New mutation affecting the synthesis of some membrane proteins and sporulation in Bacillus subtilis.

A new mutation, mpo, which affects the synthesis of some membrane proteins and sporulation in Bacillus subtilis was identified. The mpo mutation was tightly linked to the overproduction of membrane proteins MP32 and MP18 (molecular weights of 32,000 and 18,000, respectively) and the temperature-sensitive sporulation phenotype. Genetic analysis showed that the mpo mutation maps between the spoIIIB and lys loci.     

Tail-associated structural protein gp61 of Staphylococcus aureus phage φMR11 has bifunctional lytic activity

A tailed bacteriophage, φMR11 (siphovirus), was selected as a candidate therapeutic phage against Staphylococcus aureus infections. Gene 61, one of the 67 ORFs identified, is located in the morphogenic module. The gene product (gp61) has lytic domains homologous to CHAP (corresponding to an amidase function) at its N-terminus and lysozyme subfamily 2 (LYZ2) at its C-terminus. Each domain of gp61 was purified as a recombinant protein. Both the amidase [amino acids (aa) 1–150] and the lysozyme (aa 401–624) domains but not the linker domain (aa 151–400) caused efficient lysis of S . aureus . Immunoelectron microscopy localized gp61 to the tail tip of the φMR11 phage. These data strongly suggest that gp61 is a tail-associated lytic factor involved in local cell-wall degradation, allowing the subsequent injection of φMR11 DNA into the host cytoplasm. Staphylococcus aureus lysogenized with φMR11 was also lysed by both proteins. Staphylococcus aureus strains on which φMR11 phage can only produce spots but not plaques were also lysed by each protein, indicating that gp61 may be involved in 'lysis from without'. This is the first report of the presence of a tail-associated virion protein that acts as a lysin, in an S. aureus phage.     

Cryopreservation of spermatozoa of a transgenic mouse]

Spermatozoa of a homozygous transgenic mouse, in which the firefly luciferase gene was expressed under the control of beta-actin promoter, were frozen at -196 degrees C. One fourth of the frozen sperm was later thawed and used for in vitro fertilization. Thirty-six of 65 oocytes (55.4%) developed to the 2-cell stage. All the 2-cell embryos were transferred to the oviducts of pseudopregnant recipients and 23 young (63.9%, 23/36) were born. All of young analyzed carried the transgene and showed the luciferase gene expression.     

Sex pheromones in maleMelittobia parasitic wasps: Female response to conspecific and congeneric males of 3 species

Presented a choice between conspecific males and 2 congeneric males, virgin females ofMelittobia australica andM. digitata chose conspecific males disproportionately more often, whereasM. femorata females distributed themselves evenly among the choices. Empty tubes, provided as the fourth choice in the test apparatus, were entered much less often than tubes containing live males. Females of all species chose “wrong” males about equally frequently. These observations suggest that even non-conspecific males possess some degree of attractiveness to virgin females. Chemicals in the sex pheromone of the males are presumed to be the source of the males' attractancy. The incomplete species specificity is interpreted in light of the life history of this genus, and it is suggested that specific recognition cues operate primarily after the sexes come together. Supported by a grant from Kagoshima Prefecture in Japan under the exchange program of faculty members between Kagoshima University and the University of Georgia.     

Inhibition of tumor metastasis of lewis lung carcinoma in C57BL/6 mice by intrapleural administration of Lactobacillus casei

Summary The antimetastatic effect of Lactobacillus casei YIT9018 (LC 9018) against Lewis lung carcinoma (3LL) in C57BL/6 mice was determined. Intrapleural (i.pl.) administration of LC 9018 was effective in inhibiting pulmonary metastasis after s.c. inoculation of 3LL tumors into C57BL/6 mice. The combination of i.pl. and intralesional or i.v. injections of LC 9018 also markedly inhibited pulmonary metastasis in 3LL-bearing mice. The i.pl. administration of LC 9018 into mice induced an increase in the number of thoracic exudate cells (TEC) and the cell population in the TEC was mainly polymorphonuclear leukocytes in the early stage, while macrophages were dominant in the late stage. In addition, in vitro cytolytic activity against 3LL cells and natural killer cell activity of TEC were augmented by the i.pl. administration of LC 9018. Furthermore, i.pl. administration of LC 9018 into the mice rendered their lung macrophages tumoricidal for 3LL cells in vitro. These results show that TEC induced by i.pl. administration of LC 9018 played a key role in the inhibtion of metastasis in 3LL-bearing mice.     

The different effects of recombinant human tumor necrosis factor on rat fibrosarcoma sublines

Summary The antitumor effect of recombinant human tumor necrosis factor (rH-TNF) on two clones of rat fibrosarcoma with different metastatic potential to lymph nodes was examined. The colony formation of clone A, which has high metastatic potential, was completely inhibited by continuous exposure to rH-TNF at 50 U/ml. In contrast, colony formation of clone G, which has low metastatic potential, was not inhibited by high concentrations of rH-TNF (10,000 U/ml). The inhibitory effect of rH-TNF on colony formation by clone A was also observed with a 1-h exposure to rH-TNF. This effect was time and concentration dependent, as determined by the colony assay, ³H-thymidine uptake assay, and ⁵¹Cr-release assay. ³H-thymidine and ³H-uridine uptake per cell of clone A exposed to rH-TNF was not decreased. This suggests that the mechanisms of the antitumor effect of rH-TNF were not due to inhibition of DNA and RNA synthesis of tumor cells. In vivo growth and lymph node metastases of clone A inoculated i.p. to Donryu strain rats were completely suppressed by 14 consecutive i.p. injections of 10⁵ or 10⁶ U/kg per day of rH-TNF. On the other hand the growth of clone G was not influenced by rH-TNF administration.     

Distribution of spermine in bacilli and lactic acid bacteria

Obligate moderately thermophilic bacilli and obligate moderately thermoacidophilic bacilli contained spermine as the major polyamine in addition to putrescine and spermidine. The identity of spermine was confirmed by thin-layer chromatography and high-performance liquid chromatography before and after treatment with putrescine oxidase. Using these methods, thermospermine and spermine can be separated; thermospermine was not present in these organisms. On the other hand, various facultative thermophiles and mesophilic strains of the genus Bacillus, including alkalophiles and halophiles, lack spermine and other tetraamines. No spermine was detected in several strains of mesophilic or facultative slightly thermophilic lactic acid bacteria, Lactobacillus and Streptococcus.     

Opioid-binding protein (OBCAM) is rich in β-sheets

Based on circular dichroism (CD) and the sequence-predictive method, the opioid-binding cell adhesion molecule (OBCAM) consisted of one half -sheets and one fourth -helices. This is consistent with significant sequence homology of the protein to several members of the immunoglobulin (Ig) superfamily, particularly cell adhesion molecules, which are rich in -sheets. Hydropathy analysis suggests that hydrophobic and hydrophilic regions were evenly distributed along the sequence, but the NH₂- and COOH-termini were hydrophobic. Hydrophobic moments and Fourier-transform amphipathic analyses further suggest that residues 23–30 and 83–93 were amphiphathie -sheets. The overall conformation of OBCAM was unaltered by adding linoleic acid, which is required for opioid ligand binding.