Modulation of cysteine biosynthesis in chloroplasts of transgenic tobacco overexpressing cysteine synthase [O-acetylserine(thiol)-lyase]

Cysteine synthase [O-acetyl-L-serine(thiol)-lyase, EC 4.2.99.8] (CSase), which is responsible for the terminal step of cysteine biosynthesis, catalyzes the formation of L-cysteine from O-acetyl-L-serine (OAS) and hydrogen sulfide. Three T-DNA vectors carrying a spinach (Spinacia oleracea) cytoplasmic CSase A cDNA (K. Saito, N. Miura, M. Yamazaki, H. Horano, I. Murakoshi [1992] Proc Natl Acad Sci USA 89: 8078-8082) were constructed as follows: pCSK3F, cDNA driven by the cauliflower mosaic virus (CaMV) 35S RNA promoter with a sense orientation; pCSK3R, cDNA driven by the CaMV 355 promoter with an antisense orientation; pCSK4F, cDNA fused with the sequence for chloroplast-targeting transit peptide of pea ribulose-1,5-biphosphate carboxylase small subunit driven by the CaMV 35S promoter with a sense orientation. These chimeric genes were transferred into tobacco (Nicotiana tabacum) with Agrobacterium-mediated transformation, and self-fertilized progeny were obtained. CSase activities in cell-free extracts of pCSK3F and pCSK4F transformants were 2- to 3-fold higher than those of control and pCSK3R plants. CSase activities in chloroplasts of pCSK4F transformants were severalfold higher than those of control and pCSK3F plants, indicating that the foreign CSase protein is transported and accumulated in a functionally active form in chloroplasts of pCSK4F plants. Isolated chloroplasts of a pCSK4F transformant had a more pronounced ability to form cysteine in response to addition of OAS and sulfur compounds than those of a control plant. In particular, feeding of OAS and sulfite resulted in enhanced cysteine formation, which required photoreduction of sulfite in chloroplasts. The enhanced cysteine formation in a pCSK4F plant responding to sulfite was also observed in leaf discs. In addition, these leaf discs were partially resistant to sulfite toxicity, possibly due to metabolic detoxification of sulfite by fixing into cysteine. These results suggested that overaccumulated foreign CSase in chloroplasts could modulate biosynthetic flow of cysteine in response to sulfur stress.     

Control aspects of the mitsubishi continuous process

The Mitsubishi process for the continuous smelting and converting of copper holds many advantages over conventional processes, where reactions must be conducted in numerous steps and melts must be tapped frequently from the furnaces. The furnaces operate like steady-state reactors with constant melt volume, composition, and temperature. Therefore, optimal control of the process is straightforward, with one operator controlling smelting and converting simultaneously. Recent improvements in temperature control by using newly developed sensors have extended furnace campaign life, and enhanced control over melt compositions has helped further stabilize operations. Applications of the environmentally clean smelting technology are increasing internationally. Moto Goto earned his Dr. Eng. in metallurgical engineering at Tokyo University in 1984. He is currently senior managing director at Mitsubishi Materials Corporation. Dr. Goto is also a member of TMS. Eiki Oshima earned his M.S. in metallurgical engineering at Tohoku University in 1968. He is currently general manager of the Naoshima smelter at Mitsubishi Materials Corporation. Mr. Oshima is also a member of TMS. Mineo Hayashi earned his M.S. in metallurgical engineering at Tohoku University in 1971. He is currently general manager of the process and technology department, International Copper Project Division, Mitsubishi Materials Corporation. Mr. Hayashi is also a member of TMS.     

Applications of electrostatic stretch-and-positioning of DNA

The authors have previously reported that the electrostatic orientation and the dielectrophoresis (DEP) of DNA occur under ≈1 MHz, >1×10⁶ V/m field, by which DNA strands are stretched straight along field lines and positioned onto electrode edges. This paper presents some application of this stretch-and-positioning method to genetic engineering. It is shown that the DNA size distribution, as well as the activities of nuclease, can be determined by the measurement of the apparent length of stretched DNA. Several methods are developed to immobilize stretched DNA onto a substrate, including: (1) immobilization onto a conducting substrate for observations with the scanning tunneling microscopy, (2) anchoring onto a substrate only at both ends of the DNA using special electrode configuration, and/or molecular binding between avidin and biotin. The DNA can be held without contact to the substrate in the latter method, so that it does not cause steric hindrances to the DNA-binding enzymes. A novel fluid integrated circuit (FIC) device is proposed in which stretched DNA is cut by laser beam for the successive sequencing. A method to obtain unidirectionally oriented DNA is developed. The spatial resolution, and the small number of molecules required, are the advantages of the assays and measurements using electrostatic DNA manipulations over conventional biochemical methods. It is hoped that the methods may open a way to a novel category of “molecular biochemistry with spatial resolution.”     

A cylindrical micro ultrasonic motor using PZT thin film deposited by single process hydrothermal method (phi2.4 mm, L=10 mm stator transducer)

A micro ultrasonic motor using PZT (lead zirconate titanate) thin film has been improved by a single process hydrothermal method. The hydrothermal method is a unique method for depositing PZT thin film in a solution. An earlier reported hydrothermal method consisted of two linked processes. Our new method, however, has only a single process. Hence, less distribution of chemical components of the PZT film contributes to a higher efficiency of the stator transducer. The piezoelectric factor d(31) was -30 pC/N for this new method, which is six times larger than that of the previous method. The output torque of the micro ultrasonic motor fabricated by the single process hydrothermal method was measured. The output torque was 7.0 muNm, and the maximum revolution speed was 880 rpm at 15 Vp-p driving voltage.     

Effects of amrinone on left ventricular function following open heart surgery--analysis with left ventricular pressure volume loops

The effects of Amrinone on cardiac function soon after extracorporeal circulation (ECC) were studied in 5 patients including mitral valvuloplasty, VSD closure, Fontan operation and coronary AV fistel closure. In all patients, left ventricular volume load decreased postoperatively. To evaluate the efficacy, we obtained left ventricular pressure-volume loops (P-V loop) before and after ECC and after intravenous administration of Amrinone (1 mg/kg) following ECC. P-V loops were produced by measuring left ventricular pressure using a Miller catheter which was retrogradely advanced from the ascending aorta into the left ventricle and by measuring left ventricular diameter to calculate left ventricular volume with Teichholtz' formula. Although no apparent difference of Emax was recognized before and after ECC, Emax increased from 3.2 +/- 2.5 mmHg/cm3 to 5.9 +/- 4.7 mmHg/cm3 after the administration of Amrinone. The left ventricular "systolic" pressure-volume area (PVA) which is the sum of stroke work (SW) and elastic potential energy decreased from 34.4 +/- 16.4 gm to 30.9 +/- 17.8 gm after Amrinone. No difference was also recognized in left ventricular end-diastolic pressure. Ejection fraction increased from 50 +/- 17.5% to 56.1 +/- 17.3%. These results suggested that Amrinone could improve the left ventricular function without prominent change in myocardial oxygen consumption immediately after open heart surgery.     

Hydrogen analysis of mineral samples at University of Tsukuba

This study reports the present status of our work on the hydrogen analysis of mineral and rock samples. The preparation of a standard material by means of ion implantation, a method of its calibration and the application of the method are described. The number of hydrogen atoms per unit volume in the standard material can be determined from the simultaneous observation of -particles, γ rays and –γ coincidence events for the ¹H(¹⁹F,γ) reaction at the 16.44 MeV resonance energy. The hydrogen content in a natural obsidian determined with the method mentioned above is in agreement with that obtained by FTIR. A heavy-ion microbeam system under construction, which consists of a Russian-type quadrupole magnet for beam focusing, a beam defining slit system and a γ-ray detector, is also described.     

Effect of the diaphragm of free-air ionisation chamber for X-ray air-kerma measurements

Free-air ionisation chambers are widely used at standards laboratories as primary standards for absolute measurements of air kerma in X-ray fields. The area of the diaphragm aperture of a free-air ionisation chambers is an important factor for absolute measurements because it defines the size of the X-ray beam incident on the free-air chamber. In this study, correction factors for the contribution of X rays transmitted through the diaphragm of a free-air ionisation chamber and those scattered from the surface of the diaphragm aperture are obtained by Monte Carlo simulation for two different sized free-air ionisation chambers and for various diaphragm aperture sizes, X-ray energies and source-to-chamber distances.