RESUMO
Anoikis is a form of anchorage-dependent apoptosis, and cancer cells adopt anokis-resistance molecular machinery to conduct metastasis. Here, we report that N-acetylglucosaminyltransferase V gene expression confers anoikis resistance during cancer progression. Overexpression of N-acetylglucosaminyltransferase V protected detached cancer cells from apoptotic death, and suppression or knockout of the gene sensitized cancer cells to the apoptotic death. The gene expression also stimulated anchorage-dependent as well as anchorage-independent colony formation of cancer cells following anoikis stress treatments. Importantly, treatment with the lectin from Sambucus sieboldiana significantly sensitized anoikis-induced cancer cell deaths in vitro as well as in vivo. We propose that the lectin alone or an engineered form could offer a new therapeutic treatment option for cancer patients with advanced tumors.
Assuntos
Anoikis/efeitos dos fármacos , Neoplasias do Colo/metabolismo , Neoplasias do Colo/patologia , N-Acetilglucosaminiltransferases/metabolismo , Lectinas de Plantas/farmacologia , Sambucus/química , Animais , Anoikis/genética , Apoptose/efeitos dos fármacos , Apoptose/genética , Linhagem Celular Tumoral , Proliferação de Células , Neoplasias do Colo/genética , Modelos Animais de Doenças , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Técnicas de Inativação de Genes , Humanos , Camundongos , N-Acetilglucosaminiltransferases/genética , Metástase Neoplásica , Transcriptoma , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
TALE-nuclease chimeras (TALENs) can bind to and cleave specific genomic loci and, are used to engineer gene knockouts and additions. Recently, instead of using the FokI domain, epigenetically active domains, such as TET1 and LSD1, have been combined with TAL effector domains to regulate targeted gene expression via DNA and histone demethylation. However, studies of histone methylation in the TALE system have not been performed. Therefore, in this study, we established a novel targeted regulation system with a TAL effector domain and a histone methylation domain. To construct a TALE-methylation fusion protein, we combined a TAL effector domain containing an E-Box region to act as a Snail binding site and the SET domain of EHMT 2 to allow for histone methylation. The constructed TALE-SET module (TSET) repressed the expression of E-cadherin via by increasing H3K9 dimethylation. Moreover, the cells that overexpressed TSET showed increased cell migration and invasion. This is the first phenotype-based study of targeted histone methylation by the TALE module, and this new system can be applied in new cancer therapies to reduce side effects.
Assuntos
Caderinas/metabolismo , Metilação de DNA/genética , Chaperonas de Histonas/genética , Proteínas de Homeodomínio/genética , Neoplasias/genética , Proteínas Repressoras/genética , Fatores de Transcrição/genética , Sítios de Ligação/genética , Linhagem Celular Tumoral , Movimento Celular/genética , Proteínas de Ligação a DNA/genética , Transição Epitelial-Mesenquimal/genética , Células HCT116 , Células HeLa , Histonas/metabolismo , Humanos , Invasividade Neoplásica/genética , Estrutura Terciária de Proteína , Proteínas Recombinantes de Fusão/genética , CicatrizaçãoRESUMO
N-Acetylglucosaminyltransferase V (GnT-V) is an enzyme that catalyzes the formation of a ß1,6-N-acetylglucosamine (GlcNAc) side chain to a core mannosyl residue in N-linked glycoproteins. Besides its direct function of producing aberrant glycoproteins, it promotes cancer progression by its involvement in the stimulation of oncoproteins. Herein, we report that GnT-V guided the transcriptional activation of membrane-type matrix metalloproteinase-1 (MT1-MMP) in cancer cells. The activated MT1-MMP expression had dual effects on cancer progression. It not only promoted proteolytic activity for cancer cells per se, but also led to the activation of MMP-2. Consequently, the activation of the two MMPs triggered by GnT-V intensified the invasive potential. A quantitative analysis using clinical tissues revealed a relatively strong correlation between GnT-V overexpression and MT1-MMP upregulation. In this study, we report for the first time that GnT-V directs cancer progression by modulating MMPs in cancer.
Assuntos
Metaloproteinase 14 da Matriz/genética , N-Acetilglucosaminiltransferases/metabolismo , Neoplasias/patologia , Ativação Transcricional , Regulação Enzimológica da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Humanos , Metaloproteinase 2 da Matriz/genética , N-Acetilglucosaminiltransferases/genética , Invasividade Neoplásica , Metástase Neoplásica , Neoplasias/enzimologia , Neoplasias/genética , Células Tumorais CultivadasRESUMO
There has been ongoing debate over whether tissue inhibitor of metalloproteinase-1 (TIMP-1) is pro- or anti-oncogenic. We confirmed that TIMP-1 reinforced cell proliferation in an αvß3 integrin-dependent manner and conferred resistance against cytotoxicity triggered by TNF-α and IL-2 in WiDr colon cancer cells. The cell-proliferative effects of TIMP-1 contributed to clonogenicity and tumor growth during the onset and early phase of tumor formation in vivo and in vitro. However, mass-produced TIMP-1 impeded further tumor growth by tightly inhibiting the activities of collagenases, which are critical for tumor growth and malignant transformation. Tumor cells could overcome this impasse by overexpression of N-acetylglucosaminyltransferase V, which deteriorates TIMP-1 into an aberrant glycoform. The aberrant glycoform of TIMP-1 was responsible for the mitigated inhibition of collagenases. The outbalanced activities of collagenases can degrade the basement membrane and the interstitial matrix, which act as a physical barrier for tumor growth and progression more efficiently. The concomitant overexpression of TIMP-1 and N-acetylglucosaminyltransferase V enabled WiDr cells to show a higher tumor growth rate as well as more malignant behaviors in a three-dimensional culture system.
Assuntos
Proliferação de Células , Neoplasias do Colo/metabolismo , Integrina alfaVbeta3/biossíntese , N-Acetilglucosaminiltransferases/biossíntese , Proteínas de Neoplasias/biossíntese , Inibidor Tecidual de Metaloproteinase-1/biossíntese , Linhagem Celular Tumoral , Neoplasias do Colo/genética , Neoplasias do Colo/patologia , Glicosilação , Humanos , Integrina alfaVbeta3/genética , N-Acetilglucosaminiltransferases/genética , Proteínas de Neoplasias/genética , Inibidor Tecidual de Metaloproteinase-1/genéticaRESUMO
Recent studies have demonstrated that the cell adhesion molecule, L1, is expressed in several malignant tumor types and its expression correlates with tumor progression and metastasis. However, the role of L1 in gallbladder carcinoma (GBC) remains unclear. Here, we demonstrate that L1 is expressed in GBC cells and plays an important role in the growth, motility, invasiveness, and adhesiveness of GBC cells. Specific depletion or overexpression of L1 in the GBC cell lines JCRB1033 and SNU-308, respectively, was achieved by lentivirus-mediated transduction and expression of an L1 mRNA-specific short hairpin RNA or full-length human L1. Stable depletion of L1 led to a significant decrease in GBC cell proliferation, migration and invasion, as well as decreased intracellular signaling through AKT and FAK. Overexpression of L1 in GBC cells enhanced these cellular activities. In a GBC xenograft nude mouse model, suppression of L1 markedly reduced tumor growth and increased the survival of tumor-bearing mice whereas L1 overexpression stimulated tumorigenicity. Taken together, these results suggest that L1 plays a crucial role in GBC progression and may be a novel therapeutic target in GBC treatment.
