ABSTRACT
Cln Three Requiring 9 (CTR9), a scaffold protein of the polymerase-associated factor-1 (PAF1) complex (PAF1c), is primarily localized in the nucleus of cells. Recent studies show that CTR9 plays essential roles in the development of various human cancers and their occurrence; however, its regulatory roles and precise mechanisms in hepatocellular carcinoma (HCC) remain unclear. In this study, we investigated the roles of CTR9 using in vitro assays and a xenograft mouse model. We found that CTR9 protein is upregulated in tumor tissues from HCC patients. Knockdown of CTR9 substantially reduced HCC cell proliferation, invasion, and migration, whereas its overexpression promoted these activities. In addition, in vitro results revealed that CTR9 silencing dramatically increased cell cycle regulators, p21 and p27, but markedly decreased matrix metalloproteinases, MMP2 and MMP9, with these outcomes reversed upon CTR9 overexpression. Furthermore, the underlying molecular mechanism suggests that CTR9 promoted the oncogene paternally expressed gene 10 (PEG10) transcription via its promoter region. Finally, the oncogenic roles of CTR9 were confirmed in a xenograft mouse model. This study confirms that CTR9, an oncoprotein that promotes HCC cell proliferation, invasion, and migration, increases tumor growth in a xenograft mouse model. CTR9 could be a novel therapeutic target. Further investigation is warranted to verify CTR9 potential in novel therapies for HCC.
Subject(s)
Carcinoma, Hepatocellular , Liver Neoplasms , Phosphoproteins , Transcription Factors , Animals , Apoptosis Regulatory Proteins/genetics , Apoptosis Regulatory Proteins/metabolism , Carcinogenesis/genetics , Carcinoma, Hepatocellular/pathology , Cell Line, Tumor , Cell Movement , Cell Proliferation/genetics , DNA-Binding Proteins/genetics , Gene Expression Regulation, Neoplastic , Humans , Liver Neoplasms/pathology , Mice , Phosphoproteins/metabolism , RNA-Binding Proteins/metabolism , Transcription Factors/metabolismABSTRACT
The fibroblast growth factors (FGFs) are important for embryo development, wound healing, hematopoiesis, and angiogenesis. FGF-1, a member of FGF family, is involved in both receptor-dependent pathways and an intracrine pathway. Studies have recently shown that FGF-1 is overexpressed in the early stages of several kinds of cancer. Thus, FGF-1 is a candidate for cancer immunotargeting. To study the potential use of therapeutic antibodies against FGF-1, a monoclonal hybridoma 1C9 secreting monoclonal antibody specific for FGF-1 was developed. Then, a single-chain variable fragment (scFv) antibody was genetically engineered from hybridama 1C9. The binding of the scFv1C9 to the antigen FGF-1 was demonstrated by ELISA and immunoprecipitation assays. Functional analysis showed that the overexpressed scFv1C9 in MCF-7 cells targeted endogenous FGF-1 and prevented the translocation of FGF-1 into the nucleus, resulting in the blockade of the intracrine pathway of FGF-1, which caused the G1 arrest by p21 up-regulation. These results suggest that the generated scFv1C9 is an effective inhibitor of the intracrine pathway of FGF-1 and has a potential application as anti-tumoral agent in breast cancer.
Subject(s)
Antineoplastic Agents/pharmacology , Biomarkers, Tumor/metabolism , Breast Neoplasms/immunology , Breast Neoplasms/metabolism , Fibroblast Growth Factor 1/metabolism , Signal Transduction/drug effects , Single-Chain Antibodies/pharmacology , Animals , Antineoplastic Agents/immunology , Antineoplastic Agents/therapeutic use , Base Sequence , Biomarkers, Tumor/genetics , Breast Neoplasms/drug therapy , Breast Neoplasms/genetics , Breast Neoplasms/pathology , Cell Cycle/drug effects , Cell Line, Tumor , Cell Proliferation/drug effects , Cyclin-Dependent Kinase Inhibitor p21/genetics , Cyclin-Dependent Kinase Inhibitor p21/metabolism , Escherichia coli , Female , Fibroblast Growth Factor 1/genetics , Gene Expression , Humans , Hybridomas/chemistry , Hybridomas/metabolism , Immunomodulation , Mice , Molecular Sequence Data , Molecular Targeted Therapy/methods , Recombinant Proteins/genetics , Recombinant Proteins/metabolism , Single-Chain Antibodies/immunology , Single-Chain Antibodies/therapeutic use , Up-RegulationABSTRACT
Myosin X (Myo X), an unconventional myosin with a tail homology 4-band 4.1/ezrin/radixin/moesin (MyTH4-FERM) tail, is expressed ubiquitously in various mammalian tissues. In addition to the full-length Myo X (Myo X FL), a headless form is synthesized in the brain. So far, little is known about the function of this motor-less Myo X. In this study, the role of the headless Myo X was investigated in immortalized gonadotropin-releasing hormone (GnRH) neuronal cells, NLT. NLT cells overexpressing the headless Myo X formed fewer focal adhesions and spread more slowly than the wild-type NLT cells and GFP-expressing NLT cells. In chemomigration assays, the NLT cells overexpressing the headless Myo X migrated shorter distances and had fewer migratory cells compared with the control NLT cells.
Subject(s)
Cell Movement/physiology , Gonadotropin-Releasing Hormone/metabolism , Myosins/metabolism , Animals , Cell Adhesion/physiology , Cell Line , Mice , Myosins/genetics , Neurons/cytology , Neurons/metabolism , Recombinant Fusion Proteins/genetics , Recombinant Fusion Proteins/metabolismABSTRACT
The chromosome samples of Rana nigromaculata were prepared by peripheral blood lymphocyte cultured in vitro. Karyotype, C-banding and silver-staining nucleolus organizer regions (Ag-NORs) were observed on chromosomes samples. The result showed that: (1) the chromosome number of Rana nigromaculata is 26 (2n=26), i.e., 10 large and 16 small chromosomes, the large chromosomes are matched into 5 homologous pairs and small chromosomes are matched into 8 homologous pairs, and further indicates the karyotype of Rana nigromaculata is bi-modal; (2) Metaphases of female and male were observed separately, the chromosome of No.11 has a pronounced secondary constriction in the middle of long arms. But the position of secondary constriction in the aberrant type individuals is in the middle of long arms of the No.8; (3) it has only one obvious C-banding close to the telomere, which is located in the long arms of the No.5; (4) The chromosome No.11 is a pair of homologous chromosomes with Ag-NORs, and the position of Ag-NORs in female and male is identical.
