Somatic Cell and Molecular Genetics最新文献

The hemagglutinating virus of Japan (HVJ) fused with liposomes provides a unique transfection vehicle with characteristics of both virus vector and liposome. Here we investigate the efficiency and safety of the HVJ-liposome technique in delivering foreign genes and oligonucleotides into the lung of the Wistar rat. A plasmid vector containing the Escherichia coli beta-galactosidase (beta-gal) gene and the chicken beta-actin promoter was transfected via the trachea using the HVJ-liposome method. Cytochemical staining showed expression of exogenous beta-gal activity in airway epithelial cells, alveolar macrophages, and alveolar type II cells. This activity persisted at least 28 days after administration of the genes. FITC-labeled oligonucleotides also were introduced into the same types of lung cells as those expressing beta-gal. After instillation of HVJ-liposome, anti-HVJ antibodies were detected in the sera of the rats, but even after repeated administration of HVJ-liposome, no marked histopathologic change was observed while exogenous beta-gal expression was detected in pulmonary cells.

The cytogenetical reaction of human peripheral lymphocytes to okadaic acid and calyculin A was examined. Calyculin A could induce PCC about 20 times more effectively than okadaic acid. Their mechanisms of PCC induction were judged similar by their dose-dependent manner and chromosome morphology. Contrary to earlier studies suggesting that chemicals could not induce PCC in G1 cells where little cyclin B is present, the present study showed that calyculin A could induce PCC in lymphocytes not only at S and G2/M but also at the second G1 phase after PHA stimulation in vitro. PCC was induced slightly in lymphocytes both at G0 and the first in vitro G1 phase even when the calyculin A concentration increased one hundred fold. It was found that calcium ionophore A23187 increased frequencies of G0-PCC induced by calyculin A, although a further refinement is necessary to obtain a suitable morphology of G0-PCC for cytogenetic studies.

A number of human homologues of yeast cell cycle checkpoint control genes have been identified recently. In this study, the sequence alterations in six of such novel human genes (hRAD1, hRAD9, hRAD17, hHUS1, CHK1 and CHES1) were analyzed by PCR-single-strand conformational polymorphism (PCR-SSCP) method on a panel of 25 human tumor cell lines in an attempt to search for possible in vivo cases where any of the checkpoint-related genes are altered in human systems. For hRAD9, hHUS1 or CHK1, no SSCP variant was detected in any of the cell lines tested, indicating a high stability of these genes in human cancer. Most of the SSCP variants found in the other three genes were due to single nucleotide base substitutions. Two cell lines were found to be homozygous for missense-type base substitutions, i.e., Saos-2 was homoallelic for 1637T-->G in hRAD17; and COLO320DM for 1189G-->A in CHES1, indicating a possible use of these cell lines for further study. The former nucleotide change in hRAD17, which causes a change of amino acid from arginine to lysine at codon 546, was supposed to be polymorphic. Considering that lysine, but not arginine, is the amino acid that is well conserved among fission yeast, mouse and monkey at the corresponding position, coexistence of both alleles in human may have a functional or selectional implication.

RNA helicase A is the homolog of the Drosophila maleless protein, an essential factor involved in dosage compensation, and plays a crucial role in early development in mammals. Here, we have mapped the human RNA helicase A (DDX9) gene to the major susceptibility locus for prostate cancer at chromosome band 1q25, and its pseudogene (DDX9P) to the band 13q22 by fluorescence in situ hybridization, somatic cell hybrid analysis, and assignment of YAC clones, respectively.

Many anticancer drugs inhibit DNA replication. To investigate the mechanism of permanent growth inhibition after transient arrest of DNA replication, we selected genetic suppressor elements (GSEs) conferring resistance to replication inhibitor Aphidicolin. Starting from a retroviral expression library carrying normalized fragments of human cell cDNA, we isolated four GSEs which, when introduced as a combination, produced resistance to Aphidicolin, doxorubicin and hydroxyurea in HT1080 fibrosarcoma cells. The four GSEs were derived from ORFX bromodomain protein gene, WIZ zinc finger protein gene, the gene for subunit 3 of cytochrome c oxidase, and the gene corresponding to an EST with no known function. A cell line carrying all four GSEs showed a weaker induction of the senescence-like phenotype after treatment with Aphidicolin or doxorubicin; the resistance of this cell line was not associated with decreased doxorubicin accumulation. These results indicate that combined effects of GSEs derived from these four genes increase cellular resistance to replication-inhibiting drugs, possibly by inhibiting drug-induced senescence.