 3' terminal region, binds to the EED adaptor of the Polycomb repressor complex 2 (PRC2), promoting repressive histone H3K27me modifications at TGF-β-target gene promoters. Silencing EED or inhibiting PRC2 methylation activity partially rescues TGFB2-AS1-mediated gene repression. Thus, the TGF-β-induced TGFB2-AS1 lncRNA exerts inhibitory functions on TGF-β/BMP signaling output, supporting auto-regulatory negative feedback that balances TGF-β/BMP-mediated responses.
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http://dx.doi.org/10.1016/j.celrep.2019.08.028DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6859500PMC
September 2019

LXRα limits TGFβ-dependent hepatocellular carcinoma associated fibroblast differentiation.

Oncogenesis 2019 May 16;8(6):36. Epub 2019 May 16.

Department of Medical Biochemistry and Microbiology, Science for Life Laboratory, Uppsala University, Box 582, Biomedical Center, SE-751 23, Uppsala, Sweden.

Transforming growth factor β (TGFβ) is deposited in the extracellular space of diverse tissues. Resident fibroblasts respond to TGFβ and undergo myofibroblastic differentiation during tissue wound healing and cancer progression. Cancer-associated fibroblasts (CAFs) communicate with tumor cells during cancer progression, under the guidance of TGFβ signaling. We report that agonist-activated liver X receptors (LXR) limit the expression of key components of myofibroblast differentiation, including the α-smooth muscle actin (αSMA) gene in liver cancer cells. CAFs derived from hepatocellular carcinoma (HCC) express high αSMA and low LXRα levels, whereas hepatocarcinoma cells exhibit an inverse expression pattern. All hepatoma cells analyzed responded to the LXRα agonist T0901317 by inducing fatty acid synthase (FASN) expression. On the other hand, T0901317 antagonized TGFβ-induced fibroblastic marker responses, such as fibronectin and calponin, in a subset of hepatoma cells and all CAFs analyzed. Mechanistically, LXRα antagonized TGFβ signaling at the transcriptional level. Smad3 and LXRα were recruited to adjacent DNA motifs of the ACTA2 promoter. Upon cloning the human ACTA2 promoter, we confirmed its transcriptional induction by TGFβ stimulation, and LXRα overexpression repressed the promoter activity. Hepatosphere formation by HCC cells was enhanced upon co-culturing with CAFs. T0901317 suppressed the positive effects exerted on hepatosphere growth by CAFs. Taken together, the data suggest that LXRα agonists limit TGFβ-dependent CAF differentiation, potentially limiting primary HCC growth.
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http://dx.doi.org/10.1038/s41389-019-0140-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6522550PMC
May 2019

Genomewide binding of transcription factor Snail1 in triple-negative breast cancer cells.

Mol Oncol 2018 06 21;12(7):1153-1174. Epub 2018 May 21.

Department of Medical Biochemistry and Microbiology, Science for Life Laboratory, Ludwig Institute for Cancer Research, Uppsala University, Sweden.

Transcriptional regulation mediated by the zinc finger protein Snail1 controls early embryogenesis. By binding to the epithelial tumor suppressor CDH1 gene, Snail1 initiates the epithelial-mesenchymal transition (EMT). The EMT generates stem-like cells and promotes invasiveness during cancer progression. Accordingly, Snail1 mRNA and protein is abundantly expressed in triple-negative breast cancers with enhanced metastatic potential and phenotypic signs of the EMT. Such high endogenous Snail1 protein levels permit quantitative chromatin immunoprecipitation-sequencing (ChIP-seq) analysis. Snail1 associated with 185 genes at cis regulatory regions in the Hs578T triple-negative breast cancer cell model. These genes include morphogenetic regulators and signaling components that control polarized differentiation. Using the CRISPR/Cas9 system in Hs578T cells, a double deletion of 10 bp each was engineered into the first exon and into the second exon-intron junction of Snail1, suppressing Snail1 expression and causing misregulation of several hundred genes. Specific attention to regulators of chromatin organization provides a possible link to new phenotypes uncovered by the Snail1 loss-of-function mutation. On the other hand, genetic inactivation of Snail1 was not sufficient to establish a full epithelial transition to these tumor cells. Thus, Snail1 contributes to the malignant phenotype of breast cancer cells via diverse new mechanisms.
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http://dx.doi.org/10.1002/1878-0261.12317DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6026864PMC
June 2018

Snail regulates BMP and TGFβ pathways to control the differentiation status of glioma-initiating cells.

Oncogene 2018 05 16;37(19):2515-2531. Epub 2018 Feb 16.

Department of Medical Biochemistry and Microbiology, Science for Life Laboratory, Box 582, Biomedical Center, Uppsala University, SE-75123, Uppsala, Sweden.

Glioblastoma multiforme is a brain malignancy characterized by high heterogeneity, invasiveness, and resistance to current therapies, attributes related to the occurrence of glioma stem cells (GSCs). Transforming growth factor β (TGFβ) promotes self-renewal and bone morphogenetic protein (BMP) induces differentiation of GSCs. BMP7 induces the transcription factor Snail to promote astrocytic differentiation in GSCs and suppress tumor growth in vivo. We demonstrate that Snail represses stemness in GSCs. Snail interacts with SMAD signaling mediators, generates a positive feedback loop of BMP signaling and transcriptionally represses the TGFB1 gene, decreasing TGFβ1 signaling activity. Exogenous TGFβ1 counteracts Snail function in vitro, and in vivo promotes proliferation and re-expression of Nestin, confirming the importance of TGFB1 gene repression by Snail. In conclusion, novel insight highlights mechanisms whereby Snail differentially regulates the activity of the opposing BMP and TGFβ pathways, thus promoting an astrocytic fate switch and repressing stemness in GSCs.
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http://dx.doi.org/10.1038/s41388-018-0136-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5945579PMC
May 2018

Chemical regulators of epithelial plasticity reveal a nuclear receptor pathway controlling myofibroblast differentiation.

Sci Rep 2016 07 19;6:29868. Epub 2016 Jul 19.

Ludwig Institute for Cancer Research, Science for Life Laboratory, Uppsala University, Box 595, Biomedical Center, SE-751 24 Uppsala, Sweden.

Plasticity in epithelial tissues relates to processes of embryonic development, tissue fibrosis and cancer progression. Pharmacological modulation of epithelial transitions during disease progression may thus be clinically useful. Using human keratinocytes and a robotic high-content imaging platform, we screened for chemical compounds that reverse transforming growth factor β (TGF-β)-induced epithelial-mesenchymal transition. In addition to TGF-β receptor kinase inhibitors, we identified small molecule epithelial plasticity modulators including a naturally occurring hydroxysterol agonist of the liver X receptors (LXRs), members of the nuclear receptor transcription factor family. Endogenous and synthetic LXR agonists tested in diverse cell models blocked α-smooth muscle actin expression, myofibroblast differentiation and function. Agonist-dependent LXR activity or LXR overexpression in the absence of ligand counteracted TGF-β-mediated myofibroblast terminal differentiation and collagen contraction. The protective effect of LXR agonists against TGF-β-induced pro-fibrotic activity raises the possibility that anti-lipidogenic therapy may be relevant in fibrotic disorders and advanced cancer.
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http://dx.doi.org/10.1038/srep29868DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4949434PMC
July 2016

Single Chain Antibodies as Tools to Study transforming growth factor-β-Regulated SMAD Proteins in Proximity Ligation-Based Pharmacological Screens.

Mol Cell Proteomics 2016 06 29;15(6):1848-56. Epub 2016 Feb 29.

From the ‡Dept. of Immunology, Genetics and Pathology, Science for Life Laboratory, Uppsala University, Uppsala SE-751 85, Sweden;

The cellular heterogeneity seen in tumors, with subpopulations of cells capable of resisting different treatments, renders single-treatment regimens generally ineffective. Accordingly, there is a great need to increase the repertoire of drug treatments from which combinations may be selected to efficiently target sets of pathological processes, while suppressing the emergence of resistance mutations. In this regard, members of the TGF-β signaling pathway may furnish new, valuable therapeutic targets. In the present work, we developed in situ proximity ligation assays (isPLA) to monitor the state of the TGF-β signaling pathway. Moreover, we extended the range of suitable affinity reagents for this analysis by developing a set of in-vitro-derived human antibody fragments (single chain fragment variable, scFv) that bind SMAD2 (Mothers against decapentaplegic 2), 3, 4, and 7 using phage display. These four proteins are all intracellular mediators of TGF-β signaling. We also developed an scFv specific for SMAD3 phosphorylated in the linker domain 3 (p179 SMAD3). This phosphorylation has been shown to inactivate the tumor suppressor function of SMAD3. The single chain affinity reagents developed in the study were fused tocrystallizable antibody fragments (Fc-portions) and expressed as dimeric IgG-like molecules having Fc domains (Yumabs), and we show that they represent valuable reagents for isPLA.Using these novel assays, we demonstrate that p179 SMAD3 forms a complex with SMAD4 at increased frequency during division and that pharmacological inhibition of cyclin-dependent kinase 4 (CDK4)(1) reduces the levels of p179SMAD3 in tumor cells. We further show that the p179SMAD3-SMAD4 complex is bound for degradation by the proteasome. Finally, we developed a chemical screening strategy for compounds that reduce the levels of p179SMAD3 in tumor cells with isPLA as a read-out, using the p179SMAD3 scFv SH544-IIC4. The screen identified two kinase inhibitors, known inhibitors of the insulin receptor, which decreased levels of p179SMAD3/SMAD4 complexes, thereby demonstrating the suitability of the recombinant affinity reagents applied in isPLA in screening for inhibitors of cell signaling.
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http://dx.doi.org/10.1074/mcp.M115.055756DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5083110PMC
June 2016

The protein kinase LKB1 negatively regulates bone morphogenetic protein receptor signaling.

Oncotarget 2016 Jan;7(2):1120-43

Ludwig Institute for Cancer Research, Science for Life Laboratory, Uppsala University, Uppsala, Sweden.

The protein kinase LKB1 regulates cell metabolism and growth and is implicated in intestinal and lung cancer. Bone morphogenetic protein (BMP) signaling regulates cell differentiation during development and tissue homeostasis. We demonstrate that LKB1 physically interacts with BMP type I receptors and requires Smad7 to promote downregulation of the receptor. Accordingly, LKB1 suppresses BMP-induced osteoblast differentiation and affects BMP signaling in Drosophila wing longitudinal vein morphogenesis. LKB1 protein expression and Smad1 phosphorylation analysis in a cohort of non-small cell lung cancer patients demonstrated a negative correlation predominantly in a subset enriched in adenocarcinomas. Lung cancer patient data analysis indicated strong correlation between LKB1 loss-of-function mutations and high BMP2 expression, and these two events further correlated with expression of a gene subset functionally linked to apoptosis and migration. This new mechanism of BMP receptor regulation by LKB1 has ramifications in physiological organogenesis and disease.
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http://dx.doi.org/10.18632/oncotarget.6683DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4811448PMC
January 2016

Negative regulation of TGFβ signaling by the kinase LKB1 and the scaffolding protein LIP1.

J Biol Chem 2011 Jan 25;286(1):341-53. Epub 2010 Oct 25.

Ludwig Institute for Cancer Research, Uppsala University, Box 595, Biomedical Center, Uppsala University, SE-751 24 Uppsala, Sweden.

Signal transduction by the Smad pathway elicits critical biological responses to many extracellular polypeptide factors, including TGFβ and bone morphogenetic protein. Regulation of Smad signaling imparts several cytoplasmic and nuclear mechanisms, some of which entail protein phosphorylation. Previous work established a protein complex between Smad4 and the scaffolding protein LKB1-interacting protein 1 (LIP1). LKB1 is a well studied tumor suppressor kinase that regulates cell growth and polarity. Here, we analyzed the LKB1-LIP1 and the Smad4-LIP1 protein complexes and found that LIP1 can self-oligomerize. We further demonstrate that LKB1 is capable of phosphorylating Smad4 on Thr(77) of its DNA-binding domain. LKB1 inhibits Smad4 from binding to either TGFβ- or bone morphogenetic protein-specific promoter sequences, which correlates with the negative regulatory effect LKB1 exerts on Smad4-dependent transcription. Accordingly, LKB1 negatively regulates TGFβ gene responses and epithelial-mesenchymal transition. Thus, LKB1 and LIP1 provide negative control of TGFβ signaling.
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http://dx.doi.org/10.1074/jbc.M110.190660DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3012991PMC
January 2011

Transforming growth factor beta promotes complexes between Smad proteins and the CCCTC-binding factor on the H19 imprinting control region chromatin.

J Biol Chem 2010 Jun 28;285(26):19727-37. Epub 2010 Apr 28.

Ludwig Institute for Cancer Research, Uppsala University, SE-751 24 Uppsala, Sweden.

Whether signal transduction pathways regulate epigenetic states in response to environmental cues remains poorly understood. We demonstrate here that Smad3, signaling downstream of transforming growth factor beta, interacts with the zinc finger domain of CCCTC-binding factor (CTCF), a nuclear protein known to act as "the master weaver of the genome." This interaction occurs via the Mad homology 1 domain of Smad3. Although Smad2 and Smad4 fail to interact, an alternatively spliced form of Smad2 lacking exon 3 interacts with CTCF. CTCF does not perturb well established transforming growth factor beta gene responses. However, Smads and CTCF co-localize to the H19 imprinting control region (ICR), which emerges as an insulator in cis and regulator of transcription and replication in trans via direct CTCF binding to the ICR. Smad recruitment to the ICR requires intact CTCF binding to this locus. Smad2/3 binding to the ICR requires Smad4, which potentially provides stability to the complex. Because the CTCF-Smad complex is not essential for the chromatin insulator function of the H19 ICR, we propose that it can play a role in chromatin cross-talk organized by the H19 ICR.
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http://dx.doi.org/10.1074/jbc.M109.088385DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2888383PMC
June 2010

Regulating the stability of TGFbeta receptors and Smads.

Cell Res 2009 Jan;19(1):21-35

Ludwig Institute for Cancer Research, Uppsala University, Box 595, Biomedical Center, SE-751 24 Uppsala, Sweden.

Transforming growth factor beta (TGFbeta) controls cellular behavior in embryonic and adult tissues. TGFbeta binding to serine/threonine kinase receptors on the plasma membrane activates Smad molecules and additional signaling proteins that together regulate gene expression. In this review, mechanisms and models that aim at explaining the coordination between several components of the signaling network downstream of TGFbeta are presented. We discuss how the activity and duration of TGFbeta receptor/Smad signaling can be regulated by post-translational modifications that affect the stability of key proteins in the pathway. We highlight links between these mechanisms and human diseases, such as tissue fibrosis and cancer.
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http://dx.doi.org/10.1038/cr.2008.308DOI Listing
January 2009

Degradation of the tumor suppressor Smad4 by WW and HECT domain ubiquitin ligases.

J Biol Chem 2005 Jun 6;280(23):22115-23. Epub 2005 Apr 6.

Ludwig Institute for Cancer Research, Box 595, Biomedical Center, Uppsala University, SE-751 24 Uppsala, Sweden.

Smad4 mediates signaling by the transforming growth factor-beta (TGF-beta) superfamily of cytokines. Smad signaling is negatively regulated by inhibitory (I) Smads and ubiquitin-mediated processes. Known mechanisms of proteasomal degradation of Smads depend on the direct interaction of specific E3 ligases with Smads. Alternatively, I-Smads elicit degradation of the TGF-beta receptor by recruiting the WW and HECT domain E3 ligases, Smurfs, WWP1, or NEDD4-2. We describe an equivalent mechanism of degradation of Smad4 by the above E3 ligases, via formation of ternary complexes between Smad4 and Smurfs, mediated by R-Smads (Smad2) or I-Smads (Smad6/7), acting as adaptors. Smurfs, which otherwise cannot directly bind to Smad4, mediated poly-ubiquitination of Smad4 in the presence of Smad6 or Smad7. Smad4 co-localized with Smad7 and Smurf1 primarily in the cytoplasm and in peripheral cell protrusions. Smad2 or Smad7 mutants defective in Smad4 interaction failed to induce Smurf1-mediated down-regulation of Smad4. A Smad4 mutant defective in Smad2 or Smad7 interaction could not be effectively down-regulated by Smurf1. We propose that Smad4 is targeted for degradation by multiple ubiquitin ligases that can simultaneously act on R-Smads and signaling receptors. Such mechanisms of down-regulation of TGF-beta signaling may be critical for proper physiological response to this pathway.
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http://dx.doi.org/10.1074/jbc.M414027200DOI Listing
June 2005

Differential ubiquitination defines the functional status of the tumor suppressor Smad4.

J Biol Chem 2003 Aug 5;278(35):33571-82. Epub 2003 Jun 5.

Ludwig Institute for Cancer Research, Box 595, Biomedical Center, SE-751 24 Uppsala, Sweden.

Smad4 is an essential signal transducer of all transforming growth factor-beta (TGF-beta) superfamily pathways that regulate cell growth and differentiation, and it becomes inactivated in human cancers. Receptor-activated (R-) Smads can be poly-ubiquitinated in the cytoplasm or the nucleus, and this regulates their steady state levels or shutdown of the signaling pathway. Oncogenic mutations in Smad4 and other Smads have been linked to protein destabilization and proteasomal degradation. We analyzed a panel of missense mutants derived from human cancers that map in the N-terminal Mad homology (MH) 1 domain of Smad4 and result in protein instability. We demonstrate that all mutants exhibit enhanced poly-ubiquitination and proteasomal degradation. In contrast, wild type Smad4 is a relatively stable protein that undergoes mono- or oligo-ubiquitination, a modification not linked to protein degradation. Analysis of Smad4 deletion mutants indicated efficient mono- or oligo-ubiquitination of the C-terminal MH2 domain. Mass spectrometric analysis of mono-ubiquitinated Smad4 MH2 domain identified lysine 507 as a major target for ubiquitination. Lysine 507 resides in the conserved L3 loop of Smad4 and participates in R-Smad C-terminal phosphoserine recognition. Mono- or oligo-ubiquitinated Smad4 exhibited enhanced ability to oligomerize with R-Smads, whereas mutagenesis of lysine 507 led to inefficient Smad4/R-Smad hetero-oligomerization and defective transcriptional activity. Finally, overexpression of a mutant ubiquitin that only leads to mono-ubiquitination of Smad4 enhanced Smad transcriptional activity. These data suggest that oligo-ubiquitination positively regulates Smad4 function, whereas poly-ubiquitination primarily occurs in unstable cancer mutants and leads to protein degradation.
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http://dx.doi.org/10.1074/jbc.M300159200DOI Listing
August 2003