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Reversible Dissolution of Microdomains in Detergent-Resistant Membranes at Physiological Temperature.

Cremona A, Orsini F, Corsetto PA, Hoogenboom BW, Rizzo AM - PLoS ONE (2015)

Bottom Line: This shrinking in microdomain size was accompanied by a gradual reduction of the height difference between the microdomains and the surrounding membrane, consistent with the behaviour expected for lipids that are laterally segregated in liquid ordered and liquid disordered domains.Immunolabeling experiments demonstrated that the microdomains contained flotillin-1, a protein associated with lipid rafts.The microdomains reversibly dissolved and reappeared, respectively, on heating to and cooling below temperatures around 37 °C, which is indicative of radical changes in local membrane order close to physiological temperature.

View Article: PubMed Central - PubMed

Affiliation: Dipartimento di Scienze Farmacologiche e Biomolecolari, Università degli Studi di Milano, Milano, Italy.

ABSTRACT
The formation of lipid microdomains ("rafts") is presumed to play an important role in various cellular functions, but their nature remains controversial. Here we report on microdomain formation in isolated, detergent-resistant membranes from MDA-MB-231 human breast cancer cells, studied by atomic force microscopy (AFM). Whereas microdomains were readily observed at room temperature, they shrunk in size and mostly disappeared at higher temperatures. This shrinking in microdomain size was accompanied by a gradual reduction of the height difference between the microdomains and the surrounding membrane, consistent with the behaviour expected for lipids that are laterally segregated in liquid ordered and liquid disordered domains. Immunolabeling experiments demonstrated that the microdomains contained flotillin-1, a protein associated with lipid rafts. The microdomains reversibly dissolved and reappeared, respectively, on heating to and cooling below temperatures around 37 °C, which is indicative of radical changes in local membrane order close to physiological temperature.

No MeSH data available.


Related in: MedlinePlus

Temperature dependence of microdomain size and height.a, Plot of the relative areas of microdomains and membrane patches as a function of the temperature. b, Average of the surface area of individual microdomains as a function of temperature, normalized to the surface area of the respective microdomains at 25°C. c, Plot of the microdomain and membrane patch heights, both measured with respect to the mica, as a function of the temperature. d, Height of the microdomains measured with respect to the surrounding membrane patch surface, as a function of temperature. All data represent the mean ± SD obtained analyzing 16 different thermal cycles.
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pone.0132696.g003: Temperature dependence of microdomain size and height.a, Plot of the relative areas of microdomains and membrane patches as a function of the temperature. b, Average of the surface area of individual microdomains as a function of temperature, normalized to the surface area of the respective microdomains at 25°C. c, Plot of the microdomain and membrane patch heights, both measured with respect to the mica, as a function of the temperature. d, Height of the microdomains measured with respect to the surrounding membrane patch surface, as a function of temperature. All data represent the mean ± SD obtained analyzing 16 different thermal cycles.

Mentions: This behaviour was confirmed and further analysed in different experiments including in total 16 thermal cycles between a minimum of 23°C and a maximum of 46°C. Fig 3A reports the total relative areas on the membranes occupied by microdomains, as a function of temperature (100% = total area of all membrane patches observed in the AFM images). At 25°C, microdomains constituted about 15% of the total membrane area, a fraction that was rapidly reduced to a few % for temperatures between 32 and 36°C, and that was hardly distinguishable above 37°C. The same behaviour was observed using an alternative analysis, for which the total area of each microdomain at 25°C was defined as 100% and where the relative size of the microdomain was tracked as a function of temperature. Fig 3B shows the average relative areas for different microdomains in different experiments. We note that only in rare occasions, few and small microdomains persisted up to 44°C (see S4 Fig).


Reversible Dissolution of Microdomains in Detergent-Resistant Membranes at Physiological Temperature.

Cremona A, Orsini F, Corsetto PA, Hoogenboom BW, Rizzo AM - PLoS ONE (2015)

Temperature dependence of microdomain size and height.a, Plot of the relative areas of microdomains and membrane patches as a function of the temperature. b, Average of the surface area of individual microdomains as a function of temperature, normalized to the surface area of the respective microdomains at 25°C. c, Plot of the microdomain and membrane patch heights, both measured with respect to the mica, as a function of the temperature. d, Height of the microdomains measured with respect to the surrounding membrane patch surface, as a function of temperature. All data represent the mean ± SD obtained analyzing 16 different thermal cycles.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4493071&req=5

pone.0132696.g003: Temperature dependence of microdomain size and height.a, Plot of the relative areas of microdomains and membrane patches as a function of the temperature. b, Average of the surface area of individual microdomains as a function of temperature, normalized to the surface area of the respective microdomains at 25°C. c, Plot of the microdomain and membrane patch heights, both measured with respect to the mica, as a function of the temperature. d, Height of the microdomains measured with respect to the surrounding membrane patch surface, as a function of temperature. All data represent the mean ± SD obtained analyzing 16 different thermal cycles.
Mentions: This behaviour was confirmed and further analysed in different experiments including in total 16 thermal cycles between a minimum of 23°C and a maximum of 46°C. Fig 3A reports the total relative areas on the membranes occupied by microdomains, as a function of temperature (100% = total area of all membrane patches observed in the AFM images). At 25°C, microdomains constituted about 15% of the total membrane area, a fraction that was rapidly reduced to a few % for temperatures between 32 and 36°C, and that was hardly distinguishable above 37°C. The same behaviour was observed using an alternative analysis, for which the total area of each microdomain at 25°C was defined as 100% and where the relative size of the microdomain was tracked as a function of temperature. Fig 3B shows the average relative areas for different microdomains in different experiments. We note that only in rare occasions, few and small microdomains persisted up to 44°C (see S4 Fig).

Bottom Line: This shrinking in microdomain size was accompanied by a gradual reduction of the height difference between the microdomains and the surrounding membrane, consistent with the behaviour expected for lipids that are laterally segregated in liquid ordered and liquid disordered domains.Immunolabeling experiments demonstrated that the microdomains contained flotillin-1, a protein associated with lipid rafts.The microdomains reversibly dissolved and reappeared, respectively, on heating to and cooling below temperatures around 37 °C, which is indicative of radical changes in local membrane order close to physiological temperature.

View Article: PubMed Central - PubMed

Affiliation: Dipartimento di Scienze Farmacologiche e Biomolecolari, Università degli Studi di Milano, Milano, Italy.

ABSTRACT
The formation of lipid microdomains ("rafts") is presumed to play an important role in various cellular functions, but their nature remains controversial. Here we report on microdomain formation in isolated, detergent-resistant membranes from MDA-MB-231 human breast cancer cells, studied by atomic force microscopy (AFM). Whereas microdomains were readily observed at room temperature, they shrunk in size and mostly disappeared at higher temperatures. This shrinking in microdomain size was accompanied by a gradual reduction of the height difference between the microdomains and the surrounding membrane, consistent with the behaviour expected for lipids that are laterally segregated in liquid ordered and liquid disordered domains. Immunolabeling experiments demonstrated that the microdomains contained flotillin-1, a protein associated with lipid rafts. The microdomains reversibly dissolved and reappeared, respectively, on heating to and cooling below temperatures around 37 °C, which is indicative of radical changes in local membrane order close to physiological temperature.

No MeSH data available.


Related in: MedlinePlus