Arachidonic acid metabolism in rabbit renal cortex. Formation of two novel dihydroxyeicosatrienoic acids

[1-14C]Eicosatetraenoic (arachidonic) acid was incubated with a low speed (17,000 X g) rabbit renal cortical supernatant or with a cortical microsomal suspension fortified with NADPH for 15 min at 37 degrees C. The products which were less polar than prostaglandins on reversed phase high performance liquid chromatography were identified by gas chromatography-mass spectrometry. Both the fortified microsomes and the low speed supernatant formed significant amounts of two novel metabolites, 11,12-dihydroxy-5,8,14-eicosatrienoic acid and 14,15-dihydroxy-5,8,11-eicosatrienoic acid. Other identified products were 19- and 20-hydroxyeicosatetraenoic acid, 19-oxoeicosatetraenoic acid, and in the low speed supernatant, eicosatetraen-1,20-dioic acid. The metabolites were not formed in significant amounts by high speed cortical supernatant or by nonfortified cortical microsomes. Carbon monoxide inhibited formation of these compounds, indicating that they may be formed by the cytochrome P-450-linked renal monooxygenase systems.     

Calcium-dependent slow potassium conductance in rat skeletal myotubes

A prolonged hyperpolarizing afterpotential (amplitude 5–20 mV, half decay time about 400 msec at 25°C) follows the action potential in myotubes and myosacs cultured from rat skeletal muscle. This slow hyperpolarizing afterpotential (hap) is mediated by an increase in membrane K conductance, because its reversal potential follows the Nernst potential for K and is not affected by other ions. The conductance increase measured during the hap (up to four times the resting input conductance) correctly predicts the time course of the slow hap. The slow hap is Ca dependent. Its amplitude decreases when bath [Ca] is lowered, and both amplitude and duration increase when bath [Ca] is raised. The slow hap is blocked by intracellular injection of the calcium chelator, EGTA. It is inhibited by solutions containing 2–4 mM manganese or 1–5 mM barium, but is not blocked by 5–20 mM tetraethylammonium. Myotubes bathed in zero [Na], high [Ca] solutions show calcium action potentials, which are inhibited by 2–10 mM manganese, nickel or cobalt. Myotubes bathed in isotonic Ca salts (or in 2 mM Ca plus 5 mM caffeine) show long-lasting (up to 10 sec) spontaneous hyperpolarizations accompanied by prolonged contractions. These hyperpolarizations are associated with a large increase in input conductance, and they reverse in sign near the K equilibrium potential. They appear to reflect activation of the Ca-sensitive K conductance by Ca released from intracellular stores. The observation that spontaneous hyperpolarizations usually occur with no prior depolarization argues that at least a portion of the slow, Ca-sensitive K conductance system can be activated by internal Ca alone, with no requirement for plasma membrane depolarization. Cultured myotubes also have a faster K conductance system, which is inhibited by 5–20 mM tetraethylammonium or 1–5 mM barium, and is not dependent on Ca for its activation.     

Effects of red light on the growth of intact wheat and barley coleoptiles

The final lengths of intact dark-grown coleoptiles vary with species and cultivar. The growth distribution pattern in the apical 25-mm growing zone and the absolute amount of growth in each zone depend on the age and species of the coleoptile. A comparative study of several cultivars of wheat, Triticum vulgare, and barley, Hordeum vulgare, indicates that the growth distribution pattern in 30- to 38-mm coleoptiles varies with the species and cultivar. In barley, there are two patterns of growth distribution among the several cultivars, whereas in wheat, all cultivars exhibit a common zonal growth pattern. The total growth of coleoptiles, initially 30 to 38 mm in length, during a 24-hour dark incubation period is the same in dark-grown coleoptiles as in those irradiated with 3 minutes of red (660 nm) light prior to the incubation period. The growth distribution pattern in the growing zone of this 30- to 38-mm coleoptile is, however, altered by red light. Growth of the apical 5-mm zone is stimulated by red light and the zonal growth 5 to 10 mm below the apex is only slightly affected, whereas growth in the zones 10 to 15 to 20, and 20 to 25 mm below the apex is inhibited. This growth distribution pattern in irradiated coleoptiles changes as the coleoptile increases in length. The response of a zone following exposure to red light is dependent upon the age of the seedlings irradiated. The over-all effect of red light on growth of the intact coleoptile varies with the length of the coleoptile. In young seedling 20 to 29 mm in length, the cells of the coleoptile can compensate for the effects of red light, with the over-all growth of the dark-grown and irradiated coleoptile about the same. As the seedling grows older, the cells of the coleoptile can no longer make up for the effects of red light, and the over-all effect changes from compensation to pronounced inhibition.     

Electron microscopy holdings of the Protein Data Bank: the impact of the resolution revolution,new validation tools,and implications for the future

As a discipline, structural biology has been transformed by the three-dimensional electron microscopy (3DEM) “Resolution Revolution” made possible by convergence of robust cryo-preservation of vitrified biological materials, sample handling systems, and measurement stages operating a liquid nitrogen temperature, improvements in electron optics that preserve phase information at the atomic level, direct electron detectors (DEDs), high-speed computing with graphics processing units, and rapid advances in data acquisition and processing software. 3DEM structure information (atomic coordinates and related metadata) are archived in the open-access Protein Data Bank (PDB), which currently holds more than 11,000 3DEM structures of proteins and nucleic acids, and their complexes with one another and small-molecule ligands (~ 6% of the archive). Underlying experimental data (3DEM density maps and related metadata) are stored in the Electron Microscopy Data Bank (EMDB), which currently holds more than 21,000 3DEM density maps. After describing the history of the PDB and the Worldwide Protein Data Bank (wwPDB) partnership, which jointly manages both the PDB and EMDB archives, this review examines the origins of the resolution revolution and analyzes its impact on structural biology viewed through the lens of PDB holdings. Six areas of focus exemplifying the impact of 3DEM across the biosciences are discussed in detail (icosahedral viruses, ribosomes, integral membrane proteins, SARS-CoV-2 spike proteins, cryogenic electron tomography, and integrative structure determination combining 3DEM with complementary biophysical measurement techniques), followed by a review of 3DEM structure validation by the wwPDB that underscores the importance of community engagement.     

Uncoupling of Bacterioplankton and Phytoplankton Production in Fresh Waters Is Affected by Inorganic Nutrient Limitation

Pelagic bacterial production is often positively correlated, or coupled, with primary production through utilization of autotrophically produced dissolved organic carbon. Recent studies indicate that inorganic N or P can directly limit both bacterial and phytoplanktonic growth. Our mesocosm experiments, with whole communities from mesotrophic Calder Lake, test whether this apparent bacterial-algal coupling may be the result of independent responses to limiting inorganic nutrients. In systems without N additions, numbers of bacteria but not phytoplankton increased 2- to 2.5-fold in response to P fertilization (0 to 2.0 μmol of P per liter); this resulted in uncoupled production patterns. In systems supplemented with 10 μmol of NH₄NO₃ per liter, P addition resulted in up to threefold increases in bacteria and two- to fivefold increases in total phytoplankton biomass (close coupling). P limitation of pelagic bacteria occurred independently of phytoplankton dynamics, and regressions between bacterial abundance and phytoplankton chlorophyll a were nonsignificant in all systems without added N. We describe a useful and simple coupling index which predicts that shifts in phytoplankton and bacterioplankton growth will be unrelated (Δ bacteria/Δ phytoplankton → either + ∞ or - ∞) in systems with inorganic N/P (molar) ratios of <~40. In systems with higher N/P ratios (>40), the coupling index will approach 1.0 and close coupling between bacteria and phytoplankton is predicted to occur.     

Depletion and release of prolactin from rat pituitaries and pituitary tumors in vitro

Depletion of pituitary prolactin (PRL) and PRL release into culture medium were simultaneously examined over a 3.5- to 4.0-hr incubation period from anterior pituitary fragments obtained from Fischer-344 or Wistar-Furth female rats treated with estrogen for 5 days, in pituitary tumors induced by 8 weeks of diethylstilbestrol (DES) treatment in Fischer-344 rats and in MtTW15 pituitary tumors transplanted subcutaneously in Wistar-Furth rats for 4 weeks. Our objective was to determine if the event known as transformation, which we define as a loss in the tissue PRL content without a corresponding and equivalent increase in the medium PRL content, occurs in rat pituitary tumors. Our results indicated that transformation did not occur in vitro in rat anterior pituitary tumors induced in Fischer-344 rats by DES treatment but was present in pituitaries from Fischer-344 rats treated for 5 days with estrogen, which served as controls. We also observed in vitro transformation in the anterior pituitary of Wistar-Furth rats treated with estrogen for 5 days (controls) and in the pituitaries of Wistar-Furth rats inoculated with the MtTW15 tumor for 4 weeks, but not in the MtTW15 tumor itself. Although transformation was present in both Fischer-344 and Wistar-Furth rats treated acutely with estrogen the timing of the transformation was delayed 1-2 hr in the Fischer-344 rats compared with Wistar-Furth females. We concluded that transformation does not precede release of prolactin in rat pituitary tumors and that in normal pituitaries the mechanisms of transformation are induced differently between the strains of rats examined.     

Marker-assisted selection for two rust resistance genes in sunflower

In this study we report on the identification of molecular markers, OX20₆₀₀ and OO04₉₅₀, linked to the geneR _Adv in the proprietary inbred line P2. This gene confers resistance to most of the pathotypes of Puccinia helianthi identified in Australia. Analysis indicates these RAPD markers are linked to the resistance locus at 0.0 cM and 11 cM respectively. SCAR markers SCX20₆₀₀ and SCO04₉₅₀ derived from these two RAPD markers, and SCT06₉₅₀ derived from a previously reported RAPD marker linked at 4.5 cM from the R ₁ rust resistance gene were developed. SCX20₆₀₀ and SCO04₉₅₀ were linked at similar distances from their resistance locus as the RAPD markers. SCTO6₉₅₀ co-segregated completely with rust resistance. The robustness of the R ₁ SCAR marker was demonstrated through the amplification of the marker in a diverse range of sunflower germplasm considered to possess the R ₁ gene. The SCAR markers forR _Adv were not amplified in the sunflower rust differential set thereby supporting the contention that this is a novel resistance gene. They did amplify in a number of proprietary lines closely related to the line P2. This locus is under further investigation as it will be useful in our attempts to use molecular-assisted breeding to produce durable resistance in sunflower to P. helianthi.     

Comparing two fish sampling standards over time: largely congruent results but with caveats

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