1.A putative pH-dependent nuclear localization signal in the juxtamembrane region of c-Met.
Shubhash Chandra CHAUDHARY ; Min Guk CHO ; Tuyet Thi NGUYEN ; Kyu Sang PARK ; Myung Hee KWON ; Jae Ho LEE
Experimental & Molecular Medicine 2014;46(10):e119-
The C-terminal fragment of the c-Met receptor tyrosine kinase is present in the nuclei of some cells irrespective of ligand stimulation, but the responsible nuclear localization signal (NLS) has not been previously reported. Here, we report that two histidine residues separated by a 10-amino-acid spacer (H1068-H1079) located in the juxtamembrane region of c-Met function as a putative novel NLS. Deletion of these sequences significantly abolished the nuclear translocation of c-Met, as did substitution of the histidines with alanines. This substitution also decreased the association of c-Met fragment with importin beta. The putative NLS of c-Met is unique in that it relies on histidines, whose positive charge changes depending on pH, rather than the lysines or arginines, commonly found in classical bipartite NLSs, suggesting the possible 'pH-dependency' of this NLS. Indeed, decreasing the cytosolic pH either with nigericin, an Na+/H+ exchanger or pH 6.5 KRB buffer significantly increased the level of nuclear c-Met and the interaction of the c-Met fragment with importin beta, indicating that low pH itself enhanced nuclear translocation. Consistent with this, nigericin treatment also increased the nuclear level of endogenous c-Met in HeLa cells. The putative aberrant bipartite NLS of c-Met seems to be the first example of what we call a 'pH-dependent' NLS.
Active Transport, Cell Nucleus
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Amino Acid Sequence
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HeLa Cells
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Humans
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Hydrogen-Ion Concentration
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Molecular Sequence Data
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*Nuclear Localization Signals
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Protein Structure, Tertiary
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Proto-Oncogene Proteins c-met/*analysis/genetics/*metabolism
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Sequence Deletion
2.cDNA microarray-based study of gene expression profile changes in human esophageal squamous cell carcinoma.
Pei LI ; Zhi-qiang LING ; Hong-yan YANG ; You-tian HUANG ; Ji-min ZHAO ; Ming-yao ZHAO ; Zi-ming DONG
Journal of Southern Medical University 2006;26(5):632-634
OBJECTIVETo investigate the differentially expressed genes between human esophageal squamous cell carcinoma (ESCC) and normal esophageal mucosa and explore an effective method with high throughput for screening the molecular markers closely correlated with the development, invasion and metastasis of ESCC.
METHODSWith cDNA microarray and laser capture microdissection, T7-based amplification were used to detect the mRNA from both the primary carcinoma and the corresponding esophageal epithelium in 15 ESCC cases, and the results were analyzed by bioinformatics methods.
RESULTSAmong the 886 target genes, 110 (12.42%) genes were differentially expressed commonly at least twice in all the 15 samples, including 56 (6.32%) up-regulated by at least 2 folds and 54 (6.09%) down-regulated by at least 0.5 folds.
CONCLUSIONMany ESCC-associated genes were screened by the high-throughput gene chip method, and functional study of these genes may help to identify the key genes or pathways involved in the pathogenesis and development of ESCC.
Carcinoma, Squamous Cell ; genetics ; pathology ; Epithelium ; metabolism ; Esophageal Neoplasms ; genetics ; pathology ; Esophagus ; metabolism ; Female ; Gene Expression Profiling ; Gene Expression Regulation, Neoplastic ; Humans ; Male ; Middle Aged ; Oligonucleotide Array Sequence Analysis ; methods ; Proto-Oncogene Proteins ; genetics ; Proto-Oncogene Proteins c-met ; Receptors, Growth Factor ; genetics
3.Transforming variant of Met receptor confers serum independence and anti-apoptotic property and could be involved in the mouse thymic lymphomagenesis.
Cheol Min BAEK ; Soung Hoo JEON ; Ja June JANG ; Bok Soon LEE ; Jae Ho LEE
Experimental & Molecular Medicine 2004;36(4):283-291
Met tyrosine kinase receptor, the receptor of hepatocyte growth factor/scatter factor (HGF/SF), is present in mouse tissues as two major isoforms differing by a 47-aminoacid segment in the juxtamembrane domain via alternative splicing of exon 14. We found that the smaller isoform of Met (Sm-Met) was highly transformable in both in vitro and in vivo tumorigenesis assays. In this report, close examination of the transforming activity of the Sm-Met showed that the expression of Sm-Met conferred the cells serum independence and anti- apoptotic property when treated with doxorubicin. These properties of Sm-Met seemed to be originated from its far longer maintenance of tyrosine kinase activity after the binding of HGF/SF. Interestingly, the longer maintenance of activated status was accompanied with more increase of tyrosine phosphorylation of Stat3 protein. Moreover, we have tried to find (an) animal tumorigenesis model(s) showing the increase in the expression of this transforming variant of Met. In gamma-ray-induced mouse thymic lymphoma model, the expression of the mRNAs for Sm-Met was significantly increased as well as those of wild type Met and HGF/SF, suggesting a possible role of the Sm-Met in tumorigenesis in vivo.
Animals
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*Apoptosis
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Cell Proliferation
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Cell Survival
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*Cell Transformation, Neoplastic
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DNA-Binding Proteins/metabolism
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Doxorubicin/pharmacology
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Hepatocyte Growth Factor/pharmacology
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Lymphoma/*etiology/genetics/metabolism
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Mice
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NIH 3T3 Cells
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Phosphorylation
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Protein Isoforms/genetics/metabolism
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Proto-Oncogene Protein c-met/genetics/*metabolism
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RNA, Messenger/analysis/metabolism
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Research Support, Non-U.S. Gov't
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Serum/metabolism
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Thymus Gland
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Trans-Activators/metabolism
4.Inducing effects of hepatocyte growth factor on the expression of vascular endothelial growth factor in human colorectal carcinoma cells through MEK and PI3K signaling pathways.
Yu-hua ZHANG ; Wei WEI ; Hao XU ; Yan-yan WANG ; Wen-xi WU
Chinese Medical Journal 2007;120(9):743-748
BACKGROUNDVascular endothelial growth factor plays a key role in human colorectal carcinoma invasion and metastasis. However, the regulation mechanism remains unknown. Recent studies have shown that several cytokines can regulate the expression of vascular endothelial growth factor in tumor cells. In this study, we investigated whether hepatocyte growth factor can regulate the expression of vascular endothelial growth factor in colorectal carcinoma cells.
METHODSHepatocyte growth factor and vascular endothelial growth factor in human serum were measured by ELISA. The mRNA level of vascular endothelial growth factor was analyzed by reverse transcription-PCR. Western blot assay was performed to evaluate levels of c-Met and several other proteins involved in the MAPK and PI3K signaling pathways in colorectal carcinoma cells.
RESULTSSerum hepatocyte growth factor and vascular endothelial growth factor were significantly increased in colorectal carcinoma subjects. In vitro extraneous hepatocyte growth factor markedly increased protein and mRNA levels of vascular endothelial growth factor in colorectal carcinoma cells. Hepatocyte growth factor induced phosphorylation of c-Met, ERK1/2 and AKT in a dose-dependent manner. Specific inhibitors on MEK and PI3K inhibited the hepatocyte growth factor-induced expression of vascular endothelial growth factor in colorectal carcinoma cells.
CONCLUSIONThis present study indicates that hepatocyte growth factor upregulates the expression of vascular endothelial growth factor in colorectal carcinoma cells via the MEK/ERK and PI3K/AKT signaling pathways.
Butadienes ; pharmacology ; Cell Line, Tumor ; Chromones ; pharmacology ; Colorectal Neoplasms ; metabolism ; pathology ; Gene Expression Regulation ; drug effects ; Hepatocyte Growth Factor ; blood ; pharmacology ; Humans ; MAP Kinase Signaling System ; physiology ; Morpholines ; pharmacology ; Nitriles ; pharmacology ; Phosphatidylinositol 3-Kinases ; physiology ; Phosphorylation ; Proto-Oncogene Proteins c-met ; metabolism ; RNA, Messenger ; analysis ; Signal Transduction ; physiology ; Vascular Endothelial Growth Factor A ; genetics