Formation of nodules on non-nodulating soybean T201 after treatment with 2,4-dichlorophenoxyacetate

The formation of effective root nodules on a non-nodulating line (T201) of soybean (Glycine max (L.) Merr.,) was induced by a treatment with 2,4-dichlorophenoxyacetate (2,4-D). The induced nodules, inoculated with mixed Bradyrhizobium japonicum strains A1017 and IRj2101, had a normal internal structure, red in colour and the cells being filled with bacteroids. Externally, the induced nodules were of unusual shape, being paired or gourd-like in form and were attached to thickened roots. The nodules were capable of acetylene reduction (3.1–3.5 moles g^-1 fresh weight nodules h^-1), allowing the growth of plants with dark green leaves.     

Identity of soluble thiamin-binding protein with thiamin-repressible acid phosphatase in Saccharomyces cerevisiae

Two secretory glycoproteins of Saccharomyces cerevisiae, a soluble thiamin-binding protein and a thiamin-repressible acid phosphatase, were shown to be repressed to a similar extent by excess thiamin in the growth medium. Thiamin-repressible acid phosphatase was co-purified throughout the purification of the soluble thiamin-binding protein. Purified and deglycosylated soluble thiamin-binding proteins exhibited both thiamin-binding and acid phosphatase activity on non-denaturing polyacrylamide gel electrophoresis. Heat treatment of the purified soluble thiamin-binding protein caused a decrease in both activities with a similar inactivation profile. Furthermore, two thiamin-repressible acid phosphatase-defective mutants isolated had no and decreased soluble thiamin-binding activity, respectively. From the results, it was concluded that the soluble thiamin-binding protein is identical to the thiamin-repressible acid phosphatase in S. cerevisiae.     

Some properties of a Saccharomyces cerevisiae mutant resistant to 2-amino-4-methyl-5-beta-hydroxyethylthiazole

A mutant of Saccharomyces cerevisiae highly resistant to 2-amino-4-methyl-5-beta-hydroxyethylthiazole (2-aminohydroxyethylthiazole), an antimetabolite of 4-methyl-5-beta-hydroxyethylthiazole (hydroxyethylthiazole), has been isolated. Its resistance to 2-aminohydroxyethylthiazole was about 10(4) times that of the sensitive parent strain. The amount of thiamin synthesized in the cells of the resistant strain grown in minimal medium was less than half of that of the sensitive strain. The ability to synthesize thiamin from 2-methyl-4-amino-5-hydroxymethylpyrimidine (hydroxymethylpyrimidine) and hydroxyethylthiazole in the resistant strain was low compared with that of the sensitive strain. These results were found to be due to a deficiency of hydroxyethylthiazole kinase in the resistant strain: in sonic extracts of cells the enzyme activity was only 0.67% of that of the sensitive strain. Although the cells of the sensitive strain could accumulate exogenous hydroxyethylthiazole in the form of hydroxyethylthiazole monophosphate, no significant uptake of hydroxyethylthiazole by the cells of the resistant strain was observed. The possibilities that 2-aminohydroxyethylthiazole monophosphate may be the actual inhibitor of the growth of Saccharomyces cerevisiae, and that hydroxyethylthiazole may not be involved in the pathway of de novo synthesis of thiamin via hydroxyethylthiazole monophosphate, are discussed.     

On the oscillatory phenomenon in an oil/water interface

A simple theoretical model is presented for simulating the self-sustained oscillations of electric potential and pH at an oil/water interface appearing in a two-phase system composed of 2-nitropropane solution containing picrate acid and an aqueous solution of cetyltrimethylammonium bromide. In the present model, a well-known condition necessary for the occurrence of self-sustained oscillations, i.e., the presence of a positive feedback process far from equilibrium, is taken into account in a set of kinetic equations to describe simplified characters of the following two processes: (i) a cooperative formation of ion pair complexes at the interface, and (ii) supply of picrate anions and cetyltrimethylammonium cations to the interface accompanied by release of ion pair complexes to the organic phase. The numerical solutions of the present equations are shown to reproduce fairly well the characteristic properties of the oscillation of electric potential and pH such as wave forms and frequencies.     

Inactivation of rice bran thiamine-binding protein by N,N'-dicyclohexylcarbodiimide

The addition of a carboxyl-modifying reagent N,N'-dicyclohexylcarbodiimide (DCCD) to thiamine-binding protein isolated from rice bran resulted in a remarkable loss of its binding activity with [14C]thiamine. Thiamine and chloroethylthiamine substantially protected the protein against inactivation by DCCD, whereas thiamine phosphates did not. Another carboxyl reagent N-ethoxycarbonyl-2-ethoxy-1,2-dihydroquinoline (EEDQ) also inactivated rice bran thiamine-binding protein. Inactivation of the thiamine-binding protein was accompanied by covalent binding of DCCD to the protein as shown by the use of [14C]DCCD. The binding of [14C]DCCD to the thiamine-binding protein was specific, and significantly inhibited by the addition of thiamine. The loss of thiamine-binding activity was proportional to the specific binding of [14C]DCCD. For complete inactivation of the thiamine-binding activity, the binding of 2.46 mol of [14C]DCCD per mol of thiamine-binding protein was required. Furthermore, limited proteolysis of the binding protein by trypsin yielded two polypeptides with molecular weights of 35,000 (large polypeptide) and 12,500 (small polypeptide) which were separated by SDS-polyacrylamide gel electrophoresis. The binding sites of [14C]DCCD were found to be located on the large polypeptide. These results suggest that a specific carboxyl residue in the large polypeptide releasable from rice bran thiamine-binding protein by trypsin digestion when modified by DCCD is involved in the binding of thiamine.     

Purification and Heterogeneous Localization of Sarcophaga Lectin Receptor on the Surface of Imaginal Discs of Sarcophaga peregrina (Flesh Fly)

Previously, we reported autocrine involvement of Sarcophaga lectin in the development of Sarcophaga imaginal discs (Kawaguchi et al. , Dev. Biol. 144 , 86–93 (1991)). In this study, we purified Sarcophaga lectin binding protein from the membrane fraction of cultured embryonic cells of Sarcophaga to near homogeneity and raised a monoclonal antibody against it. Histochemical analysis using the monoclonal antibody revealed that this binding protein is distributed heterogeneously on the surface of leg imaginal discs. This binding protein was especially clearly localized in the central region of the basal side of leg discs which forms the junction between the leg and body, suggesting the participation of Sarcophaga lectin in morphogenesis of the basal region of the developing leg.     

Effects of physical exercise on the elasticity and elastic components of the rat aorta

To evaluate the effects of exercise on aortic wall elasticity and elastic components, young male rats underwent various exercise regimes for 16 weeks. In the exercised rats, the aortic incremental elastic modulus decreased significantly when under physiological strain. The aortic content of elastin increased significantly and the calcium content of elastin decreased significantly in the exercised group. The accumulated data from the exercised and sedentary groups revealed that the elastin calcium content was related positively to the incremental elastic modulus. We concluded that physical exercise from an early age decreases the calcium deposit in aortic wall elastin and that this effect probably produced in the exercised rats a distensible aorta.     

Possible Involvement of Phosphorylation of Occludin in Tight Junction Formation

Occludin is an integral membrane protein localizing at tight junctions in epithelial and endothelial cells. Occludin from confluent culture MDCK I cells resolved as several (>10) bands between 62 and 82 kD in SDS-PAGE, of which two or three bands of the lowest M_r were predominant. Among these bands, the lower predominant bands were essentially extracted with 1% NP-40, whereas the other higher M_r bands were selectively recovered in the NP-40–insoluble fraction. Alkaline phosphatase treatment converged these bands of occludin both in NP-40–soluble and -insoluble fractions into the lowest M_r band, and phosphoamino acid analyses identified phosphoserine (and phosphothreonine weakly) in the higher M_r bands of occludin. These findings indicated that phosphorylation causes an upward shift of occludin bands and that highly phosphorylated occludin resists NP-40 extraction. When cells were grown in low Ca medium, almost all occludin was NP-40 soluble. Switching from low to normal Ca medium increased the amount of NP-40–insoluble occludin within 10 min, followed by gradual upward shift of bands. This insolubilization and the band shift correlated temporally with tight junction formation detected by immunofluorescence microscopy. Furthermore, we found that the anti–chicken occludin mAb, Oc-3, did not recognize the predominant lower M_r bands of occludin (non- or less phosphorylated form) but was specific to the higher M_r bands (phosphorylated form) on immunoblotting. Immunofluorescence microscopy revealed that this mAb mainly stained the tight junction proper of intestinal epithelial cells, whereas other anti-occludin mAbs, which can recognize the predominant lower M_r bands, labeled their basolateral membranes (and the cytoplasm) as well as tight junctions. Therefore, we conclude that non- or less phosphorylated occludin is distributed on the basolateral membranes and that highly phosphorylated occludin is selectively concentrated at tight juctions as the NP-40–insoluble form. These findings suggest that the phosphorylation of occludin is a key step in tight junction assembly.