Order disorder phase transitions in the plasma membrane of HeLa cells measured by fluorescent analysis of solvatochromic probes

Abstract

Using the solvatochromic membrane probes Laurdan and Pro12A we investigated order-disorder phase transitions in cellular lipid membranes of HeLa cells. Due to internalization of Laurdan its fluorescence signal yields information about inner and outer cellular membranes while Pro12A exclusively stains the plasma membrane. The two different membrane-embedded dyes show an emission redshift upon increasing disorder of the surrounding lipids that can be quantified by the Generalized Polarization (GP). First, we compare the sensitivity of both probes to lipid phase transitions by analyzing GP of synthetic lipid vesicles as function of temperature. Second, we investigate the temperature dependent lipid order of HeLa cell membranes and find that the plasma membrane shows a partially reversible order-disorder transition regime at temperatures between T = 20 {degrees}C and T = 70 {degrees}C. Third, we calorimetrically detect an irreversible transition at T = 55 {degrees}C and conclude that the optically detected restructuring of the plasma membrane can be partially attributed to protein denaturation. At last, it is shown that the reversible plasma membrane transition temperature Tm shifts from Tm = 25 {degrees}C to Tm =-16 {degrees}C upon cholesterol depletion and sharpens from a half width of {Delta}TFWHM = 60 K to {Delta}TFWHM = 8 K. The reversibility and the sensitivity to cholesterol of this transition indicate a temperature-induced lipid melting within the plasma membrane of HeLa cells.