Distribution of ribonucleic acid coliphages in south and east Asia.

We investigated the distribution of ribonucleic acid (RNA) coliphages in the Philippines, Singapore, Indonesia, India, and Thailand by collecting sewage samples from domestic drainage in November 1976. Of the 221 samples collected from domestic drainage, 50 contained RNA phages (52 strains). By serological analysis, 46 of the 52 strains were found to belong to group III. It can thus be said that the most prevalent RNA phages in Southeast Asia (at least, in the Philippines, Singapore, and Indonesia) were group III phages. Investigations of sewage samples collected from domestic drainage in Japan indicate that the most prevalent RNA phages in mainland Japan (north of Kyushu) are group II phages, whereas group III phages are predominant in the southern part of Japan (south of Amamiohshima Island). We therefore propose a borderline between Kyushu and Amamiohshima Island for the geographical distribution of RNA coliphages in the domestic drainage of South and East Asia. Moreover, one strain (ID2) was inactivated to some extent with the antisera of four groups of RNA phages. This is thought to be significant from the evolutionary viewpoint.     

An endonuclease activity in Bacillus subtilis specific for UV-irradiated DNA

An enzyme activity specific for UV-DNA¹ was found in the extract of $\text{Bacillus subtilis}$(Marburg 168). The enzyme preparation obtained from the extract by ammonium sulfate precipitation acts on UV-DNA endonucleolytically and induces single strand breaks. The number of single strand breaks introduced in DNA is proportional to UV dose.     

Overexpression of CNP in chondrocytes rescues achondroplasia through a MAPK-dependent pathway

Achondroplasia is the most common genetic form of human dwarfism, for which there is presently no effective therapy. C-type natriuretic peptide (CNP) is a newly identified molecule that regulates endochondral bone growth through GC-B, a subtype of particulate guanylyl cyclase. Here we show that targeted overexpression of CNP in chondrocytes counteracts dwarfism in a mouse model of achondroplasia with activated fibroblast growth factor receptor 3 (FGFR-3) in the cartilage. CNP prevented the shortening of achondroplastic bones by correcting the decreased extracellular matrix synthesis in the growth plate through inhibition of the MAPK pathway of FGF signaling. CNP had no effect on the STAT-1 pathway of FGF signaling that mediates the decreased proliferation and the delayed differentiation of achondroplastic chondrocytes. These results demonstrate that activation of the CNP-GC-B system in endochondral bone formation constitutes a new therapeutic strategy for human achondroplasia.     

Role of the essential thiol group in the thiol-activated cytolysin from Clostridium perfringens

A hemolysin, 0-toxin, produced by Clostridium perfringens has one cysteinyl residue in the free thiol form which is essential for its hemolytic activity. The cysteinyl residue was shown to be located at a position about 5 kDa from the C terminus of the molecule by the method of cysteine-specific chemical cleavage. Modification of the residue with a thiol-blocking agent, 5,5'-dithiobis(2-nitrobenzoic acid), reduced the binding affinity of the toxin to sheep erythrocytes to 1/100 that of intact toxin, resulting in a failure of binding at low cell concentrations (0.5%). Thus the failure of hemolysis at low cell concentrations is primarily ascribed to a decreased affinity of the toxin for erythrocytes. Effects of the modification on the lytic processes were examined using high cell concentrations where considerable amounts of modified toxin bound to the cells. The modified toxin hemolyzes erythrocytes once it binds to them; however, the efficiency of hemolysis is reduced by the modification. These, and additional results indicating that modification alters the sensitivity of toxin molecules to protease digestion, show that thiol-modification inactivates the toxin by affecting both binding and the subsequent lytic processes, probably through a conformational change introduced in the toxin molecules.     

Enzymatic conversion of norsolorinic acid to averufin in aflatoxin biosynthesis

5'-Hydroxyaverantin (HAVN) was isolated from a mold, Emericella heterothallica IFO 30842. Aspergillus parasiticus NIAH-26, a UV-irradiated mutant of A. parasiticus SYS-4, produced neither aflatoxins nor precursors in yeast extract-sucrose (YES) medium. When the postmicrosome (cytosol) fraction of NIAH-26, which had been prepared from the culture in YES medium, was incubated with norsolorinic acid (NA) in the presence of NADH or NADPH, averantin (AVN) was produced. The reverse reaction from AVN to NA was promoted by the addition of NAD or NADP (dehydrogenase reaction). When the microsome fraction of NIAH-26 was incubated with AVN, HAVN was produced in the presence of NADPH (monooxygenase reaction). HAVN was, furthermore, oxidized to averufin (AVR) by the cytosol fraction of NIAH-26 in the presence of NAD or NADP (dehydrogenase reaction). In the feeding experiments with A. parasiticus NIAH-26, aflatoxins were produced from AVN, HAVN, NA, and AVR but not from averufanin or averythrin. These results indicate that the reaction sequence NA in equilibrium AVN----HAVN----AVR is involved in the biosynthetic pathway of aflatoxins. The enzyme activities described here were dependent on the culture medium, and no enzyme activities were observed in the nonaflatoxigenic strain A. oryzae SYS-2 (IFO 4251).     

粘质沙雷氏菌AS-1中SpnR功能及卤化呋喃对其群体感应的抑制

通过分泌和感知一系列信号分子,细菌能够根据自身菌体密度的变化调控基因的表达,从而控制一系列重要的表现型,包括毒力因子的产生,生物膜的形成以及菌体发光等.这种广泛存在的信号机制被称为群体感应.在沙雷氏菌种中已经发现了多套群体感应机制.粘质沙雷氏菌AS-1从土壤中分离,其中含有LuxI/LuxR的同类蛋白,被称为SpnI/SpnR.粘质沙雷氏菌AS-1合成AHLs分子N-hexanoy1-L-homoserinelactone(C6-HSL)和N-(3.oxohexanoyl)-L-homoserine lactone(3-oxo-C6-HSL)作为其信号分子,通过群体感应感知菌体密度来控制基因的表达.通过基因替代的方法制得了spnR基因破坏的变异株,命名为粘质沙雷氏菌AS-1R.对粘质沙雷氏菌AS-1R的研究表明SpnR蛋白消极的调控沙雷氏菌红色色素的产生,运动性以及生物膜的形成等一系列由群体感应控制的性状:另一方面,作为一种天然的群体感应抑制剂,卤化呋喃能够有效的抑制粘质沙雷氏菌AS-1的群体感应,但并不干扰AHL-SpnR的相互作用.为运用粘质沙雷氏菌群体感应调节抑制其致病性提供了方法和依据,同时也为卤化呋喃对群体感应抑制机理的研究提供了新的思路.     

Microvascular and interstitial PO(2) measurements in rat skeletal muscle by phosphorescence quenching.

To clarify the transport of O(2) across the microvessels in skeletal muscle, we designed an intravital laser microscope that utilizes a phosphorescence quenching technique to determine both the microvascular and tissue PO(2). After we injected the phosphorescent probe into systemic blood, phosphorescence excited by a N(2)-dye pulse laser was detected with a photomultiplier over a 10 microm in diameter area. In vitro and in vivo calibrations confirmed that the present method is accurate for PO(2) measurements in the range of 7-90 Torr (r = 0.958) and has a rapid response time. This method was then used to measure the PO(2) of microvessels with different diameters (40-130 microm) and of interstitial spaces in rat cremaster muscle. These measurements showed a significant drop in PO(2) in the arterioles after branching (from 74.6 to 46.6 Torr) and the presence of a large PO(2) gradient at the blood-tissue interface of arterioles (15-20 Torr). These findings suggest that capillaries are not the sole source of oxygen supply to surrounding tissue.     

Phytosulfokine stimulates somatic embryogenesis in Cryptomeria japonica

Phytosulfokine (PSK), which has been identified as a plant growth factor, had a dramatic stimulatory effect on the formation of somatic embryos of sugi (Cryptomeria japonica) in the presence of polyethylene glycol. The resultant somatic embryos germinated with synchronous sprouting of cotyledons, hypocotyls and roots, and most of the seedlings grew normally. A cDNA clone for the precursor to the PSK peptide of C. japonica was identified in an expressed sequence tags database. Our results support the existence of a PSK signaling pathway in C. japonica.