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Exploring the biophysical properties of biological membranes: from ceramide domains to lipid droplets
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Abstract(s)
Actualmente é reconhecido que a organização da membrana e as suas propriedades são reguladores de funções celulares. As alterações na organização membranar e a formação de domínios ordenados estão implicados em diversos processos biológicos, tais como na sinalização celular. As ceramidas constituem as cadeias estruturais de todos os esfingolípidos complexos, e são considerados lípidos bioactivos capazes de modular vários processos celulares e implicados no desenvolvimento de várias doenças. O mecanismo molecular subjacente ao modo de acção das ceramidas parece estar relacionado com o modo como estes lípidos alteram as propriedades biofísicas das membranas, nomeadamente através da formação de domínios enriquecidos em ceramida. No entanto, o efeito da ceramida nas propriedades das membranas são complexos e dependem tanto da composição lipídica da membrana como da estrutura da ceramida, e estão pouco caracterizados em situações que mimetizem comportamentos biológicos. Por isso, este estudo tem como objectivo investigar o impacto biofísico das ceramidas em membranas biológicas e em condições fisiologicamente relevantes, e identificar as consequências biológicas associadas a estas alterações. Vários estudos foram feitos tendo como objectivo i) melhorar a caracterização das ceramidas em membranas artificiais e ii) desenvolver e implementar metodologias biofísicas para caracterizar as propriedades de membranas e organelos em células e em condições fisiológicas. Os resultados obtidos neste estudo confirmam a complexidade associadas às alterações promovidas pela ceramida nas propriedades biofísicas das membranas, que passam pela formação de fases gel metaestáveis interdigitadas. Para além disso, também é mostrado que a formação de ceramida induzida por factores de stress promove alterações na fluidez da membrana plasmática, nos lisossomas e na célula em geral. A capacidade da ceramida promover um aumento da ordem da membrana, causado pela formação dos domínios gel de ceramida, está coordenado com o aumento da internalização de sondas lipofílicas, e é acompanhado por um aumento na internalização de vesiculas enriquecidas em ceramida, as quais são transportadas através da via endo/lisossomal. Além disso, este estudo mostra que a formação de ceramida e as alterações biofísicas estão associadas ao metabolismo e propriedades das gotículas lipídicas (“lipid droplets”). Resumindo, este estudo demostra que as acções biológicas da ceramida poderão estar associadas às alterações biofísicas induzidas nas membranas. O efeito das ceramidas nas propriedades das membranas propaga-se para além do local onde são formadas, e envolvem alterações em vários organelos. Estas evidências sugerem que activação de alvos intracelulares relacionados com as ceramidas possa ser mediada pelas vesiculas enriquecidas em ceramida com propriedades biofísicas únicas.
It is nowadays recognized that membrane organization and properties are key regulators of cell function. Changes in lateral organization of the membranes and formation of ordered membrane domains have been implicated in many biological processes, including cell signaling. Ceramides, the backbone of all complex sphingolipids, are bioactive lipids involved in many cellular processes and implicated in several diseases. It is hypothesized that the biological actions of this highly hydrophobic lipid occur at the membrane level, through modulation of membrane biophysical properties, including the formation of ceramide-enriched domains. However, the effects of ceramides on membrane properties are complex and depend both on membrane lipid composition and ceramide structure and are poorly characterized in biological relevant settings. Therefore, this study aimed to investigate the biophysical impact of ceramides in biological membranes under physiological relevant conditions and identify biological consequences associated to these changes. To that end, multiple studies were performed that i) enabled a better characterization of the biophysical properties of ceramides in artificial membranes and ii) the development and implementation of biophysical methodologies to characterize the properties of membranes and organelles in live cells under physiological relevant conditions. The results obtained in this study further confirmed the complexity of ceramides associated changes on membrane biophysical properties, which encompass the formation of metastable interdigitated gel phases. Moreover, it is shown that stress-induced ceramide generation causes alterations in the fluidity of the plasma membrane, but also of the lysosomes and whole cells. Membrane ordering caused by ceramide-gel domains formation is coordinated with increased internalization of lipophilic probes and accompanied by increased internalization of ceramide-enriched vesicles that traffic via the endo-lysosomal pathway. Moreover, this study further shows that ceramide-generation and membrane biophysical alterations are linked to lipid droplet metabolism and properties. In summary, this study demonstrates that ceramide biological actions might be linked to the alterations induced by these lipids in the membranes. The effects of ceramides extend beyond the site where they are formed, reaching multiple intracellular sites. This evidence further suggests that ceramides might reach and activate their intracellular targets via formation of ceramide-enriched vesicles with distinct biophysical properties.
It is nowadays recognized that membrane organization and properties are key regulators of cell function. Changes in lateral organization of the membranes and formation of ordered membrane domains have been implicated in many biological processes, including cell signaling. Ceramides, the backbone of all complex sphingolipids, are bioactive lipids involved in many cellular processes and implicated in several diseases. It is hypothesized that the biological actions of this highly hydrophobic lipid occur at the membrane level, through modulation of membrane biophysical properties, including the formation of ceramide-enriched domains. However, the effects of ceramides on membrane properties are complex and depend both on membrane lipid composition and ceramide structure and are poorly characterized in biological relevant settings. Therefore, this study aimed to investigate the biophysical impact of ceramides in biological membranes under physiological relevant conditions and identify biological consequences associated to these changes. To that end, multiple studies were performed that i) enabled a better characterization of the biophysical properties of ceramides in artificial membranes and ii) the development and implementation of biophysical methodologies to characterize the properties of membranes and organelles in live cells under physiological relevant conditions. The results obtained in this study further confirmed the complexity of ceramides associated changes on membrane biophysical properties, which encompass the formation of metastable interdigitated gel phases. Moreover, it is shown that stress-induced ceramide generation causes alterations in the fluidity of the plasma membrane, but also of the lysosomes and whole cells. Membrane ordering caused by ceramide-gel domains formation is coordinated with increased internalization of lipophilic probes and accompanied by increased internalization of ceramide-enriched vesicles that traffic via the endo-lysosomal pathway. Moreover, this study further shows that ceramide-generation and membrane biophysical alterations are linked to lipid droplet metabolism and properties. In summary, this study demonstrates that ceramide biological actions might be linked to the alterations induced by these lipids in the membranes. The effects of ceramides extend beyond the site where they are formed, reaching multiple intracellular sites. This evidence further suggests that ceramides might reach and activate their intracellular targets via formation of ceramide-enriched vesicles with distinct biophysical properties.
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Keywords
ceramida gotículas lipídicas propriedades biofísicas espectroscopia e microscopia de fluorescência metabolismo celular ceramide lipid droplets biophysical properties fluorescence spectroscopy and microscopy cell metabolismo