High level of divergence of male-reproductive-tract proteins, between Drosophila melanogaster and its sibling species, D. simulans

We compared male-reproductive-tract polypeptides of Drosophila melanogaster and D. simulans by using two-dimensional gel electrophoresis. Approximately 64% of male-reproductive-tract polypeptides were identical between two randomly chosen isofemale lines from these two species, compared with 83% identity for third-instar imaginal wing-disc polypeptides. Qualitatively similar differences were found between reproductive tracts and imaginal discs when D. sechellia was compared with D. melanogaster and with D. simulans. When genic polymorphism was taken into account, approximately 10% of male- reproductive-tract polypeptides were apparently fixed for different alleles between D. melanogaster and D. simulans; this proportion is the same as that found for soluble enzymes by one-dimensional gel electrophoresis. Strikingly, approximately 20% of male-reproductive- tract polypeptides of either D. melanogaster or D. simulans had no detectable homologue in the other species. We propose that proteins of the Drosophila male reproductive tract may have diverged more extensively between species than have other types of proteins and that much of this divergence may involve large changes in levels of polypeptide expression.      

Some properties of human and bovine brain cathepsin B

Cathespin B has been purified 750-fold to apparent homogeneity from human and bovine brain cortex using ammonium sulfate fractionation (30–70%), chromatography on Sephadex G-100, CM-Sephadex C-50, and concanavalin A-Sepharose. Enzyme was assayed fluorometrically at pH 4.0 with pyridoxyl-hemoglobin in the presence of 1 mM DTT and 1 mM EDTA. Properties of the enzyme from the two sources proved to be similar. On disc PAGE the purified preparation produced two bands associated with proteinase activity that are due to existence of two multiple forms of brain cathepsin B with pI 6.1 and 6.8. The enzyme is completely inactivated by thiol-blocking reagents, leupeptin, E-64, and demands thiol compounds for its ultimate activity. Z-Phe-Ala-CHN₂ is a potent inhibitor of the enzyme (K _2nd=1280 M⁻¹s⁻¹) in contrast to Z-Phe-Phe-CHN₂ (K _2nd=264 M⁻¹s⁻¹). pH optimum in the reaction of hydrolysis of Pxy-Hb is 4.0–6.0,K _M(app.) =10⁻⁵ M. Cathepsin B splits azocasein: pH optimum 5.0–6.0,K _M(app.)=2.2·10⁻⁵ M, but inclusion of urea in the incubation medium depresses the azocaseinolytic activity of the enzyme 1.5-fold. It does not split Lys-NNap, Arg-NMec and is not inhibited by bestatin. The specific activity of brain cathepsin B with Z-Arg-Arg-NNapOMe at pH 6.0 is 10-fold higher than with Bz-Arg-NNap, Z-Gly-Gly-Arg-NNap is a poor substrate. With Z-Arg-Arg-NMec and Bz-Phe-Val-Arg-NMec the specific acitivity is 80 and 35%, respectively of that with Z-Phe-Arg-NMec. Special Issue dedicated to Dr. Eugene Kreps.     

Osmosensitivity of the 2H+−K+-exchange and the H+-ATPase complex F0F1 in anaerobically grownEscherichia coli

Escherichia coli grown anaerobically for osmotic studies upon increased osmolarity in alkaline medium carried out H⁺–K⁺-exchange in two steps, the first of which was DCCD¹ sensitive and osmo-dependent and had the 2H⁺/K⁺ stoichiometry. H⁺-efflux in the presence of protonophore (CCCP) upon increase of osmolarity was shown to be high and inhibited by DCCD, whereas H⁺-efflux induced by a decrease of osmolarity was small and not inhibited by DCCD. The 2H⁺/K⁺-exchange was absent intrkA anduncA mutants. InuncB mutant 2H⁺/K⁺-exchange was not DCCD-and osmosensitive. Competition between DCCD and osmoshock on inhibition of 2H⁺/K⁺-exchange was found. Osmosensitivity of this exchange disappeared in spheroplasts. Osmosensitivity of both 2H⁺/K⁺-exchange and the F₀F₁ and osmoregulation of the F₀F₁ via F₀ and a periplasmic space are postulated.Abbreviations F₀F₁ H⁺-ATPase complex - F₀ H⁺-channel, proteolipid - F₁ H⁺-ATPase - Trk constitutive system for K⁺ uptake - PV periplasmic protein valve - DCCD N,N-dicyclohexylcarbodiimide - CCCP carbonylcyanide-m-chlorophenylhydrazone - _H or _K transmembrane electrochemical gradient for H⁺ or K⁺ respectively - membrane potential - upshock or downshock increase or decrease of medium osmolarity, respectively - CGSC E. coli Genetic Stock Center, Yale University, USA     

Microelectrode studies of the active Na transport pathway of frog skin

When the outer surface of short-circuited frog skin was penetrated with microelectrodes, stable negative potentials that averaged near -100 mV were recorded consistently, confirming the results of Nagel (W. Nagel. 1975. Abstracts of the 5th International Biophysics Congress, Copenhagen. P-147.). The appearance of these stable potentials, V(O), concurrent with the observations that (a) a high resistance outer barrier R(O) accounting for approximately 75 percent or more of the transcellular resistance of control skins had been penetrated and that (b) 10(-5) M amiloride and reduced [Na] outside caused the values of V(O) to increase towards means value near -130 mV while the values of percent R(O) increased to more than 90 percent. It was of relationships were the same as the values of E(1) observed in studies of the current-voltage relationships were the same as the values of E’(1) defined as the values of voltage at the inner barrier when the V(O) of the outer barrier was reduced to zero by voltage clamping of the skins. Accordingly, these data are interpreted to mean that the values of E(1), approximately 130 mV, represent the E(Na) of the sodium pump at the inner barrier. 2,4-DNP was observed to decrease the values of transepithelial voltage less than E(1) the V(O) was negative. These data can be interpreted with a simple electrical equivalent circuit of the active sodium transport pathway of the frog skin that includes the idea that the outer membrane behaves as an electrical rectifier for ion transport.     

Effects of hydrocortisone on the hypercalcemia and plasma levels of 13,14-dihydro-15-keto-prostaglandin E2 in mice bearing the HSDM1 fibrosarcoma

In mice bearing the prostaglandin-producing HSDM₁ fibrosarcoma, the plasma concentration of 13,14-dihydro-15-keto-PGE₂ was elevated before the development of hypercalcemia, and the magnitude of the rise was greater than that of PGE₂. When hydrocortisone, which inhibits synthesis of PGE₂ by HSDM₁ cells in culture, was administered to tumor-bearing mice, the steroid hormone prevented the rises in plasma PGE₂ metabolite and calcium concentrations. At the dose levels used, hydrocortisone did not inhibit the calcium-mobilizing action of parathyroid hormone $\text{in vivo}$ or the bone resorption-stimulating activity of $\text{PGE}{}_2\text{in vitro}$. These findings are consistent with our hypothesis that the hypercalcemic syndrome in HSDM₁ tumor-bearing mice is due to the secretion of PGE₂ by the tumor.     

Thyrotropin-releasing hormone inreases prolactin mRNA activity in the cytoplasm of GH-cells as measured by translation in a wheat germ cell-free system

A wheat germ embryo extract was used to translate cytoplasmic RNA isolated from rat pituitary tumor cells (GH-cells). This RNA directed the synthesis of a radioactive product which was precipitated with antiserum specific for rat prolactin. The molecular weight of this immunoprecipitated product was 24,500 as determined by electrophoresis in denaturing gels. Prolactin secreted by intact GH-cells had a molecular weight identical to standard pituitary prolactin, reported to be about 22,500. Our finding that a larger form of prolactin is made by the wheat germ system is similar to results recently described by Maurer, Stone and Gorski (J. Biol. Chem., in press). Thyrotropin-releasing hormone (TRH) stimulates prolactin synthesis in GH-cells, and cytoplasmic RNA isolated from cells treated with TRH directed the synthesis in wheat germ extracts of larger amounts of prolactin than RNA isolated from control cells. The increase in translatable cytoplasmic mRNA for prolactin corresponded to the increase in prolactin synthesis which suggests that the increase in prolactin synthesis in TRH-treated cells is a result of the accumulation of cytoplasmic mRNA for prolactin.     

Comparable Effects of Low-intensity Electromagnetic Irradiation at the Frequency of 51.8 and 53 GHz and Antibiotic Ceftazidime on Lactobacillus acidophilus Growth and Survival

The effects of low-intensity electromagnetic irradiation (EMI) with the frequencies of 51.8 and 53 GHz on Lactobacillus acidophilus growth and survival were revealed. These effects were compared with antibacterial effects of antibiotic ceftazidime. Decrease in bacterial growth rate by EMI was comparable with the inhibitory effect of ceftazidime (minimal inhibitory concentration—16 μM) and no enhanced action was observed with combined effects of EMI and the antibiotic. However, EMI-enhanced antibiotic inhibitory effect on bacterial survival. The kinetics of the bacterial suspension oxidation–reduction potential up to 24 h of the growth was changed by EMI and ceftazidime. The changes were more strongly expressed by combined effects of EMI and antibiotic especially up to 12 h. Moreover, EMI did not change overall energy (glucose)-dependent H⁺ efflux across the membrane but it increased N,N′-dicyclohexylcarbodiimide (DCCD)-inhibited H⁺ efflux. In contrast, this EMI in combination with ceftazidime decreased DCCD-sensitive H⁺ efflux. Low-intensity EMI had inhibitory effect on L. acidophilus bacterial growth and survival. The effect on bacterial survival was more significant in the combination with ceftazidime. The H⁺-translocating F ₀ F ₁-ATPase, for which DCCD is specific inhibitor, might be a target for EMI and ceftazidime. The revealed bactericide effects on L. acidophilus can be applied in biotechnology, food producing and safety technology.     

Area-Level Attributes and Esophageal Adenocarcinoma in Surveillance,Epidemiology and End Results Registries

Purpose

To examine the associations between area-level socioeconomic attributes and stage of esophageal adenocarcinoma diagnoses in 16 SEER cancer registries during 2000-2007.

Methods

Odds ratios (OR) and 95% confidence intervals (CI) were calculated using multivariable logistic regression models to assess the relationship between distant-stage esophageal adenocarcinoma and individual, census tract, and county-level attributes.

Results

Among cases with data on birthplace, no significant association was seen between reported birth within versus outside the United States and distant-stage cancer (adjusted OR=1.02, 95% CI: 0.85-1.22). Living in an area with a higher percentage of residents born outside the United States than the national average was associated with distant-stage esophageal adenocarcinoma; census tract level: >11.8%, (OR=1.10, 95% CI:1.01–1.19), county level: >11.8%, (OR=1.14, 95% CI:1.05-1.24). No association was observed between median household income and distant-stage cancer at either census tract or county levels.

Conclusion

The finding of greater odds of distant-stage esophageal adenocarcinoma among cases residing in SEER areas with higher proportion of non-U.S. Natives suggests local areas where esophageal cancer control efforts might be focused. Missing data at the individual level was a limitation of the present study. Furthermore, inconsistent associations with foreign birth at individual- versus area-levels cautions against using area-level attributes as proxies for case attributes.     

Oxidative and Reductive Routes of Glycerol and Glucose Fermentation by Escherichia coli Batch Cultures and Their Regulation by Oxidizing and Reducing Reagents at Different pHs

Glycerol and glucose fermentation redox routes by Escherichia coli and their regulation by oxidizing and reducing reagents were investigated at different pHs. Cell growth was followed by decrease of pH and redox potential (E _h ). During glycerol utilization at pH 7.5 ?pH, the difference between initial and end pH, was lower compared with glucose fermentation. After 8 h growth, during glycerol utilization E _h dropped down to negative values (?150 mV) but during glucose fermentation it was positive (+50 mV). In case of glycerol H₂ was evolved at the middle log phase while during glucose fermentation H₂ was produced during early log phase. Furthermore, upon glycerol utilization, oxidizer potassium ferricyanide (1 mM) inhibited both cell growth and H₂ formation. Reducing reagents dl-dithiothreitol (3 mM) and dithionite (1 mM) inhibited growth but stimulated H₂ production. The findings point out the importance of reductive conditions for glycerol fermentation and H₂ production by E. coli.