Differential Reduction of Tellurite by Growing Colonies of Normal Yeast and Respiration-Deficient Mutants.

Nagai, Susumu (National Women's University, Nara, Japan). Differential reduction of tellurite by growing colonies of normal yeast and respiration-deficient mutants. J. Bacteriol. 90:220-222. 1965.-A differential reduction of sodium tellurite was observed between normal and respiration-deficient mutant colonies of several species of Saccharomyces. Normal colonies turned black in contrast to mutant colonies which remained nearly white when grown on an agar medium containing 30 to 40 mg per liter of tellurite. Schopfer's medium enriched with yeast extract and a mixture of vitamins was most suitable to develop such black-and-white contrast. The difference was far less obvious when the asparagine of this medium was replaced by other nitrogen sources such as glutamate, peptone, or Casamino Acids. Addition of ammonium sulfate to the medium weakened and sometimes completely reversed the contrast. The usefulness of tellurite medium for diagnostic color differentiation of respiration deficiency was considered.     

Survival of cultured cells and somatic embryos of Asparagus officinalis cryopreserved by vitrification

Cultured cells and somatic embryos derived from the mesophyll tissue of asparagus (Asparagus officinalis L.) were cryopreserved by vitrification. The vitrification solution (PVS) contains (w/v) 22% glycerol, 15% ethylene glycol, 15% propylene glycol and 7% DMSO in Murashige-Skoog medium enriched with 0.5M sorbitol. After initial cryoprotection with sorbitol supplemented MS medium containing 12% ethylene glycol, cells or embryos were exposed stepwise to 85% PVS at 0°C. They were loaded into 0.5 ml transparent straws, and were then plunged directly into liquid nitrogen. After rapid warming, PVS was removed and diluted stepwise. The highest survivals of vitrified cells and embryos were about 65 and 50%, respectively. Surviving embryos developed into plantlets.Abbreviations DMSO dimetyl sulfoxide - PVS vitrification solution - LN liquid nitrogen - DSC differential scanning calorimeter - MS Murashige-Skoog salt medium - NAA naphthalene acetic acid - BA 6-benzyladenine     

The role of thromboxane A2 (TxA2) in allergic cutaneous reactions and the effect of (E)-3-[p-(1H-imidazol-1-ylmethyl) phenyl]-2-propenoic acid hydrochloride (OKY-046), a TxA2 synthetase inhibitor

To study the role of thromboxane A2 (TxA2) in cutaneous allergic reactions, the effect of (E)-3-[p-(1H-Imidazol-1-ylmethyl)phenyl]-2-propenoic acid hydrochloride (OKY-046), a selective TxA2 synthetase inhibitor, on cutaneous reactions in rats and mice was studied. Simultaneously, the effect of 9,11-methanoepoxy-prostaglandin H2 (U-46619), a stable analogue of TxA2, on capillary permeability in mouse and rat skin was investigated. Passive cutaneous anaphylaxis (PCA) in mouse ear was clearly inhibited by OKY-046 but not by indomethacin. The inhibitory action of OKY-046 was not influenced by pretreatment with indomethacin. Moreover, prostaglandin I2, which accumulated as a result of the inhibition of TxA2 synthetase, did not affect the PCA. But, the dye leakages caused by histamine, serotonin and leukotriene C4 in mouse ear were clearly inhibited by OKY-046. In addition, OKY-046 inhibited rat reversed cutaneous anaphylaxis, but its inhibitory action was not affected by pretreatment with indomethacin. Contrary to the above results, rat footpad passive Arthus reaction and mouse footpad tuberculin delayed hypersensitivity reaction were not affected by OKY-046. Additionally, U-46619 did not cause an increase of capillary permeability in either mouse and rat skin. These results suggest a slight role of TxA2 in cutaneous allergic reactions in mice and rats and the efficacy of OKY-046 on Type I and II reactions regardless of the inhibition of TxA2 synthetase activity.     

Primary structure of chicken pituitary prolactin deduced from the cDNA sequence. Conserved and specific amino acid residues in the domains of the prolactins

The perform of chicken prolactin (PRL) deduced from the cDNA sequence contains a signal peptide of 30 amino acid residues followed by a mature PRL of 199 residues. Chicken PRL shows 77, 68, 67, 58, and 31% identity of amino acid sequence with whale, human, ovine, rat, and salmon PRLs, respectively. Elucidation of the primary structure of avian PRL enabled extended analysis of the specific and conserved amino acid residues and domains of the PRL molecules. The mammalian, teleostean, and avian PRLs share 32 common residues, and these conserved residues are observed to cluster in four distinct domains (PD1 to PD4), corresponding to four of five conserved domains of the growth hormones. Of the 32 residues, 8 residues in the PD2 and PD4 domains, including 4 cysteines, are conserved by other members of the growth hormone family, which indicates that these 8 residues may be essential for common structural features of the gene family. On the other hand, 13 other residues distributed among all four domains are conserved almost exclusively in the PRLs, suggesting that these residues are indispensable for specific binding of the PRLs to their receptors.     

Identification of two types of smooth muscle myosin heavy chain isoforms by cDNA cloning and immunoblot analysis

We previously reported the characterization of a rabbit uterus cDNA clone (SMHC29) which encoded part of the light meromyosin of smooth muscle myosin heavy chain (Nagai, R., Larson, D.M., and Periasamy, M. (1988) Proc. Natl. Acad. Sci. U. S. A. 85, 1047-1051). We have now characterized a second cDNA clone (SMHC40) which also encodes part of the light meromyosin but differs from SMHC29 in the following respects. Nucleotide sequence analysis demonstrates that the two myosin heavy chain mRNAs are identical over 1424 nucleotides but differ in part of the 3'-carboxyl coding region and a portion of the 3'-nontranslated sequence. Specifically, SMHC40 cDNA encodes a unique stretch of 43 amino acids at the carboxyl terminus, whereas SMHC29 cDNA contains a shorter carboxyl terminus of 9 unique amino acids which is the result of a 39-nucleotide insertion. Recent peptide mapping of smooth muscle myosin heavy chain identified two isotypes with differences in the light meromyosin fragment that were designated as SM1 (204 kDa) and SM2 (200 kDa) type myosin (Eddinger, T. J., and Murphy, R.A. (1988) Biochemistry 27, 3807-3811). In this study we present direct evidence that SMHC40 and SMHC29 mRNA encode the two smooth muscle myosin heavy chain isoforms, SM1 and SM2, respectively, by immunoblot analysis using antibodies against specific carboxyl terminus sequences deduced from SMHC40 and SMHC29 cDNA clones.     

Diacylglycerol, but not inositol 1,4,5-trisphosphate, accounts for platelet-derived growth factor-stimulated proliferation of BALB 3T3 cells

Recently we found that an intracellular event related to phosphatidylinositol 4,5-bisphosphate (PIP2) is crucial for platelet-derived growth factor (PDGF)-induced mitogenesis in fibroblastic cells (Matuoka, K., et al.: Science 239:640-643, 1988). In the present study we examined the mitogenic effects of PIP2 and its hydrolysis products introduced into the cytoplasm of BALB 3T3 cells by micro-injection to confirm the role of PIP2 hydrolysis in PDGF stimulation of cell proliferation. Injection of 1,2-dioleylglycerol (diolein) into serum-deprived quiescent cells induced DNA synthesis with the same time course as that induced by exposure of the cells to PDGF and, in the presence of PDGF, caused no additional increase in the cell population entering S phase. The injection of PIP2, inositol 1,4,5-trisphosphate, or 1,2-dioleylphosphatidic acid into the cells did not induce mitogenesis. Consistent results were obtained in experiments in which the cells were exposed to 1-oleyl-2-acetylglycerol (OAG) and ionomycin; namely, OAG stimulated proliferation of BALB 3T3 cells, but ionomycin did not induce any mitogenesis. Desensitization of the protein kinase C pathway by prolonged exposure of the cells to phorbol ester abolished the induction of cell proliferation by subsequent injection of diolein or exposure to phorbol ester or OAG as well as by PDGF challenge. These findings strongly suggest that activation of the protein kinase C system following formation of diacylglycerol by PIP2 hydrolysis is mainly responsible for the mitogenic action of PDGF on BALB 3T3 cells.