Using a Saccharomyces cerevisiae strain having the activities of serine O-acetyl-transferase (SATase), O-acetylserine/ O-acetylhomoserine sulphydrylase (OAS/OAH SHLase), cystathionine-synthase (-CTSase) and cystathionine-lyase (-CTLase), we individually disrupted CYS3 (coding for-CTLase) and CYS4 (coding for-CTSase). The obtained gene disruptants were cysteine-dependent and incorporated the radioactivity of 35 S-sulphate into homocysteine but not into cysteine or glutathione. We concluded, therefore, that SATase and OAS/OAH SHLase do not constitute a cysteine biosynthetic pathway and that cysteine is synthesized exclusively through the pathway constituted with-CTSase and-CTLase; note that OAS/OAH SHLase supplies homocysteine to this pathway by acting as OAH SHLase. From further investigation upon the cys3-disruptant, we obtained results consistent with our earlier suggestion that cysteine and OAS play central roles in the regulation of sulphate assimilation. In addition, we found that sulphate transport activity was not induced at all in the cys4-disruptant, suggesting that CYS4 plays a role in the regulation of sulphate assimilation.

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BackgroundS100 family proteins have recently been identified as biomarkers in various cancers. Of this protein family, S100A14 and S100A16 are also believed to play an important role in tumor progression. The aim of the present study was to clarify the clinical significance and functional role of these molecules in breast cancer.MethodsIn a clinical study, an immunohistochemical analysis of S100A14 and S100A16 expression in archival specimens of primary tumors of 167 breast cancer patients was performed. The relationship of S100A14 and S100A16 expression to patient survival and clinicopathological variables was statistically analyzed. In an experimental study, the subcellular localization and function of these molecules was examined by using the human breast cancer cell lines MCF7 and SK-BR-3, both of which highly express S100A14 and S100A16 proteins. Cells transfected with expression vectors and siRNA for these genes were characterized using in vitro assays for cancer invasion and metastasis.ResultsImmunohistochemical analysis of 167 breast cancer cases showed strong cell membrane staining of S100A14 (53% of cases) and S100A16 (31% of cases) with a significant number of cases with co-expression (p < 0.001). Higher expression levels of these proteins were significantly associated with a younger age (<60 years), ER-negative status, HER2-positive status and a poorer prognosis. Co-expression of the two proteins showed more aggressive features with poorer prognosis. In the human breast cancer cell lines MCF7 and SK-BR-3, both proteins were colocalized on the cell membrane mainly at cell-cell attachment sites. Immunoprecipitation and immunofluorescence analyses demonstrated that the 100A14 protein can bind to actin localized on the cell membrane in a calcium-independent manner. A Boyden chamber assay showed that S100A14 and S100A16 knockdown substantially suppressed the invasive activity of both cell lines. Cell motility was also inhibited by S100A14 knockdown in a modified dual color wound-healing assay.ConclusionsTo our knowledge, this is the first report showing the correlation of expression of S100A14, S100A16, and co-expression of these proteins with poor prognosis of breast cancer patients. In addition, our findings indicate that S100A14 and S100A16 can promote invasive activity of breast cancer cells via an interaction with cytoskeletal dynamics. S100A14 and S100A16 might be prognostic biomarkers and potential therapeutic targets for breast cancer.Electronic supplementary materialThe online version of this article (doi:10.1186/s12885-015-1059-6) contains supplementary material, which is available to authorized users.

Background: We have previously described an alternative invasion-independent pathway of cancer metastasis in a murine mammary tumor model. This pathway is initiated by intravasation of tumor nests enveloped by endothelial cells of sinusoidal vasculature within the tumor. In this study, we examined whether evidence for the invasion-independent pathway of metastasis is present in human cancers.

Muscle wasting or sarcopenia contributes to morbidity and mortality in patients with cancer, renal failure, or heart failure, and in elderly individuals. Na+-K+-2Cl− cotransporter 1 (NKCC1) is highly expressed in mammalian skeletal muscle, where it contributes to the generation of membrane ion currents and potential. However, the physiologic function of NKCC1 in myogenesis is unclear. We investigated this issue using the NKCC1 inhibitors bumetanide and furosemide, which are commonly used loop diuretics. NKCC1 protein levels increased during C2C12 murine skeletal myoblast differentiation, similarly to those of the myogenic markers myogenin and myosin heavy chain (MHC). NKCC1 inhibitors markedly suppressed myoblast fusion into myotubes and the expression of myogenin and MHC. Furthermore, phosphorylated and total NKCC1 levels were elevated in mouse skeletal muscles after 6 weeks’ voluntary wheel running. Immunofluorescence analyses of myofiber cross-sections revealed more large myofibers after exercise, but this was impaired by daily intraperitoneal bumetanide injections (0.2 or 10 mg/kg/day). NKCC1 plays an essential role in myogenesis and exercise-induced skeletal muscle hypertrophy, and sarcopenia in patients with renal or heart failure may be attributable to treatment with loop diuretics.