UV micro-irradiation of the Chinese hamster cell nucleus and caffeine post-treatment immunocytochemical localization of DNA photolesions in cells with partial and generalized chromosome shattering

UV micro-irradiation of a small part of the Chinese hamster nucleus and caffeine post-incubation often results in shattered chromosomes at the first post-irradiation mitosis. In some of these mitotic cells, chromosome shattering is restricted to a few chromosomes spatially related in a small area of the metaphase spread; in others, shattering includes the whole chromosome complement. These 2 types of damage have been called partial and generalized chromosome shattering (PCS and GCS).Using antisera that specifically react with UV-irradiated DNA, we identified micro-irradiated chromatin in interphase nuclei and in mitotic cells with PCS or GCS by indirect immunofluorescence microscopy. In PCS, immunofluorescence staining was found in the damaged area, while the surrounding intact chromosomes were not stained. In GCS, staining was also restricted to a small region of the shattered chromosome complement. In other experiments, cells synchronized in G1 were micro-irradiated in the nucleus, pulse-labelled with [³H]thymidine and post-incubated with caffeine. Autoradiographs of cells with GCS showed unscheduled DNA synthesis restricted to a small chromatin region.Our data present direct evidence that the distribution of DNA photolesions does not coincide with the sites of chromosomal damage in GCS. As a working hypothesis, we propose that an indirect mechanism is involved in the induction of GCS by which DNA photolesions in a small nuclear segment induce shattering of both micro-irradiated and non-irradiated chromosomes.     

Distribution of branched-chain fatty acid in the skin surface lipid of laboratory animals

1. The distribution of the branched-chain fatty acid (BCFA) was studied in skin surface lipids of laboratory animals (rat, mouse, hamster and rabbit) and the experimental animal for the study of the metabolic fate of BCFA was chosen. 2. The monoester fraction resistant to microbial degradation was the index of which fatty acids were identified and the compositions were analyzed by capillary gas chromatography and mass spectrometry (GC-MS). 3. The contents of monoester fractions in rat, mouse, hamster and rabbit were 78.9, 15.9, 30.4 and 45.6% of the total skin surface lipid, respectively. 4. BCFAs were exclusively identified to be either iso- or anteiso-series type by means of mass spectrometry. 5. The sum of iso- and anteiso-acid was the highest in hamster comprising 53% in male and 38% in female of the monoester acid. 6. Sex related differences of BCFA concentration were not so evident in the other three species. 7. BCFA contents in the monoester fractions were as follows: rat 32%, mouse 25% and rabbit 3%. 8. Concentrations of iso-series fatty acid were consistently higher than that of anteiso-series type in all animals studied. 9. Abundance of 2-hydroxy fatty acid in the rabbit monoester fraction was noted for the first time. 10. Suitability of these laboratory animals for the study of BCFA metabolism was discussed.     

Activation of cellular protein kinase C and mode of inhibitory action of phospholipid-interacting compounds

12-O-Tetradecanoylphorbol-13-acetate (TPA), that is intercalated into the cell membrane, binds a roughly stoichiometric amount of protein kinase C to produce a catalytically active complex with phospholipid. Local anesthetics and other phospholipid-interacting compounds such as chlorpromazine inhibit profoundly this complex formation in a manner competitive with phospholipid but not with TPA. A tiny change of the membrane phospholipid bilayer structure that is caused by TPA appears to facilitate this unique phospholipid-protein kinase C interaction.     

Molecular cloning of engrailed: a gene involved in the development of pattern in Drosophila melanogaster

The engrailed gene acts early in Drosophila embryogenesis and plays an essential role in the processes that establish and maintain the repeating segmental pattern. To begin molecular analysis of the role of the engrailed gene in embryonic pattern formation, we used a chromosomal walk to clone genomic sequences that encompass the locus, and have physically mapped the positions of 15 engrailed mutations. The positions of engrailed rearrangement mutations indicate that the engrailed complementation unit includes a minimum of 70 kb. The locus can be divided into two regions. Rearrangement mutations interrupting the centromere proximal 50 kb of the locus result in embryonic lethality while mutants altered in the distal 20 kb of the locus survive to show morphological abnormalities in several adult segments. It appears that long-range cis interactions play a role in the function of the engrailed gene.     

Induction of rat liver angiotensinogen mRNA following acute inflammation

Inflammatory responses of the angiotensinogen mRNA in rat liver and brain were examined by RNA blot-hybridization analysis with use of a cDNA probe specific for rat angiotensinogen. The angiotensinogen mRNA in the liver increased rapidly during the first 5 h following the administration of Escherichia coli lipopolysaccharide, and at maximum level of induction, the mRNA increased approximately 5-fold over its normal level. The levels of the mRNA increased with increasing doses of lipopolysaccharide, the half-maximal dose being approximately 1 microgram/100 g body weight. In contrast, no such increase was observed in the brain angiotensinogen mRNA. Thus, the expression of the rat angiotensinogen mRNA is regulated in a tissue-specific manner in response to induction of acute inflammation.     

Filamentous vibriophage fs2 encoding the rstC gene integrates into the same chromosomal region as cholera toxin X

The genome of the filamentous phage of Vibrio cholerae fs2 was found to contain rst C and rst B1 (truncated) genes downstream of ORF500. att -fs2-dir and att- fs2-rev sequences homologous to that of att -CTXφ were found between orf 500 and rst C of the fs2 genome. This prompted us to search for the integration site of fs2 in the genomes of V. cholerae O1 and O139. The genome of fs2 was found to integrate downstream of att RS of the CTXφ phage, which integrated into chromosome I of V. cholerae O1 and O139. When infected with fs2, a fimbriate strain of V. cholerae O1 appeared to reduce fimbrial production in an adult rabbit ileal loop assay.