The protein-tyrosine phosphatase Shp2 plays an essential role in growth factor and integrin signaling, and Shp2 mutations cause developmental defects and/or malignancy. Previous work has placed Shp2 upstream of Ras. However, the mechanism of Shp2 action and its substrate(s) are poorly defined. Additional Shp2 functions downstream of, or parallel to, Ras/Erk activation also are proposed. Here, we show that Shp2 promotes Src family kinase (SFK) activation by regulating the phosphorylation of the Csk regulator PAG/Cbp, thereby controlling Csk access to SFKs. In Shp2-deficient cells, SFK inhibitory C-terminal tyrosines are hyperphosphorylated, and the tyrosyl phosphorylation of multiple SFK substrates, including Plcgamma1, is decreased. Decreased Plcgamma1 phosphorylation leads to defective Ras activation on endomembranes, and may help account for impaired Erk activation in Shp2-deficient cells. Decreased phosphorylation/activation of other SFK substrates may explain additional consequences of Shp2 deficiency, including altered cell spreading, stress fibers, focal adhesions, and motility.

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contributed equally to this workIn Caenorhabditis elegans, histone acetyltransferase CBP-1 counteracts the repressive activity of the histone deacetylase HDA-1 to allow endoderm differentiation, which is speci®ed by the E cell. In the sister MS cell, the endoderm fate is prevented by the action of an HMG box-containing protein, POP-1, through an unknown mechanism. In this study, we show that CBP-1, HDA-1 and POP-1 converge on end-1, an initial endoderm-determining gene. In the E lineage, an essential function of CBP-1 appears to be the activation of end-1 transcription. We further identify a molecular mechanism for the endoderm-suppressive effect of POP-1 in the MS lineage by demonstrating that POP-1 functions as a transcriptional repressor that inhibits inappropriate end-1 transcription. We provide evidence that POP-1 represses transcription via the recruitment of HDA-1 and UNC-37, the C.elegans homolog of the co-repressor Groucho. These ®ndings demonstrate the importance of the interplay between acetyltransferases and deacetylases in the regulation of a critical cell fate-determining gene during development. Furthermore, they identify a strategy by which concerted actions of histone deacetylases and other co-repressors ensure maximal repression of inappropriate cell type-speci®c gene transcription. Keywords: C.elegans embryonic development/CBP-1/ histone acetylase/histone deacetylase/transcriptional repression

Loss of TGFBI, a secreted protein induced by transforming growth factor-b, has been implicated in cell proliferation, tumor progression, and angiogenesis by in vitro studies. However, in vivo antitumor functions of TGFBI as well as the underlying molecular mechanism are not well understood. To these aims, we have generated a mouse model with disruption of TGFBI genomic locus. Mice lacking TGFBI show a retarded growth and are prone to spontaneous tumors and 7,12-dimethylbenz(a)anthracene-induced skin tumors. In relation to wild-type (WT) mouse embryonic fibroblasts (MEF), TGFBI À/À MEFs display increased frequencies of chromosomal aberration and micronuclei formation and exhibit an enhanced proliferation and early S-phase entry. Cyclin D1 is up-regulated in TGFBI À/À MEFs, which correlates with aberrant activation of transcription factor cyclic AMPresponsive element binding protein (CREB) identified by chromatin immunoprecipitation and luciferase reporter assays. TGFBI reconstitution in TGFBI À/À cells by either retroviral infection with WT TGFBI gene or supplement with recombinant mouse TGFBI protein in the culture medium leads to the suppression of CREB activation and cyclin D1 expression, and further inhibition of cell proliferation. Cyclin D1 up-regulation was also identified in most of the tumors arising from TGFBI À/À mice. Our studies provide the first evidence that TGFBI functions as a tumor suppressor in vivo.

Controversy has arisen as to the role of transforming growth factor-β-induced protein (TGFBI) in the regulation of tumor metastasis. Using lung and breast cancer cell lines (H522 and MCF7, respectively), we established that TGFBI induced cell adhesion to extracellular matrix proteins by activating adhesion-associated signaling and subsequent structure reformation, ultimately leading to cells less motile; whereas TGFBI reduced abilities of colony formation in soft agar, penetration through matrix gel, and activation of matrix metalloproteinases 2 and 9. Furthermore, injection of TGFBI-expressing cells into immuno-deficient mice resulted in a significant reduction in tumor metastasis in vivo. Taken together, these data suggest that TGFBI moderates the metastatic potential of cancer cells.

Background:Although radiation-induced bystander effects have been confirmed using a variety of endpoints, the mechanism(s) underlying these effects are not well understood, especially for in vivo study.Methods:A 1-cm2 area (1 cm × 1 cm) in the lower abdominal region of gpt delta transgenic mice was irradiated with 5 Gy of 300 keV X-rays, and changes in out-of-field lung and liver were observed.Results:Compared with sham-treated controls, the Spi− mutation frequency increased 2.4-fold in non-targeted lung tissues at 24 h after partial body irradiation (PBIR). Consistent with dramatic Cyclooxygenase 2 (COX-2) induction in the non-targeted bronchial epithelial cells, increasing levels of prostaglandin, together with 8-hydroxydeoxyguanosine, in the out-of-field lung tissues were observed after PBIR. In addition, DNA double-strand breaks and apoptosis were induced in bystander lung tissues after PBIR.Conclusion:The PBIR induces DNA damage and mutagenesis in non-targeted lung tissues, especially in bronchial epithelial cells, and COX-2 has an essential role in bystander mutagenesis.

Background Transforming growth factor β induced (TGFBI) product, an extracellular matrix (ECM) protein, has been implicated as a putative tumor suppressor in recent studies. Our previous findings revealed that expression of TGFBI gene is down-regulated in a variety of cancer cell lines and clinical tissue samples. In this study, ectopic expression of TGFBI was used to ascertain its role as a tumor suppressor and to determine the underlying mechanism of mesothelioma and breast cancer. Methods Cells were stably transfected with pRc/CMV2-TGFBI and pRc/CMV2-empty vector with Lipofectamine Plus. Ectopic expression of TGFBI was quantified by using quantitative PCR and Western-blotting. Characterization of cell viability was assessed using growth curve, clonogenic survival and soft agar growth. The potential of tumor formation was evaluated by an in vivo mouse model. Cell cycle was analyzed via flow cytometry. Expressions of p21, p53, p16 and p14 were examined using Western-blotting. Senescent cells were sorted by using a Senescence β-Galactosidase Staining Kit. Telomerase activity was measured using quantitative telomerase detection kit. Results In this study, an ectopic expression of TGFBI in two types of cancer cell lines, a mesothelioma cell line NCI-H28 and a breast cancer cell line MDA-MB-231 was found to have reduced the cellular growth, plating efficiency, and anchorage-independent growth. The tumorigenicity of these cancer cell lines as determined by subcutaneous inoculation in nude mice was similarly suppressed by TGFBI expression. Likewise, TGFBI expression reduced the proportion of S-phase while increased the proportion of G1 phase in these cells. The redistribution of cell cycle phase after re-expression of TGFBI was correspondent with transiently elevated expression of p21 and p53. The activities of senescence-associated β-galactosidase and telomerase were enhanced in TGFBI-transfected cells. Conclusion Collectively, these results imply that TGFBI plays a suppressive role in the development of mesothelioma and breast cancer cells, possibly through inhibitions of cell proliferation, delaying of G1-S phase transition, and induction of senescence.