Blue-light-activated phototropin2 trafficking from the cytoplasm to Golgi/post-Golgi vesicles

Phototropins are plasma membrane-localized UVA/blue light photoreceptors which mediate phototropism, inhibition of primary hypocotyl elongation, leaf positioning, chloroplast movements, and stomatal opening. Blue light irradiation activates the C-terminal serine/threonine kinase domain of phototropin which autophosphorylates the receptor. Arabidopsis thaliana encodes two phototropins, phot1 and phot2. In response to blue light, phot1 moves from the plasma membrane into the cytosol and phot2 translocates to the Golgi complex. In this study the molecular mechanism and route of blue-light-induced phot2 trafficking are demonstrated. It is shown that Atphot2 behaves in a similar manner when expressed transiently under 35S or its native promoter. The phot2 kinase domain but not blue-light-mediated autophosphorylation is required for the receptor translocation. Using co-localization and western blotting, the receptor was shown to move from the cytoplasm to the Golgi complex, and then to the post-Golgi structures. The results were confirmed by brefeldin A (an inhibitor of the secretory pathway) which disrupted phot2 trafficking. An association was observed between phot2 and the light chain2 of clathrin via bimolecular fluorescence complementation. The fluorescence was observed at the plasma membrane. The results were confirmed using co-immunoprecipitation. However, tyrphostin23 (an inhibitor of clathrin-mediated endocytosis) and wortmannin (a suppressor of receptor endocytosis) were not able to block phot2 trafficking, indicating no involvement of receptor endocytosis in the formation of phot2 punctuate structures. Protein turnover studies indicated that the receptor was continuously degraded in both darkness and blue light. The degradation of phot2 proceeded via a transport route different from translocation to the Golgi complex

Hot air stream reduces cytotoxicity of light-cured calcium hydroxide based cements

The light-cured calcium hydroxide based cements have incomplete polymerization and unconverted monomers can cause pulp cell damage. The aim of this study was to evaluate the influence of a warm and hot air stream on the cytotoxicity of light-cured calcium hydroxide based cements. The materials Dycal (conventional cement), Biocal, Hidrox-Cal, and Ultra-Blend Plus (light-cured calcium hydroxide cements) were submitted to cytotoxicity analysis after polymerization, without vs. with previous heat treatment with a warm (37°C) and a hot (60°C) air stream. Following polymerization, cements were maintained in culture medium for 24 hours and 7 days, and subjected to the MTT test. Data were analyzed using analysis of variance (ANOVA) followed by post-hoc Student-Newman-Keuls (<0.05). The results indicated significant differences between the materials according to their composition, i.e., light-cured cements treated with a jet of warm air showed similar cytotoxicity levels to those observed for conventional cement, suggesting that they may be considered alternatives in cases requiring pulp-capping treatment. Application of a hot air stream reduced cytotoxicity of materials tested

Characteristics of blue light-induced chloroplast movement and expression of GFP : myosin XI-H C-terminal domain construct in Nicotiana benthamiana leaves

Światło stanowi dla roślin źródło energii niezbędnej do prawidłowego funkcjonowania. Wykorzystywane jest przez chloroplasty w fazie jasnej fotosyntezy do produkcji wysokoenergetycznych ATP oraz NADPH. W zależności od natężenia absorbowanego światła jego wpływ może być zarówno dobroczynny jak i szkodliwy. Rośliny wykształciły zatem mechanizmy pozwalające na jego wykorzystanie w sposób jak najbardziej wydajny. Jednym z nich jest aktywowany światłem niebieskim ruch chloroplastów, na który składają się reakcja akumulacji (indukowana światłem słabym) oraz ucieczki (indukowana światłem silnym). W procesach tych pośredniczą fototropiny (receptory światła niebieskiego) oraz białka motoryczne odpowiedzialne m.in. za transport organelli wzdłuż mikrofilamentów (prawdopodobnie miozyny).Do głównych celów niniejszej pracy należało zbadanie reakcji ruchowych chloroplastów Nicotiana benthamiana w odpowiedzi na światło niebieskie oraz określenie stopnia udziału miozyny XI-H w tych przemieszczeniach.Wykazano, że kinetyka oraz amplitudy odpowiedzi ruchowych dla roślin młodych (2-tygodniowych) oraz starszych (6-tygodniowych) są bardzo zbliżone. Podobnie jak w Arabidopsis thaliana światło słabe (0,4; 1,6 μEm-2s-1) wywołuje reakcję akumulacji chloroplastów natomiast silne (120 μEm-2s-1) reakcję ucieczki, przy czym szybkość reakcji ucieczki jest 5-krotnie większa, zgodnie z jej rolą w ochronie aparatu fotosyntetycznego. W przeciwieństwie do A. thaliana, w przypadku N. benthamiana światło niebieskie o natężeniu 20 μEm-2s-1 wywołuje słabą reakcję akumulacji.W liściach A. thaliana wykazano ekspresję miozyny XI-H, której poziom nie zależy od światła. Skonstruowano wektor ekspresyjny pGreen-p35S-smGFP-ogonXI-H-t35S, następnie w liściach N. benthamiana uzyskano ekspresję białka fuzyjnego składającego się z białka reporterowego GFP oraz C-końcowej domeny miozyny XI-H. Poziom ekspresji był jednak zbyt niski na kontynuację badań, w tym weryfikację możliwego udziału miozyny XI-H w mechanizmie ruchu chloroplastów.Light is an energy source necessary for proper functioning of plants. It is used by chloroplasts in the light reactions of photosynthesis for production of energy-rich ATP and NADPH. Depending on the intensity of absorbed light its impact can be either beneficial or harmful. Therefore, plants have evolved mechanisms for more effective utilization of light. One of them is a blue light-controlled chloroplast redistribution which consists of an accumulation response (induced by weak light) and an avoidance response (induced by strong light). These processes are mediated by phototropins (blue light-specific photoreceptors) and motor proteins responsible inter alia for organelle trafficking along microfilaments (probably myosins).The main objectives of this work were to examine the Nicotiana benthamiana chloroplast responses to blue light and to determine the role of myosin XI-H in these relocations.It has been shown that the chloroplast movement kinetics and amplitudes are very similar for both young (2-week old) and older (6-week old) plants. Low light intensities (0,4; 1,6 μEm-2s-1) trigger the accumulation response and strong light (120 μEm-2s-1) induces the avoidance response. The velocity of the avoidance response is 5-fold higher than the accumulation reaction velocity consistent with a role of the former response in photoprotection of the photosynthetic apparatus. In contrast to A. thaliana, 20 μEm-2s-1 blue light causes a weak accumulation response in N. benthamiana.The expression of XI-H myosin was observed in A. thaliana leaves and its level did not depend on light. The construction of the expression vector pGreen-p35S-smGFP-tailXI-H-t35S allowed the expression of the fusion protein (GFP with a myosin C-terminal region) in N. benthamiana leaves. However, the expression level was too low to continue the research, in particular to verify a possible contribution of myosin XI-H in the mechanism of chloroplast movements

Relação entre o estilo cognitivo dependência-independência de campo e prática desportiva

This research investigated the relation between field dependence-independence and the practice of closed-skill and open-skill sports. Open-skill sports are characterized by the interaction between opponents, and the closed-skills sports lack external interferences on the athlete’s performance. Field dependence-independence is a cognitive style dimension associated with the degree of body boundaries differentiation. The Group Embedded Figures Test (GEFT) measured field independence in 149 athletes, 84 athletes of open-skill sports and 65 athletes of closed-skill sports, 77 men and 72 women. Men of closed-skill sports showed more field independence than men of open-skill sports; this difference didn’t happen in the feminine group. Within open-skills athletes women revealed higher field independence than men. The results showed the interaction between sex and sport type for differences in field independence.Esta pesquisa investigou a relação entre o estilo cognitivo dependência-independência de campo e a prática de esportes de habilidades abertas ou fechadas. Os esportes de habilidades abertas caracterizam-se pela interação de adversários em jogo, e os de habilidades fechadas pela ausência de perturbação externa sobre o desempenho do atleta. A dependência-independência de campo é uma dimensão de estilo cognitivo associada ao grau de diferenciação das fronteiras corporais. O Teste de Figuras Mascaradas (GEFT) foi aplicado em 149 sujeitos, 84 atletas de esportes de habilidades abertas e 65 de habilidades fechadas, 77 homens e 72 mulheres. Homens em esportes de habilidades fechadas mostraram-se mais independentes de campo do que os de habilidades abertas; esta diferença não ocorreu no grupo feminino. Entre os atletas de habilidades abertas, mulheres revelaram maior independência de campo do que homens. Conclui-se que há interação entre sexo e tipo de esporte para diferenças em independência de campo.Esta pesquisa investigó la relación entre el estilo cognitivo dependencia-independencia de campo y la práctica de deportes de habilidades cerradas o abiertas. Deportes de habilidades abiertas son caracterizados por la interacción entre oponentes, y deportes de habilidades cerradas son caracterizados por la ausencia de interferencias externas sobre el desempeño del atleta. Dependencia-independencia de campo es una dimensión de estilo cognitivo asociada con el grado de diferenciación de las fronteras corporales. El Teste de Figuras Enmascaradas (GEFT) mensuró la independencia de campo en 149 atletas, 84 atletas de deportes de habilidades abiertas y 65 atletas de deportes de habilidades cerradas, 77 hombres y 72 mujeres. Hombres en deportes de habilidades cerradas mostraran mayor independencia de campo do que hombres en deportes de habilidades abiertas; esta diferencia no ocurrió en el grupo femenino. En los atletas de deportes de habilidades abiertas las mujeres revelaran mayor independencia de campo do que hombres. Los resultados apuntan la interacción entre estilo cognitivo y sexo para diferencias en independencia de campo