Genetically controlled differences in the effects of chlormequat on tetraploid wheat (Triticum turgidum)

Two crosses between Triticum turgidum wheat lines differing in their response to chlormequat (CCC) were tested. In the F₂ population of one cross, which was segregating for the Rht1 dwarfing allele, each plant was cloned by separation of two tillers, one of which was treated with CCC. The tall (rht1/rht1) and the intermediate (Rht1/rht1) genotypes showed a greater response to CCC than the semi-dwarf (Rht1/Rht1) genotype, as expressed by culm length and date of ear emergence. The F₃ families of another cross and their two semi-dwarf parents were grown in a three-replicated field test in paris of rows, one of which was treated with CCC. In one of the parents and in 1/4 of the F₃ families CCC induced a wide-angled tiller growth, suggesting a monogenic control of this growth habit in response to CCC.Based on an M.Sc. thesis presented by the senior author to the Faculty of Agriculture of The Hebrew University of Jerusalem.     

CCDC65 Mutation Causes Primary Ciliary Dyskinesia with Normal Ultrastructure and Hyperkinetic Cilia

Background

Primary ciliary dyskinesia (PCD) is a genetic disorder characterized by impaired ciliary function, leading to chronic sinopulmonary disease. The genetic causes of PCD are still evolving, while the diagnosis is often dependent on finding a ciliary ultrastructural abnormality and immotile cilia. Here we report a novel gene associated with PCD but without ciliary ultrastructural abnormalities evident by transmission electron microscopy, but with dyskinetic cilia beating.

Methods

Genetic linkage analysis was performed in a family with a PCD subject. Gene expression was studied in Chlamydomonas reinhardtii and human airway epithelial cells, using RNA assays and immunostaining. The phenotypic effects of candidate gene mutations were determined in primary culture human tracheobronchial epithelial cells transduced with gene targeted shRNA sequences. Video-microscopy was used to evaluate cilia motion.

Results

A single novel mutation in CCDC65, which created a termination codon at position 293, was identified in a subject with typical clinical features of PCD. CCDC65, an orthologue of the Chlamydomonas nexin-dynein regulatory complex protein DRC2, was localized to the cilia of normal nasal epithelial cells but was absent in those from the proband. CCDC65 expression was up-regulated during ciliogenesis in cultured airway epithelial cells, as was DRC2 in C. reinhardtii following deflagellation. Nasal epithelial cells from the affected individual and CCDC65-specific shRNA transduced normal airway epithelial cells had stiff and dyskinetic cilia beating patterns compared to control cells. Moreover, Gas8, a nexin-dynein regulatory complex component previously identified to associate with CCDC65, was absent in airway cells from the PCD subject and CCDC65-silenced cells.

Conclusion

Mutation in CCDC65, a nexin-dynein regulatory complex member, resulted in a frameshift mutation and PCD. The affected individual had altered cilia beating patterns, and no detectable ultrastructural defects of the ciliary axoneme, emphasizing the role of the nexin-dynein regulatory complex and the limitations of certain methods for PCD diagnosis.     

Alpha-synuclein and polyunsaturated fatty acids promote clathrin-mediated endocytosis and synaptic vesicle recycling

α-Synuclein (αS) is an abundant neuronal cytoplasmic protein implicated in Parkinson's disease (PD), but its physiological function remains unknown. Consistent with its having structural motifs shared with class A1 apolipoproteins, αS can reversibly associate with membranes and help regulate membrane fatty acid composition. We previously observed that variations in αS expression level in dopaminergic cultured cells or brains are associated with changes in polyunsaturated fatty acid (PUFA) levels and altered membrane fluidity. We now report that αS acts with PUFAs to enhance the internalization of the membrane-binding dye, FM 1-43. Specifically, αS expression coupled with exposure to physiological levels of certain PUFAs enhanced clathrin-mediated endocytosis in neuronal and non-neuronal cultured cells. Moreover, αS expression and PUFA-enhanced basal and -evoked synaptic vesicle (SV) endocytosis in primary hippocampal cultures of wild type (wt) and genetically depleted αS mouse brains. We suggest that αS and PUFAs normally function in endocytic mechanisms and are specifically involved in SV recycling upon neuronal stimulation.     

GIRK channel activation involves a local rearrangement of a preformed G protein channel complex

G protein-coupled signaling is one of the major mechanisms for controlling cellular excitability. One of the main targets for this control at postsynaptic membranes is the G protein-coupled potassium channels (GIRK/Kir3), which generate slow inhibitory postsynaptic potentials following the activation of Pertussis toxin-sensitive G protein-coupled receptors. Using total internal reflection fluorescence (TIRF) microscopy combined with fluorescence resonance energy transfer (FRET), in intact cells, we provide evidence for the existence of a trimeric G protein-channel complex at rest. We show that activation of the channel via the receptor induces a local conformational switch of the G protein to induce channel opening. The presence of such a complex thus provides the means for a precise temporal and highly selective activation of the channel, which is required for fine tuning of neuronal excitability.     

Modernity,Cultural Anesthesia,and Sensory Agency: Technologies of the Listening Self in a US Collegiate Jazz Music Program

In this article, I rely on Michel Foucault's notion of ‘technologies of the self’ to theorize the micro-practices by which individuals actively negotiate the reconfiguration of their sensory skills as a result of modernization processes. In doing so, I draw on ethnographic fieldwork I conducted in a collegiate jazz music program in the USA. By exploring a number of interactional games in which jazz students attempt to negotiate the challenge of cultivating aural skills in a pedagogical context that embraces visually mediated modes of knowledge production and transmission as a result of the professionalization and rationalization of jazz training, I inquire into the conditions of possibility for sensory agency under modernity.     

Effect of Chronic Lithium Treatment on 5-Hydroxytryptamine Autoreceptors and Release of 5-[3H]Hydroxytryptamine from Rat Brain Cortical, Hippocampal, and Hypothalamic Slices

The effect of acute and chronic lithium treatments on 5-hydroxytryptamine (5-HT, serotonin) release and on its regulation by presynaptic 5-HT autoreceptors was studied in [3H]5-HT preloaded superfused rat brain slices. The [3H]5-HT overflow evoked by a 30-s exposure to 65 mM K+ was increased after 3 weeks of ingestion of lithium-containing diet in the three brain areas examined. Acute injection of 4 mEq/kg lithium chloride did not affect 5-HT release. The K+-induced release observed in both control and chronically lithium-treated animals was Ca2+-dependent. Chronic lithium treatment was also found to be associated with a decrease in basal [3H]5-HT overflow in the cortex and hypothalamus but not in hippocampus [corrected]. The Ca2+-independent overflow induced by fenfluramine was also decreased in cortical slices from lithium-treated animals. The sensitivity of the inhibitory 5-HT autoreceptors was assessed by the response to the 5-HT agonist 5-methoxytryptamine. The results indicate a marked reduction in the maximal inhibition of [3H]5-HT release induced by 5-methoxytryptamine in slices obtained from animals which have been treated with lithium for 3 weeks. These data suggest that the functional down regulation of the prejunctional 5-HT sites may be responsible for the increase in K+-stimulated 5-HT overflow in brain slices of animals treated chronically with lithium.     

SKF83959 selectively regulates phosphatidylinositol-linked D1 dopamine receptors in rat brain

Previously a distinct D1-like dopamine receptor (DAR) that selectively couples to phospholipase C/phosphatidylinositol (PLC/PI) was proposed. However, lack of a selective agonist has limited efforts aimed at characterizing this receptor. We characterized the in vitro and in vivo effects of SKF83959 in regulating PI metabolism. SKF83959 stimulates (EC50, 8 micro m) phosphatidylinositol 4,5-biphosphate hydrolysis in membranes of frontal cortex (FC) but not in membranes from PC12 cells expressing classical D1A DARs. Stimulation of FC PI metabolism was attenuated by the D1 antagonist, SCH23390, indicating that SKF83959 activates a D1-like DAR. The PI-linked DAR is located in hippocampus, cerebellum, striatum and FC. Most significantly, administration of SKF83959 induced accumulations of IP3 in striatum and hippocampus. In contrast to other D1 DAR agonists, SKF83959 did not increase cAMP production in brain or in D1A DAR-expressing PC12 cell membranes. However, SKF83959 inhibited cAMP elevation elicited by the D1A DAR agonist, SKF81297, indicating that the compound is an antagonist of the classical D1A DAR. Lastly, we demonstrated that SKF83959 enhances [35S]guanosine 5'-O-(3-thiotriphosphate) binding to membrane Galphaq and Galphai proteins, suggesting that PI stimulation is mediated by activation of these guanine nucleotide-binding regulatory proteins. Results indicate that SKF83959 is a selective agonist for the PI-linked D1-like DAR, providing a unique tool for investigating the functions of this brain D1 DAR subtype.     

Macrophages function as a ferritin iron source for cultured human erythroid precursors

Iron is essential for the survival as well as the proliferation and maturation of developing erythroid precursors (EP) into hemoglobin-containing red blood cells. The transferrin-transferrin receptor pathway is the main route for erythroid iron uptake. Using a two-phase culture system, we have previously shown that placental ferritin as well as macrophages derived from peripheral blood monocytes could partially replace transferrin and support EP growth in a transferrin-free medium. We now demonstrate that in the absence of transferrin, ferritin synthesized and secreted by macrophages can serve as an iron source for EP. Macrophages trigger an increase in both the cytosolic and the mitochondrial labile iron pools, in heme and in hemoglobin synthesis, along with a decrease in surface transferrin receptors. Inhibiting macrophage exocytosis, binding extracellular ferritin with specific antibodies, inhibiting EP receptor-mediated endocytosis or acidification of EP lysosomes, all resulted in a decreased EP growth when co-cultured with macrophages under transferrin-free conditions. The results suggest that iron taken up by macrophages is incorporated mainly into their ferritin, which is subsequently secreted by exocytosis. Nearby EP are able to take up this ferritin probably through clathrin-dependent, receptor-mediated endocytosis into endosomes, which following acidification and proteolysis release the iron from the ferritin, making it available for regulatory and synthetic purposes. Thus, macrophages support EP development under transferrin-free conditions by delivering essential iron in the form of metabolizable ferritin.