Pulmonary metastases are the main cause of death in patients with osteosarcoma, however, the molecular mechanisms of metastasis are not well understood. To detect lung metastasis-related microRNA (miRNA) in human osteosarcoma, we compared parental (HOS) and its subclone (143B) human osteosarcoma cell lines showing lung metastasis in a mouse model. miR-143 was the most downregulated miRNA (P < 0.01), and transfection of miR-143 into 143B significantly decreased its invasiveness, but not cell proliferation. Noninvasive optical imaging technologies revealed that intravenous injection of miR-143, but not negative control miRNA, significantly suppressed lung metastasis of 143B (P < 0.01). To search for miR-143 target mRNA in 143B, microarray analyses were performed using an independent RNA pool extracted by two different comprehensive miR-143-target mRNA collecting systems. Western blot analyses revealed that MMP-13 was mostly protein downregulated by miR-143. Immunohistochemistry using clinical samples clearly revealed MMP-13-positive cells in lung metastasis-positive cases, but not in at least three cases showing higher miR-143 expression in the no metastasis group. Taken together, these data indicated that the downregulation of miR-143 correlates with the lung metastasis of human osteosarcoma cells by promoting cellular invasion, probably via MMP-13 upregulation, suggesting that miRNA could be used to develop new molecular targets for osteosarcoma metastasis.

A number of chromosomal abnormalities including 19q deletions have been associated with the formation of human gliomas. In this study, we employed a proteomics-based approach to identify possible genes involved in glioma tumorigenesis which may serve as potential diagnostic molecular markers for this type of cancer. By comparing protein spots from gliomas and non-tumor tissues using two-dimensional (2D) gel electrophoresis, we identified 11 up-regulated proteins and four down-regulated proteins in gliomas. Interestingly, we also discovered that a group of cytoskeleton-related proteins are differentially regulated in gliomas, suggesting the involvement of cytoskeleton modulation in glioma pathogenesis. We then focused on the cytoskeleton-related protein, SIRT2 (sirtuin homologue 2) tubulin deacetylase, which was down-regulated in gliomas. SIRT2 is located at 19q13.2, a region known to be frequently deleted in human gliomas. Subsequent Northern blot analysis revealed that RNA expression of SIRT2 was dramatically diminished in 12 out of 17 gliomas and glioma cell lines, in agreement with proteomic data. Furthermore, ectopic expression of SIRT2 in glioma cell lines led to the perturbation of the microtubule network and caused a remarkable reduction in the number of stable clones expressing SIRT2 as compared to that of a control vector in colony formation assays. These results suggest that SIRT2 may act as a tumor suppressor gene in human gliomas possibly through the regulation of microtubule network and may serve as a novel molecular marker for gliomas. Additional proteins were also identified, whose function in gliomas was previously unsuspected.

We previously identified SIRT2, an nicotinamide adenine dinucleotide (NAD)-dependent tubulin deacetylase, as a protein downregulated in gliomas and glioma cell lines, which are characterized by aneuploidy. Other studies reported SIRT2 to be involved in mitotic progression in the normal cell cycle. We herein investigated whether SIRT2 functions in the mitotic checkpoint in response to mitotic stress caused by microtubule poisons. By monitoring chromosome condensation, the exogenously expressed SIRT2 was found to block the entry to chromosome condensation and subsequent hyperploid cell formation in glioma cell lines with a persistence of the cyclin B/cdc2 activity in response to mitotic stress. SIRT2 is thus a novel mitotic checkpoint protein that functions in the early metaphase to prevent chromosomal instability (CIN), characteristics previously reported for the CHFR protein.We further found that histone deacetylation, but not the aberrant DNA methylation of SIRT2 5 0 untranslated region is involved in the downregulation of SIRT2. Although SIRT2 is normally exclusively located in the cytoplasm, the rapid accumulation of SIRT2 in the nucleus was observed after treatment with a nuclear export inhibitor, leptomycin B and ionizing radiation in normal human fibroblasts, suggesting that nucleo-cytoplasmic shuttling regulates the SIRT2 function. Collectively, our results suggest that the further study of SIRT2 may thus provide new insights into the relationships among CIN, epigenetic regulation and tumorigenesis. Oncogene (2007) 26, 945-957.

Sir2, an NAD+-dependent protein deacetylase, extends the lifespan in diverse species from yeast to flies. Mammals have seven homologs of Sir2, SIRT1-7, which affect aging and metabolism and which are potential targets for pharmacologic intervention. We identified SIRT2, which preferentially deacetylates tubulin and histone H4, as a downregulated protein in gliomas due to its epigenetic aberration. We herein discuss the role of SIRT2 in the mitotic checkpoint function and show that it may be as a potential target of anti-cancer drugs.

Aureobasidin A (AbA) is a novel cyclic depsipeptide antifungal antibiotic. The antifungal activity of AbAwas studied in vitro and in vivo in comparison with clinically effective antifungal agents, amphotericin B and fluconazole. AbAwas highly active in vitro against many pathogenic fungi, including Candida albicans, Cryptococcus neoformans, Blastomyces dermatitidis and Histoplasma capsulatum. The activity was superior to amphotericin B in most cases. AbAexhibited fungicidal action toward growing cultures of C. albicans. It was highly tolerated by mice and showed good efficacy in the treatment of murine systemic candidiasis whengiven orally or subcutaneously. AbA's fungicidal action in mice with candidiasis was more effective than fluconazole and amphotericin B.The incidence of systemic fungal infections are increasing in immunecompromisedpatients with several underlying diseases, such as leukemia and acquired immunodeficiency syndrome (AIDS), and in patients receiving anti-neoplastic agents, immunosuppressive agents or broad spectrum antibacterial antibiotics. Amphotericin B, flucytosine, and two imidazole drugs, miconazole and ketoconazole, have been the drugs of choice for systemic antifungal chemotherapy. Recently, two neworal triazoles, fluconazole and itraconazole,1) have been successfully developed and are now available for the treatment of some systemic fungal infections. However, clinical usefulness of all of these drugs is limited to insufficient therapeutic efficacy and/or toxicity. Thus there is a need for novel antifungal agents with a greater clinical efficacy.Aureobasidin A (AbA) is a new antifungal cyclic depsipeptide antibiotic (Fig. 1) produced by Aureobasidium pullulans R106.2~4) The characteristic of the structure of AbAis that of a peptide composed of eight L-form amino acids linked through one hydroxy acid to form a ring structure. There are some cyclic peptide antibiotics with antifungal activity in vitro, including valinomycin,5) syringomycin,6'7) calophycin8) and aculeacin/echinocandin family.9~1 1} Manyof them effect cell membrane permeability and have toxicity to eukaryotic cells. Several antifungal cyclic peptides with a long-chain fatty acid, such as aculeacin A, echinocandin B and their analogs, inhibit fungal cell wall synthesis with low acute toxicity. 9' 1 0) However, they show a narrow spectrum antifungal activity, ineffectiveness to murine candidiasis when given orally, and hemolytic effect. On the other hand, AbAhas a broader spectrum and higher

No abstract

Members of the Id subfamily of helix-loop-helix (HLH) proteins play important roles in promoting cell cycle entry, enhancing apoptosis, stimulating proliferation, and blocking cellular differentiation (reviewed in references 21, 28, and 30). The founding member of this subfamily, Idl, was originally identified as a protein that inhibits the DNA-binding activity of basic HLH (bHLH) proteins (4). Subsequently, three further genes that encode the related proteins Id2 (5, 41), Id3 (8, 11), and Id4 (35) were identified. Like Id1, the other Id proteins (Id2, Id3, and Id4) also inhibit DNA binding by bHLH proteins (reviewed in references 21, 28, and 30). Mechanistically, the Id proteins are thought to inhibit bHLH proteins by sequestering them in inactive heterodimers which are incapable of DNA binding due to the absence of the basic region in the Id proteins (4, 41; reviewed in references 21, 28, and 30). In addition to their association with bHLH transcription factors, Id proteins have also been shown to interact with several non-HLH proteins, including the retinoblastoma protein (pRB) and related pocket proteins (19,22,23), MIDA1 (20,38), and, more recently, members of the TCF subfamily of ETS-domain transcription factors (48). Id proteins inhibit DNA binding by the TCF proteins through interaction with their ETS DNA-binding domains. This interaction also leads to the dissociation of TCFs from ternary TCF-SRF-SRE complexes and hence to the inhibition of c-fos promoter activity (48).A subset of ETS-domain transcription factors, including Elk-1, can also form ternary complexes with the paired-domain transcription factor Pax-5 and the B-cell-specific mb-1 promoter (15). In this case, Pax-5, rather than SRF, serves to recruit the ETS-domain proteins to the promoter. Pax-5 is a member of a subfamily of Pax proteins which also contains Pax-2 and Pax-8 (reviewed in references 25 and 40). This subfamily is characterized by the presence of an octapeptide motif and a partial homeodomain in addition to the N-terminal paired DNA-binding domain. Pax-5 plays an important role in regulating B-cell development (reviewed in references 7 and 27). Several target genes have been identified, which are either up-regulated (mb-1, N-myc, and LEF-1) or down-regulated (PD-1) in keeping with the observation that Pax-5 can function as both a transcriptional activator and repressor. In the case of the mb-1 and LEF-1 genes, the paired domain of Pax-5 is sufficient to up-regulate their expression (31).As Id proteins are also expressed during B-cell development and function as negative regulators of B lymphopoiesis (9,41,42, 44), we tested whether Id proteins could affect the activity of ETS-domain protein complexes that form on the mb-1 promoter. By analogy with the ternary complex that forms on the c-fos SRE, it was expected that Id-mediated dissociation of the ETS-domain protein component might be observed.

Cytokines play several roles in developing and/or reinforcing premature cellular senescence of young cells. One such cytokine, interleukin-6 (IL-6), regulates senescence in some systems in addition to its known functions of immune regulation and promotion of tumorigenesis. In this review, we describe recent advances in studies on the roles of IL-6 and its downstream signal transducer and activator of transcription 3 (STAT3) in regulating premature cellular senescence. IL-6/sIL-6Rα stimulation forms a senescence-inducing circuit involving the STAT3-insulin-like growth factor-binding protein 5 (IGFBP5) as a key axis triggering and reinforcing component in human fibroblasts. We describe how cytokines regulate the process of senescence by activating STAT3 in one system and anti-senescence or tumorigenesis in other systems. The roles of other STAT members in premature senescence also will be discussed to show the multiple mechanisms leading to cytokine-induced senescence.