1. Sodium fluorescein and Evans Blue, commonly used tracers in the study of blood-brain barrier disturbances, revealed considerable differences in their respective protein binding capacity in the plasma, passage through the barrier and in the rate of their elimination from the brain parenchyma. 2. In the plasma a considerable portion of the sodium fluorescein remains free and behaves like a micromolecular barrier tracer. On the other hand, almost complete binding of the Evans Blue to albumin confers to it properties of a protein tracer. 3. Following the extravasation of the tracers, the sodium fluorescein is relatively soon eliminated, whereas Evans Blue remains in the cellular elements of the brain parenchyma for a considerable time, although the protein moiety of the tracer is removed much sooner from the cytoplasm of glial cells, presumably by the lysosomal digestion.
Rescue of the ribosomes from dead-end translation complexes, such as those on truncated (non-stop) mRNA, is essential for the cell. Whereas bacteria use trans-translation for ribosome rescue, some Gram-negative species possess alternative and release factor (RF)-dependent rescue factors, which enable an RF to catalyze stop-codon-independent polypeptide release. We now discover that the Gram-positive Bacillus subtilis has an evolutionarily distinct ribosome rescue factor named BrfA. Genetic analysis shows that B. subtilis requires the function of either trans-translation or BrfA for growth, even in the absence of proteotoxic stresses. Biochemical and cryo-electron microscopy (cryo-EM) characterization demonstrates that BrfA binds to non-stop stalled ribosomes, recruits homologous RF2, but not RF1, and induces its transition into an open active conformation. Although BrfA is distinct from E. coli ArfA, they use convergent strategies in terms of mode of action and expression regulation, indicating that many bacteria may have evolved as yet unidentified ribosome rescue systems.
Regulatory nascent peptides participate in the regulation of cellular functions by the mechanisms involving regulated translation arrest. A class of them in bacteria, called monitoring substrates, feedback-regulates the expression of a specific component of protein localization machinery. Three monitoring substrates, SecM, MifM and VemP have previously been identified. Here, we attempt at identifying additional arrest peptides in bacteria. Our bioinformatic searches over more than 400 bacterial genomic sequences for proteins that have the common characteristic features shared by the known monitoring substrates and subsequent in vitro and in vivo characterization of the highlighted sequences allowed the identification of three arrest peptides termed ApcA, ApdA and ApdP. ApcA and ApdA homologs are conserved among a subset of actinobacteria, whereas ApdP has homologs in a subset of α-proteobacteria. We demonstrate that these arrest peptides, in their ribosome-tethered nascent states, inhibit peptidyl transfer. The elongation arrest occurs at a specific codon near the 3′ end of the coding region, in a manner depending on the amino acid sequence of the nascent chain. Interestingly, the arrest sequences of ApcA, ApdA and ApdP share a sequence R-A-P-G/P that is essential for the elongation arrest.
SUMMARYThe incidence of cerebral lesions in stroke-prone spontaneously hypertensive rats appears to depend on the severity of the hypertension and nutritional factors. Comparison of American and Japanese commercial rat diets revealed a much higher incidence of stroke in rats receiving the Japanese diet (88% vs 30% by 9 months of age). Analyses of the diets indicate that perhaps the most important difference in the two diets is the protein content. Based on complete amino acid analyses of the protein in these diets, it appears that the American diet contains about 22% protein as compared to about 15% for the Japanese diet. Minor differences in vitamin and mineral contents are not remarkable. Comparison of the findings in this experimental rat model with epidemiologic studies suggest that nutritional factors may also play a role in the incidence of stroke in humans. INCREASED blood pressure appears to be one of the primary risk factors related to the incidence of hemorrhagic stroke in humans.' It was of interest therefore when Yamori et al, 2 reported the development of a rat strain that had very high blood pressure and a high incidence of stroke. This strain was derived from the spontaneously hypertensive rats (SHR) developed by Okamoto and Aoki 3 and exhibited an 80% to 90% incidence of cerebral lesions during the first year of life. This stroke-prone substrain is designated SHRSP and uniformly develops arterial blood pressure of over 200 mm Hg by 6 months of age. In 1975, this substrain was established in the breeding colonies of the National Institutes of Health (NIH) and used for studies on the pathogenesis of hypertension in this genetic model. It has subsequently become apparent to us that the incidence of cerebral lesions in the SHRSP in our laboratory was dramatically lower than that reported in Japan. The mean blood pressures observed in From the Section on Biochemical Pharmacology, HypertensionEndocrine Branch, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, Maryland (Dr. Lovenberg); the Department of Pathology, Shimane Medical University, Izumo, Japan (Drs. Yamori, Horie, Fujiwara, Nara, and Tanase).Present address for Dr. the NIH-SHRSP were significantly higher than in the SHR and were similar to the values reported for SHRSP in Japan. These observations suggested that perhaps environmental factors interacted with the genetic traits for hypertension to account for the high incidence of stroke in this strain of rats in Japan. We therefore undertook experiments to determine if the different incidence of stroke could be related to the diet. Materials and MethodsMale SHRSP rats were obtained from the Animal Production section of the NIH at 5 weeks of age and were assigned randomly to Groups 1 and 2 that were to receive either the Japanese rat diet (Funahashi-SP diet, Funahashi Farm, Funahashi City, Chiba Prefecture, Japan) or the NIH open formula diet (Ralston-Purina, Inc., St. Louis, Missouri). These groups were fed ad libitum and received tap water. Starting at 6 ...
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