Studies on interspecies competition between the azuki bean weevil and the southern cowpea weevil

Experimental studies was made on the interspecies competition between the azuki bean weevil, Callosobruchus chinensis and the southern cowpea weevil, C. macultatus. And the following results were obtained.

1. The reverse result of competition between the two species observed under the air-tight condition and the air-free one. That is, the population of the azuki bean weevils destroyedby that of the southern cowpea weevils under the former condition and vice versa under the latter. It is thought that such a reversal is due to the difference of sensibility of each species to the air-tight condition.
2. Under the air-free condition, change of the time interval of food-supplying had no effect on the result of competition within the limits of this experiment. The population of the southern cowpea weevils was always destroyed by that of the azuki bean weevils. But there was a certain degree of correlation between the duration of co-existence and the interval of food-supplying.
3. From the results, it is possible to say that by changing the degree of air-tightness, or the time interval of food-supplying, the co-existence period and the final result of competition can be changed.
4. The different mechanism of competition between two species in the present experiment from the experiments ofUtida (1952) andYoshida (1957) were discussed.
5. A difference in the mechanism of interspecies competition and intraspecies competition was expected from the level of total population numbers of two species and the individual weight of C. chinensis.

     

Roles of epidermal growth factor (EGF) in the growth and differentiation of human endometrium

Endometrial tissues undergo drastic changes during menstrual cycle. After menstruation, they proliferate and differentiate into cells with secretory activity in the preparation for egg implantation. Although sex steroids play an important role in the development of endometrial tissues, sequential events occurring in the endometrium can not be fully explained by the direct actions of sex steroids. In this study, we offer evidences that EGF is released from endometrial cells and they possess the receptor for EGF. These findings prompted us to explore the biological roles of EGF in endometrial tissues. Here we clearly demonstrate that EGF is involved in the proliferation of endometrial cells. Moreover, EGF is found to enhance both glycogenesis and glycogenolysis, thus increasing the supply of glucose for blastocysts. We further set forth that EGF augments the capacity of progestin receptor and release of prostaglandins in endometrial cells. In summary, this study emphasizes that EGF may participate in the development of human endometrial tissues in concert with sex steroids, thus contributing to the acquisition of receptivity of eggs in the endometrium.     

Alterations in migrating cranial neural crest cells in embryos of mice fed retinoic acid

Alterations in migrating neural crest cells induced by all-trans retinoic acid (RA) were studied morphologically and immunohistochemically in the cranial portion of 8-day-old mouse embryos which were derived from dams given 60, 40 or 0 mg kg of RA and killed 2 to 8 h later. Additionally, the embryos exposed to 4 mg/kg of actinomycin D (AD) on day 8 of gestation for 5 h were examined similarly. Light microscopy revealed that RA was cytotoxic and caused the appearance of pleomorphic nuclei, extra-large nucleoli and cytoplasmic budding which replaced lamellipodia and spike-like projections. Electron microscopy revealed pleomorphic nuclei containing nucleoli with major granular portions frequently surrounded with heterochromatin, monosomes, and phagosomes. A monosomal distribution pattern was different from that seen in the neural crest cells exposed to AD. The latter showed incomplete polyribosomal dispersion with fewer nucleolar components. Fewer neural crest cells with choline acetyltransferase-like immunoreactivity were detected in RA- and AD-exposed embryos than in the controls. These findings suggest that excess RA inhibits acetylcholine synthesis of the migrating neural crest cells, in a manner different from AD, and that it enhances phagocytosis. These phenomena modify the characteristics of neural crest cells resulting in craniofacial malformations.     

Location in Muscarinic Acetylcholine Receptors of Sites for [3H]Propylbenzilcholine Mustard Binding and for Phosphorylation with Protein Kinase C

Muscarinic acetylcholine receptors purified from porcine cerebra or atria were covalently labeled with [3H]propylbenzilylcholine mustard ([3H]PrBCM), and then the labeled receptors were subjected to limited hydrolysis with trypsin, V8 protease, and lysyl endopeptidase, followed by analysis involving sodium dodecyl sulfate-polyacrylamide gel electrophoresis, fluorography, autoradiography, or immunostaining. The labeled peptides were located on the basis of their reactivity with antibodies raised against three synthetic peptides with partial sequences of the m1 or m2 receptor, and of their sensitivity to endoglycosidase F, which was taken as evidence that they contain glycosylation sites near the N terminus. The [3H]PrBCM-binding site in both cerebral and atrial receptors was found to be located between the N terminus and the second intracellular loop, because the size of the smallest deglycosylated peptide that contained both the [3H]PrBCM-binding and glycosylation sites was approximately 16 kDa. Cerebral receptors were 32P-phosphorylated with protein kinase C, and the major phosphorylation sites in cerebral muscarinic receptors were found to be located in a C-terminal segment including a part of the third intracellular loop, because a 32P-labeled peptide of 12-14 kDa reacted with anti-(m1 C-terminal peptide) antiserum. The presence of an intramolecular disulfide bond, probably between Cys 98 and Cys 178 in the first and second extracellular loops, respectively, was suggested by the finding that a peptide of approximately 17 kDa containing the [3H]PrBCM-binding site, but not the glycosylation sites, was partly converted to a peptide of approximately 12 kDa on treatment with beta-mercaptoethanol.     

Purification and characterization of a peptide C-terminal alpha-amidating enzyme from porcine atrium

In many peptide hormones and neuropeptides, the carboxy-terminal alpha-amide structure is essential in eliciting biological activity. Here we report the purification and characterization of an alpha-amidating enzyme from porcine atrium, in which a high concentration of alpha-amidating activity was detected. The enzyme was purified to homogeneity from the membrane fraction of porcine atria by five steps of chromatography, including an affinity chromatography using a Sepharose column coupled with a substrate, Tyr-Phe-Gly. The purified enzyme was found to be composed of a single polypeptide chain with an apparent molecular weight of 92,000. This enzyme converted several synthetic peptides with C-terminal glycine to the corresponding des-glycine peptide alpha-amides.     

Insertion of a signal peptide-derived hydrophobic segment into the mature domain of OmpC, an outer membrane protein, does not interfere with the export of the following polypeptide chain across the cytoplasmic membrane of E. coli

The effects of a hydrophobic peptide segment inserted into the amino-terminal region of the mature domain of OmpC, an outer membrane protein, on its translocation across the cytoplasmic membrane was studied. Both the intact OmpC and central domain-deleted OmpC were examined. The hydrophobic segment was derived from the signal peptide of OmpF. Secretory translocation across the cytoplasmic membrane was examined by means of proteinase K treatment. Four monoclonal antibodies that recognize different regions of OmpC were used to characterize proteinase K-resistant fragments. Insertion of the hydrophobic segment did not appreciably prevent the translocation of these proteins across the cytoplasmic membrane, larger parts of them being found as mature forms, which were mostly localized outside the cytoplasmic membrane. Circumstantial evidence supports the view, on the other hand, that the inserted hydrophobic domain was retained in the cytoplasmic membrane. It is concluded, therefore, that the hydrophobic segment, although it is not exported across the cytoplasmic membrane, does not prevent the secretion of the following polypeptide chain. The secretion was dependent on the amino-terminal signal peptide. Insertion of positive charges immediately after the hydrophobic segment resulted in suppression of the translocation. Based on these results possible mechanisms by which the secretion of the polypeptide chain after the hydrophobic segment are discussed.     

Pure human inactive renin. Evidence that native inactive renin is prorenin

To clarify contradicting observations on the identity of inactive renin and prorenin, inactive renin was completely purified from native human chorion laeve and the culture medium of human chorion cells. A 720,000-fold purification with 14% recovery was achieved from chorion laeve in 6 steps, including immunoaffinity chromatography on a monoclonal antibody to human renin coupled to Protein A-Sepharose CL-4B. A 3,100-fold purification with 40% recovery was achieved from chorion culture medium in 4 steps, including immunoaffinity chromatography. Inactive renin purified from the two different sources migrated as a single protein band with the same molecular weight of 47,000 on sodium dodecyl sulfate-polyacrylamide gel electrophoresis and consisted of multiple components that could be resolved by isoelectric focusing. Both had the same pI values which shifted downward upon activation by trypsin; however, relative peak heights were different between the two preparations. The purified inactive renin from chorion laeve was completely inactive and did not bind to pepstatin-aminohexyl-Sepharose; however, that from chorion culture medium was partially active and completely bound to the pepstatin gel, indicating that each molecule is partially activated. Trypsin-activated inactive renins from both sources were identical with human renal renin in terms of pH optimum and Km. Specific activities of trypsin-activated inactive renin from chorion laeve and chorion culture medium were 529 Goldblatt units/mg of protein and 449 Goldblatt units/mg of protein, respectively. Amino acid sequence analysis of both of the purified inactive renin preparations demonstrated a leucine residue at the amino terminus. The sequence of 11 additional amino acids was identical in both and agreed with that predicted from the base sequence of the renin gene. These findings indicate that preprorenin is converted to prorenin following removal of a 23-amino acid signal peptide and that the native inactive renin, whose amino acid sequence commences with Leu-Pro-Thr..., is prorenin.     

Determination of monosaccharides and sugar alcohols in tissues from diabetic rats by high-performance liquid chromatography with pulsed amperometric detection.

A sensitive and simple high-performance liquid chromatographic method has been developed to determine the concentration of monosaccharides and sugar alcohols in animal tissues. Five neutral monosaccharides (D-glucose, D-galactose, D-mannose, D-fructose, and D-ribose) and three neutral sugar alcohols (myo-inositol, glycerol, and D-sorbitol) predominate in the renal cortices and sciatic nerves of rats. These monosaccharides and sugar alcohols were extracted with distilled water, purified by deproteinization with ethanol, a Sep-Pak C18 cartridge, and columns of Dowex 50W-X8 and Amberlite CG-400, then separated on Ca2+ and Pb2+ cation-exchange columns, eluted with deionized distilled water at 80 degrees C, and detected using integrated pulsed amperometry. About 10 pmol of each sugar was detectable with a signal-to-noise ratio of 10:1. D-Glucose, D-fructose, D-sorbitol, and D-mannose were higher in both the renal and sciatic tissues of diabetic rats than in those of normal animals. D-Ribose and glycerol were higher in the renal cortex of diabetic animals.     

Growth of the malignant and nonmalignant human squamous cells in a protein-free defined medium

Summary A novel protein-free synthetic medium has been developed for the culture of human squamous cell carcinoma cells. This medium, designated PF86-1, supports the serial subcultivation of six out of nine human squamous cell carcinoma cell lines in a protein-free, chemically defined condition without the adapting culture from serum-containing conditions. These cell lines growing in PF86-1 exhibited nearly equal potency to grow in massive culture without noticeable changes in morphology but presented a significantly decreased level of colony forming efficiency when compared with the cells cultured in serum-containing media, suggesting the implication of some autocrine mechanism. Interestingly, this medium supported the growth of normal human squamous cells of oral mucosa and skin for more than 2 mo. in the primary explant culture in spite of high levels of calcium ion concentration, where the overgrowth of fibroblasts as contaminant was not observed. These results suggest that PF86-1 supports the growth of cells derived from epidermal tissues selectively and provides the same defined condition for growth of malignant and nonmalignant human squamous cells. It seems, therefore, that PF86-1 allows investigations on the products of squamous cell carcinoma cells or on the differences of growth mechanisms between normal and neoplastic human squamous cells.