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  • Küçük Resim
    Öğe
    Covalent dimerization of interleukin-like epithelial-to-mesenchymal transition (EMT) inducer (ILEI) facilitates EMT, invasion, and late aspects of metastasis
    (2017) Kral, Maria; Klimek, Christoph; Kutay, Betül; Timelthaler, Gerald; Lendl, Thomas; Neuditschko, Benjamin; Gerner, Christopher; Sibilia, Maria; Csiszar, Agnes
    The interleukin-like epithelial-to-mesenchymal transition (EMT) inducer(ILEI)/FAM3C is a member of the highly homologous FAM3 family andis essential for EMT and metastasis formation. It is upregulated in severalcancers, and its altered subcellular localization strongly correlates withpoor survival. However, the mechanism of ILEI action, including the struc-tural requirements for ILEI activity, remains elusive. Here, we show thatILEI formed both monomers and covalent dimers in cancer cell lines andin tumors. Using mutational analysis and pulse-chase experiments, wefound that the four ILEI cysteines, conserved throughout the FAM3 fam-ily and involved in disulfide bond formation were essential for extracellularILEI accumulation in cultured cells. Modification of a fifth cysteine(C185), unique for ILEI, did not alter protein secretion, but completelyinhibited ILEI dimerization. Wild-type ILEI monomers, but not C185Amutants, could be converted into covalent dimers extracellularly uponoverexpression by intramolecular-to-intermolecular disulfide bond isomer-ization. Incubation of purified ILEI with cell culture medium showed thatdimerization was triggered by bovine serum in a dose- and time-dependentmanner. Purified ILEI dimers induced EMT and trans-well invasion ofcancer cellsin vitro. In contrast, ILEI monomers and the dimerization-defective C185A mutant affected only cell motility as detected by scratchassays and cell tracking via time-lapse microscopy. Importantly, tumorcells overexpressing wild-type ILEI caused large tumors and lung metas-tases in nude mice, while cells overexpressing the dimerization-defectiveC185A mutant behaved similar to control cells. These data show that cova-lent ILEI self-assembly is essential for EMT induction, elevated tumorgrowth, and metastasis.
  • Küçük Resim
    Öğe
    Inducible overexpression of a FAM3C/ILEI transgene has pleiotropic effects with shortened life span, liver fibrosis and anemia in mice
    (2023) Uluca, Betül; Schmidt, Ulrike; Vokic, Iva; Malik, Barizah; KolbeI, Thomas; Lassnig, Caroline; Holcmann, Martin; Moreno-Viedma, Veronica; Robl, Bernhard; Muhlberge, Carina; Gotthard, Dagmar; Sibilia, Maria; Müller, Mathias; Csisza, Agnes; Rülick, Thomas; Klymkowsky, Michael
    FAM3C/ILEI is an important factor in epithelial-to-mesenchymal transition (EMT) induction, tumor progression and metastasis. Overexpressed in many cancers, elevated ILEI levels and secretion correlate with poor patient survival. Although ILEI’s causative role in invasive tumor growth and metastasis has been demonstrated in several cellular tumor models, there are no available transgenic mice to study these effects in the context of a complex organism. Here, we describe the generation and initial characterization of a Tet-ON inducible Fam3c/ILEI transgenic mouse strain. We find that ubiquitous induction of ILEI overexpression (R26-ILEIind) at weaning age leads to a shortened lifespan, reduced body weight and microcytic hypochromic anemia. The anemia was reversible at a young age within a week upon withdrawal of ILEI induction. Vav1-driven overexpression of the ILEIind transgene in all hematopoietic cells (Vav-ILEIind) did not render mice anemic or lower overall fitness, demonstrating that no intrinsic mechanisms of erythroid development were dysregulated by ILEI and that hematopoietic ILEI hyperfunction did not contribute to death. Reduced serum iron levels of R26-ILEIind mice were indicative for a malfunction in iron uptake or homeostasis. Accordingly, the liver, the main organ of iron metabolism, was severely affected in moribund ILEI overexpressing mice: increased alanine transaminase and aspartate aminotransferase levels indicated liver dysfunction, the liver was reduced in size, showed increased apoptosis, reduced cellular iron content, and had a fibrotic phenotype. These data indicate that high ILEI expression in the liver might reduce hepatoprotection and induce liver fibrosis, which leads to liver dysfunction, disturbed iron metabolism and eventually to death. Overall, we show here that the novel Tet-ON inducible Fam3c/ILEI transgenic mouse strain allows tissue specific timely controlled overexpression of ILEI and thus, will serve as a versatile tool to model the effect of elevated ILEI expression in diverse tissue entities and disease conditions, including cancer.