Akt isoforms regulate intermediate filament protein levels in epithelial carcinoma cells

Department of Chemistry-Biology, University of Quebec at Trois-Rivieres, Trois-Rivieres, Quebec, Canada G9A 5H7.
FEBS letters (Impact Factor: 3.17). 03/2010; 584(5):984-8. DOI: 10.1016/j.febslet.2010.01.045
Source: PubMed


Keratin 8 and 18 are simple epithelial intermediate filament (IF) proteins, whose expression is differentiation- and tissue-specific, and is maintained during tumorigenesis. Vimentin IF is often co-expressed with keratins in cancer cells. Recently, IF have been proposed to be involved in signaling pathways regulating cell growth, death and motility. The PI3K/Akt pathway plays a pivotal role in these processes. Thus, we investigated the role of Akt (1 and 2) in regulating IF expression in different epithelial cancer cell lines. Over-expression of Akt1 increases K8/18 proteins. Akt2 up-regulates K18 and vimentin expression by an increased mRNA stability. To our knowledge, these results represent the first indication that Akt isoforms regulate IF expression and support the hypothesis that IFs are involved in PI3K/Akt pathway.

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Available from: Anne-Marie Fortier, Mar 12, 2014
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    • "Moreover, K8/18 complete filamentous network seems to be essential to exert the moderating role in cell motility since perinuclear reorganization of K8/18 network by sphingosylphosphorylcholine increases cellular elasticity and augments migration through limited-sized pores [142]. We also demonstrated that perinuclear reorganization of K8/18 network occurs in HeLa and HepG2 epithelial cancer cells overexpressing constitutively active Akt isoforms [143], and that is associated with increased K8/18 protein levels [143] and invasion through matrigel (Fortier et al., unpublished results). Disruption of K8/18 filamentous network in hepatocytes by transfection of mutant K18 or by proteasome inhibition also affects localization of desmoplakin, zonula occludens-1, beta-catenin and 14-3-3-zeta, which are relocated to keratin inclusions [144]. "
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    ABSTRACT: K8 and K18 (K8/18) are the major heteropolymeric intermediate filaments (IFs) present in simple layer epithelia. In hepatocytes, keratin filaments form an extensive cytoplasmic network that is denser at the cell periphery and around bile canaliculi. Keratin filaments are attached to the plasma membrane via desmosomes. K8/18 IFs have long been linked to human chronic liver diseases. In fact, modifications in keratin IFs network organization and the formation, in hepatocytes, of K8/18 containing aggregates, named Mallory-Denk bodies (MDBs), are characteristic of alcoholic and non-alcoholic steatohepatitis, copper metabolism diseases such as Wilson disease and Indian childhood cirrhosis and hepatocellular carcinoma. The formation of MDBs is the consequence of an increase of K8/18 mRNA and proteins, alterations in K8/18 post-translational modifications such as phosphorylation on multiple sites, transglutaminase mediated keratin crosslinking and a defect in K8/18 degradation by ubiquitin-proteasome pathway. The use of transgenic mouse models has allowed unravelling the significance of these changes in K8/18 dynamic in hepatocytes and revealed a function for keratins in protecting hepatocytes against mechanical and non-mechanical stresses. For instance, mice expressing an ectopic human K14, a mutated human K18 (Arg89→Cys) or K18 (Ser52→Ala) or K18-Gly(-), K8 (Ser73→Ala) or K8 deficient mice are more susceptible to various mechanical and toxic injuries. The recent observation of the existence of K8 and K18 mutations in cases of cryptogenic and non-cryptogenic forms or human liver disease is in total agreement with a role for keratin in maintaining cellular integrity under various threatening conditions. In a pursuit for understanding the molecular mechanism by which keratins could accomplish their protective role in cells, researchers have investigated the relationship between K8/18 and different regulatory pathways. There is now evidence that keratins are involved in signalling pathways regulating apoptosis, cell growth, and motility of various simple epithelial cells. It is noteworthy that expression of keratins is maintained during malignant transformation of simple epithelial cells. The PI3K/Akt pathway plays a pivotal role in apoptosis, cell growth, and motility and over-expression of the active form of Akt1 and Akt2 increase K8/18 protein levels suggesting that IFs are involved in PI3K/Akt pathway. The direct binding of K8 to Akt1 suggests that K8/18 IFs might provide a scaffold for Akt1 in cells. K8/18 interacts directly with other molecules involved in the apoptotic signalling pathway such as tumor necrosis factor receptor (TNFR), TNFR1-associated death domain protein (TRADD), Mrj co-chaperon, with Hsp/c70 and caspase 3. K8/18 also binds to key regulatory proteins of mitosis and cell proliferation such as 14-3-3, Cdc25 and Raf-1 kinase. Therefore keratins which were considered as only structural in the early 90s are now considered as key regulatory elements in modulating multiple signalling pathways.
    Keratin: Structure, Properties and Applications, 06/2012: chapter 1: pages 1-35; Nova Science Publishers, Inc., ISBN: 978-1-62100-336-6
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    • "Additional evidence that keratins may function upstream of mTOR is provided by studies in mice with ablation of all keratin genes, where embryonic lethality from severe growth retardation is associated with aberrant localization of the glucose transporters GLUT1 and GLUT3m resulting in adenosine monophosphate kinase activation and suppression of the mTORC1 downstream targets S6 kinase and 4E-BP1 (Vijayaraj et al., 2009). In an apparently reciprocal relationship, AKT isoforms regulate intermediate filament expression in epithelial cancer cell lines, as overexpression of AKT1 increases K8/K18 levels and AKT2 upregulates K18 and vimentin (Fortier et al., 2010). Thus, keratins, which are often aberrantly expressed in epithelial cancers, interact in multiple ways with the AKT/mTOR pathway, which itself is frequently abnormally activated in aggressive tumors, raising the possibility that the role of AKT in epithelial tumorigenesis is at least partially keratin mediated and/or dependent. "
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    ABSTRACT: Keratins are the intermediate filament (IF)-forming proteins of epithelial cells. Since their initial characterization almost 30 years ago, the total number of mammalian keratins has increased to 54, including 28 type I and 26 type II keratins. Keratins are obligate heteropolymers and, similarly to other IFs, they contain a dimeric central α-helical rod domain that is flanked by non-helical head and tail domains. The 10-nm keratin filaments participate in the formation of a proteinaceous structural framework within the cellular cytoplasm and, as such, serve an important role in epithelial cell protection from mechanical and non-mechanical stressors, a property extensively substantiated by the discovery of human keratin mutations predisposing to tissue-specific injury and by studies in keratin knockout and transgenic mice. More recently, keratins have also been recognized as regulators of other cellular properties and functions, including apico-basal polarization, motility, cell size, protein synthesis and membrane traffic and signaling. In cancer, keratins are extensively used as diagnostic tumor markers, as epithelial malignancies largely maintain the specific keratin patterns associated with their respective cells of origin, and, in many occasions, full-length or cleaved keratin expression (or lack there of) in tumors and/or peripheral blood carries prognostic significance for cancer patients. Quite intriguingly, several studies have provided evidence for active keratin involvement in cancer cell invasion and metastasis, as well as in treatment responsiveness, and have set the foundation for further exploration of the role of keratins as multifunctional regulators of epithelial tumorigenesis.
    Oncogene 10/2010; 30(2):127-38. DOI:10.1038/onc.2010.456 · 8.46 Impact Factor
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    ABSTRACT: Keratins 8 and 18 (K8/18) intermediate filament proteins are believed to play an essential role in the protection of hepatocytes against mechanical and toxic stress. This assertion is mainly based on increased hepatocyte fragility observed in transgenic mice deficient in K8/18, or carrying mutations on K8/18. The molecular mechanism by which keratins accomplish their protective functions has not been totally elucidated. Liver diseases such as alcoholic hepatitis and copper metabolism diseases are associated with modifications, in hepatocytes, of intermediate filament organisation and the formation of K8/18 containing aggregates named Mallory-Denk bodies. Treatment of mice with a diet containing griseofulvin induces the formation of Mallory-Denk bodies in hepatocytes. This provides a reliable animal model for assessing the molecular mechanism by which keratins accomplish their protective role in the response of hepatocytes to chemical injuries. In this study, we found that griseofulvin intoxication induced changes in keratin solubility and that there was a 5% to 25% increase in the relative amounts of soluble keratin. Keratin phosphorylation on specific sites (K8 pS79, K8 pS436 and K18 pS33) was increased and prominent in the insoluble protein fractions. Since at least six K8 phosphoepitopes were detected after GF treatment, phosphorylation sites other than the ones studied need to be accounted for. Immunofluorescence staining showed that K8 pS79 epitope was present in clusters of hepatocytes that surrounded apoptotic cells. Activated p38 MAPK was associated with, but not present in K8 pS79-positive cells. These results indicate that griseofulvin intoxication mediates changes in the physicochemical properties of keratin, which result in the remodelling of keratin intermediate filaments which in turn could modulate the signalling pathways in which they are involved by modifying their binding to signalling proteins.
    Experimental and Molecular Pathology 10/2010; 89(2):117-25. DOI:10.1016/j.yexmp.2010.07.004 · 2.71 Impact Factor
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