Wsc proteins have been identified in fungi and are believed to be stress sensors in the cell wall integrity (CWI) signaling pathway. In this study, we characterized the sensor orthologs WscA and WscB in Aspergillus nidulans. Using hemagglutinin-tagged WscA and WscB, we showed both Wsc proteins to be N-and Oglycosylated and localized in the cell wall and membrane, implying that they are potential cell surface sensors. The wscA disruptant (⌬wscA) strain was characterized by reduced colony and conidia formation and a high frequency of swollen hyphae under hypo-osmotic conditions. The deficient phenotype of the ⌬wscA strain was facilitated by acidification, but not by alkalization or antifungal agents. In contrast, osmotic stabilization restored the normal phenotype in the ⌬wscA strain. A similar inhibition was observed in the wscB disruptant strain, but to a lesser extent. In addition, a double wscA and wscB disruptant (⌬wscA ⌬wscB) strain was viable, but its growth was inhibited to a greater degree, indicating that the functions of the products of these genes are redundant. Transcription of ␣-1,3-glucan synthase genes (agsA and agsB) was significantly altered in the wscA disruptant strain, resulting in an increase in the amount of alkali-soluble cell wall glucan compared to that in the wild-type (wt) strain. An increase in mitogen-activated protein kinase (MpkA) phosphorylation was observed as a result of wsc disruption. Moreover, the transient transcriptional upregulation of the agsB gene via MpkA signaling was observed in the ⌬wscA ⌬wscB strain to the same degree as in the wt strain. These results indicate that A. nidulans Wsc proteins have a different sensing spectrum and downstream signaling pathway than those in the yeast Saccharomyces cerevisiae and that they play an important role in CWI under hypo-osmotic and acidic pH conditions.

PurposeMicrosatellite instability (MSI) in human endometrial cancer (EC) was analysed using a unique fluorescent technique. MSI is associated with various human neoplasms. However, the reported frequency of MSI differs widely in each malignancy. Methodological difficulties have in fact been pointed out in its assay techniques.MethodsWe previously established a sensitive fluorescent technique in which the major methodological problems are overcome. Application of this technique has revealed two distinct modes of microsatellite alterations, i.e. Type A and Type B. In the present study, we have applied this technique to 94 ECs.ResultsSignificant microsatellite alterations were observed in 38 (40.4 %) tumours of the panel. The two modes, Type A and Type B, were indeed observed in this malignancy. More importantly, we found that the modes more closely correlated with the molecular and clinicopathological backgrounds of the tumours than the established and widely used MSI grades, MSI-H and MSI-L. Type B MSI widely correlated with family history of hereditary non-polyposis colorectal cancer-associated cancers, whereas MSI-H only did with that of colorectal cancer. Furthermore, mutation in the KRAS oncogene, which has been regarded as generally infrequent in microsatellite-unstable tumours, was clearly associated with Type A MSI.ConclusionsOur observations may suggest a biological relevance and a potential utility of the modal classification of MSI and, furthermore, added complexities to genomic instability underlying tumourigenesis in human endometrium.Electronic supplementary materialThe online version of this article (doi:10.1007/s00432-015-2030-2) contains supplementary material, which is available to authorized users.

Biomarkers have revolutionized cancer chemotherapy. However, many biomarker candidates are still in debate. In addition to clinical studies, a priori experimental approaches are needed. Thymidylate synthase (TS) expression is a long-standing candidate as a biomarker for 5-fluorouracil (5-FU) treatment of cancer patients. Using the Tet-OFF system and a human colorectal cancer cell line, DLD-1, we first constructed an in vitro system in which TS expression is dynamically controllable. Quantitative assays have elucidated that TS expression in the transformant was widely modulated, and that the dynamic range covered 15-fold of the basal level. 5-FU sensitivity of the transformant cells significantly increased in response to downregulated TS expression, although being not examined in the full dynamic range because of the doxycycline toxicity. Intriguingly, our in vitro data suggest that there is a linear relationship between TS expression and the 5-FU sensitivity in cells. Data obtained in a mouse model using transformant xenografts were highly parallel to those obtained in vitro. Thus, our in vitro and in vivo observations suggest that TS expression is a determinant of 5-FU sensitivity in cells, at least in this specific genetic background, and, therefore, support the possibility of TS expression as a biomarker for 5-FU-based cancer chemotherapy.

Formalin-ˆxed para‹n-embedded (FFPE) tissues are one of the most eŠective tools for human identiˆcation in forensic science. To evaluate the eŠectiveness for human identiˆcation, we compared the following six methods for the extraction of DNA from FFPE tissues; EZ1 DNA Investigator Kit, ReliaPrep FFPE gDNA Miniprep System, DNA Isolator PS-Rapid Reagent, QIAamp DNA FFPE Tissue Kit, NucleoSpin DNA FFPE XS and phenol-chloroform method. DNA extracted using each method from two FFPE tissue blocks were quantiˆed by three diŠerent DNA quantiˆcation methods (Quant-it dsDNA HS Assay Kit and quantitative PCR based on 207 bp and 98 bp regions in the D17Z1 locus), and short tandem repeat (STR) typing using AmpFlSTR Identiˆler Plus PCR Ampliˆcation Kit was conducted.All extracts from the FFPE tissues were highly degraded, and large diŠerences were observed among total DNA yields determined by these three quantiˆcation methods. DNA obtained using QIAamp DNA FFPE Tissue Kit had the highest total DNA yield among all the quantiˆcation methods. In STR typing with 1 ng DNA template estimated by quantitative PCR based on 98 bp region in the D17Z1 locus, the greatest number of detected alleles was observed using QIAamp DNA FFPE Tissue Kit. In contrast, EZ1 DNA Investigator Kit and DNA Isolator PS-Rapid Reagent were inferior to the other methods in terms of the DNA yield and the number of detected alleles via STR typing.In conclusion, QIAamp DNA FFPE Tissue Kit was the most eŠective method

No abstract