ABSTRACT
Approximately 40% of patients with lung adenocarcinoma (LUAD) often develop bone metastases during the course of their disease. However, scarcely any in vivo model of LUAD bone metastasis has been established, leading to a poor understanding of the mechanisms underlying LUAD bone metastasis. Here, we established a multiorgan metastasis model via the left ventricular injection of luciferase-labeled LUAD cells into nude mice and then screened out lung metastasis (LuM) and bone metastasis (BoM) cell subpopulations. BoM cells exhibited greater stemness and epithelial-mesenchymal transition (EMT) plasticity than LuM cells and initially colonized the bone and subsequently disseminated to distant organs after being reinjected into mice. Moreover, a CD74-ROS1 fusion mutation (C6; R34) was detected in BoM cells but not in LuM cells. Mechanistically, BoM cells bearing the CD74-ROS1 fusion highly secrete the C-C motif chemokine ligand 5 (CCL5) protein by activating STAT3 signaling, recruiting macrophages in tumor microenvironment and strongly inducing M2 polarization of macrophages. BoM cell-activated macrophages produce a high level of TGF-ß1, thereby facilitating EMT and invasion of LUAD cells via TGF-ß/SMAD2/3 signaling. Targeting the CD74-ROS1/CCL5 axis with Crizotinib (a ROS1 inhibitor) and Maraviroc (a CCL5 receptor inhibitor) in vivo strongly impeded bone metastasis and secondary metastasis of BoM cells. Our findings reveal the critical role of the CD74-ROS1/STAT3/CCL5 axis in the interaction between LUAD bone metastasis cells and macrophages for controlling LUAD cell dissemination, highlighting the significance of the bone microenvironment in LUAD bone metastasis and multiorgan secondary metastasis, and suggesting that targeting CD74-ROS1 and CCL5 is a promising therapeutic strategy for LUAD bone metastasis.
Subject(s)
Adenocarcinoma of Lung , Bone Neoplasms , Epithelial-Mesenchymal Transition , Lung Neoplasms , Macrophages , Protein-Tyrosine Kinases , Proto-Oncogene Proteins , Animals , Humans , Mice , Bone Neoplasms/secondary , Bone Neoplasms/genetics , Bone Neoplasms/metabolism , Bone Neoplasms/pathology , Macrophages/metabolism , Macrophages/pathology , Lung Neoplasms/secondary , Lung Neoplasms/pathology , Lung Neoplasms/genetics , Lung Neoplasms/metabolism , Adenocarcinoma of Lung/genetics , Adenocarcinoma of Lung/pathology , Adenocarcinoma of Lung/secondary , Adenocarcinoma of Lung/metabolism , Epithelial-Mesenchymal Transition/genetics , Protein-Tyrosine Kinases/metabolism , Protein-Tyrosine Kinases/genetics , Proto-Oncogene Proteins/genetics , Proto-Oncogene Proteins/metabolism , Antigens, Differentiation, B-Lymphocyte/metabolism , Antigens, Differentiation, B-Lymphocyte/genetics , Chemokine CCL5/metabolism , Chemokine CCL5/genetics , Histocompatibility Antigens Class II/genetics , Histocompatibility Antigens Class II/metabolism , Mice, Nude , Cell Line, Tumor , Tumor Microenvironment , Oncogene Proteins, Fusion/genetics , Oncogene Proteins, Fusion/metabolism , STAT3 Transcription Factor/metabolism , STAT3 Transcription Factor/genetics , Signal TransductionABSTRACT
Mammalian target of rapamycin (mTOR) kinase functions as a central regulator of cell growth and metabolism, and its complexes mTORC1 and mTORC2 phosphorylate distinct substrates. Dysregulation of mTOR signaling is commonly implicated in human diseases, including cancer. Despite three decades of active research in mTOR, much remains to be determined. Here, we demonstrate that prolyl 4-hydroxylase alpha-2 (P4HA2) binds directly to mTOR and hydroxylates one highly conserved proline 2341 (P2341) within a kinase domain of mTOR, thereby activating mTOR kinase and downstream effector proteins (e.g. S6K and AKT). Moreover, the hydroxylation of P2341 strengthens mTOR stability and allows mTOR to accurately recognize its substrates such as S6K and AKT. The growth of lung adenocarcinoma cells overexpressing mTORP2341A is significantly reduced when compared with that of cells overexpressing mTORWT. Interestingly, in vivo cell growth assays show that targeting P4HA2-mTOR significantly suppresses lung adenocarcinoma cell growth. In summary, our study reveals an undiscovered hydroxylation-regulatory mechanism by which P4HA2 directly activates mTOR kinase, providing insights for therapeutically targeting mTOR kinase-driven cancers.
Subject(s)
Adenocarcinoma of Lung , Cell Proliferation , Lung Neoplasms , TOR Serine-Threonine Kinases , Humans , Hydroxylation , TOR Serine-Threonine Kinases/metabolism , Adenocarcinoma of Lung/metabolism , Adenocarcinoma of Lung/pathology , Adenocarcinoma of Lung/genetics , Adenocarcinoma of Lung/drug therapy , Lung Neoplasms/metabolism , Lung Neoplasms/pathology , Lung Neoplasms/genetics , Lung Neoplasms/drug therapy , Animals , Mice , Cell Line, Tumor , Signal Transduction , Procollagen-Proline Dioxygenase/metabolism , Procollagen-Proline Dioxygenase/geneticsABSTRACT
The molecular underpinnings of lung adenocarcinoma (LUAD) metastasis remain poorly defined. Here, using human LUAD cell lines, we find that transcriptional intermediary factor 1 γ (TIF1γ) binds to TATA box binding protein (TBP) in competition with TBP-associated factor 15 (TAF15) and impedes TAF15/TBP-mediated interleukin 6 (IL-6) transactivation. TIF1γ modifies TAF15 through multi-mono-ubiquitylation and drives nuclear export of TAF15. Functionally, TAF15 accelerates epithelial-mesenchymal transition (EMT) and metastasis of LUAD cells, acting in just the opposite way as TIF1γ. Low TIF1γ or high TAF15 expression levels are shown in metastatic LUAD specimens and correlate with poor survival of individuals with LUAD. Our findings suggest that the TAF15/TBP complex is required for IL-6 activation-induced EMT and invasion, which are inhibited by TIF1γ. This study highlights the crucial interaction between TIF1γ and the TAF15/TBP complex for regulating EMT and metastasis in LUAD.
Subject(s)
Adenocarcinoma of Lung , Lung Neoplasms , TATA-Binding Protein Associated Factors , Humans , Epithelial-Mesenchymal Transition , Interleukin-6 , Lung Neoplasms/pathology , TATA-Binding Protein Associated Factors/genetics , TATA-Box Binding ProteinABSTRACT
Quaking (QKI) proteins belong to the signal transduction and activation of RNA (STAR) family of RNA-binding proteins that have multiple functions in RNA biology. Here, we show that QKI-5 is dramatically decreased in metastatic lung adenocarcinoma (LUAD). QKI-5 overexpression inhibits TGF-ß-induced epithelial-mesenchymal transition (EMT) and invasion, whereas QKI-5 knockdown has the opposite effect. QKI-5 overexpression and silencing suppresses and promotes TGF-ß-stimulated metastasis in vivo, respectively. QKI-5 inhibits TGF-ß-induced EMT and invasion in a TGFßR1-dependent manner. KLF6 knockdown increases TGFßR1 expression and promotes TGF-ß-induced EMT, which is partly abrogated by QKI-5 overexpression. Mechanistically, QKI-5 directly interacts with the TGFßR1 3' UTR and causes post-transcriptional degradation of TGFßR1 mRNA, thereby inhibiting TGF-ß-induced SMAD3 phosphorylation and TGF-ß/SMAD signaling. QKI-5 is positively regulated by KLF6 at the transcriptional level. In LUAD tissues, KLF6 is lowly expressed and positively correlated with QKI-5 expression, while TGFßR1 expression is up-regulated and inversely correlated with QKI-5 expression. We reveal a novel mechanism by which KLF6 transcriptionally regulates QKI-5 and suggest that targeting the KLF6/QKI-5/TGFßR1 axis is a promising targeting strategy for metastatic LUAD.
Subject(s)
Adenocarcinoma of Lung , Lung Neoplasms , Adenocarcinoma of Lung/genetics , Cell Line, Tumor , Cell Movement/genetics , Epithelial-Mesenchymal Transition/genetics , Gene Expression Regulation, Neoplastic , Humans , Lung Neoplasms/genetics , RNA-Binding Proteins , Transforming Growth Factor beta/genetics , Transforming Growth Factor beta/metabolism , Transforming Growth Factor beta1/metabolismABSTRACT
Polymorphic and morphological transformations of cellulosic materials are strongly associated to their properties and applications, especially in the case of emerging nanocelluloses. Related changes that take place upon treatment of cellulose nanocrystals (CNC) in alkaline conditions are studied here by XRD, TEM, AFM, and other techniques. The results indicate polymorphic transformation of CNC proceeds gradually in a certain range of alkali concentrations, i.e. from about 8% to 12.5% NaOH. In such transition alkali concentration, cellulose I and II allomorphs coexists. Such value and range of the transition concentration is strongly interdependent with the crystallite size of CNCs. In addition, it is distinctively lower than that for macroscopic fibers (12-15% NaOH). Transmission electron microscopy and particle sizing reveals that after mercerization CNCs tend to associate. Furthermore, TEMPO-oxidized mercerized CNC reveals the morphology of individual nanocrystal of the cellulose II type, which is composed of some interconnected granular structures. Overall, this work reveals how the polymorphism and morphology of individual CNC change in alkali conditions and sheds light onto the polymorphic transition from cellulose I to II.
