Basic–Alimentary TractIsolation and sequencing of a novel tropomyosin isoform preferentially associated with colon cancer☆,☆☆,★
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Isolation and characterization of cDNA clones
A λZAPII phage library prepared from T84 human colon cancer cell poly(A)+ RNAs was purchased from Stratagene (La Jolla, CA). A 29-mer oligonucleotide probe common to all of known hTM messenger RNAs was used to screen this library as described previously.4 After plaque purification, 28 positive clones were obtained. Plasmids from positive plaques were isolated from their Uni-ZAP XR vector by in vivo excision with helper phage as described by the manufacturer. The cDNA inserts were polymerase
Identification of cDNA clones encoding tropomyosin isoforms from T84 colon cancer cells
Previously, we designed a common oligonucleotide probe (REN29) and showed that this REN29 probe was capable of recognizing messenger RNAs derived from all 4 hTM genes.4 Using this probe to extensively screen a cDNA library prepared from T84 cancer cells, 28 positive clones were obtained. After restriction enzyme analysis, PCR amplification with isoform-specific primers (Table 1), and nucleotide sequencing of the inserts, 15, 11, and 1 clones were classified into hTM5, hTM4, and hTM1,
Discussion
In the present study, we cloned a novel tropomyosin TC22 isoform from a colon cancer cell line T84. This isoform has 5'-untranslated sequence and most of the coding sequences (except the last coding exon, exon 9) identical to that in hTM5, suggesting that both isoforms are derived from the same human γ-TM gene. Furthermore, the divergent coding sequence and 3'-untranslated sequence of TC22 have high homology (96.1% and 77%, respectively) to NM-4 tropomyosin isoform from rat cochlea.
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2013, Archives of Biochemistry and BiophysicsCitation Excerpt :The secondary antibodies used were fluorescein isothiocyanate (FITC)-labeled goat anti-mouse IgG and trimethylrhodamine isothiocyanate (TMRITC)-labeled goat anti-rabbit IgG. The expression plasmids used in transient transfection included pCMV-HA, pCMV-HA-p120-catenin, pCMV-Myc, pCMV-Myc-mXinα, pCMV-Myc-mXinα5′, pCMV-Myc-mXinα1R, pCMV-Myc-mXinα2R, pCMV-Myc-mXinα1R2Rp, pCMV-Myc-mXinαCTR, pEGFP-C2, pEGFPTC22 (encoding GFP fused to one of tropomyosin isoform, TC22) [23] and pcDNA-vinculin-GFP (a generous gift from Dr. Wolfgang H. Goldmann, University of Erlangen, Germany). To minimize a possible competition between two plasmids for co-expression, each transfection experiment was carried out with equal molar amount of DNAs equivalent to a combination of 1.5 μg pCMV-HA and 1.5 μg pCMV-Myc.
From skeletal muscle to cancer: Insights learned elucidating the function of tropomyosin
2012, Journal of Structural BiologyEffect of Eradication of Helicobacter pylori on the Histology and Cellular Phenotype of Gastric Intestinal Metaplasia
2008, Clinical Gastroenterology and HepatologyCitation Excerpt :However, TC22 expression was essentially the same. Thus, p53 protein accumulation seems to be a late event, whereas TC22-4 expression is an earlier event, as also reported in colonic neoplasm.40 Although the effect on cancer development will need longer follow-up evaluation, the end point of the current study was to assess the changes in the expression of biomarkers in GIM, one associated with cell phenotype and the other associated with epithelial neoplasia, but not the development of gastric cancer after H pylori treatment.
Salicylate modulates oxidative stress in the rat colon: A proteomic approach
2006, Biochemical PharmacologyCitation Excerpt :Vitamin E deficiency appears to alter tropomysosin isomers. Altered tropomyosin isomer expression was observed in a previous study in colon from rats treated with the alkylating agent, dimethylhydrazine [24] and altered tropomysosin profiles are associated with colon pathology [40,41]. Salicylic acid supplementation of Vitamin E deficient rats appears to regulate some of the identified actin-binding proteins in colon (schinderin, profilin-1) in a similar way to rats sufficient in Vitamin E, with opposing effects on expression levels in Vitamin E deficient rats.
Tropomyosin isoforms: Divining rods for actin cytoskeleton function
2005, Trends in Cell Biology
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Address requests for reprints to: Kiron M. Das, M.D., Ph.D., UMDNJ–Robert Wood Johnson Medical School, 1 Robert Wood Johnson Place, New Brunswick, New Jersey 08903. Fax: (732) 235-7792.
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Supported by grants (NIDDK47673 and HD18577) from the National Institutes of Health. J.-R.Y. was partly supported by grant 2000-1-20200-003-1 from the Basic Research program of the Korean Science and Engineering Foundation. X.G. is supported in part by a research grant from the Crohn's and Colitis Center of New Jersey.
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J.L-C.L. and X.G. contributed equally to this work.