Role of membranes in mammalian stress response : sensing, lipid signals and adaptation

Abstract

It was suggested that under heat stress the accumulation of denatured proteins alone triggers the expression of heat shock proteins. However, earlier research suggested that during abrupt temperature fluctuations membranes represent the most thermally-sensitive macromolecular structures. The aim of this thesis to confirm experimentally for the membrane sensor theory in mammalian cells and to explore the mechanisms behind membrane lipid structural reorganizations. The main results are as follows: (i) I provide the first evidence that heat-analogous, chemically-induced membrane perturbation of K562 erythroleukemic cells is indeed capable of activating heat shock protein formation at the growth temperature, without causing measurable protein denaturation; (ii) I showed that the membrane fluidizer benzyl alcohol acts as a chaperone-inducer also in B16(F10) melanoma cells. Furthermore, following both alcohol and heat treatments, condensation of ordered plasma membrane domains was detected by fluorescence microscopy; (iii) lipidomic fingerprints revealed that stress achieved either by heat or benzyl alcohol resulted in pronounced and highly specific alterations of membrane lipids in B16(F10) cells. The loss in polyenes with the concomitant increase in saturated lipid species was shown to be a consequence of activation of phospholipases. The accumulation of lipid species with raft-forming properties may explain the condensation of ordered plasma membrane domains detected previously; (iv) with Laurdan two-photon microscopy it was demonstrated that, in contrast to the formation of ordered domains in surface membranes, the molecular disorder is significantly elevated within the internal membranes of cells preexposed to mild heat stress. These results were compared with those obtained by other probes and visualisation methods. It was found that the structurally different probes revealed substantially distinct alterations in membrane heterogeneity. The results highlight that even subtle changes in membrane microstructure may play a role in temperature sensing and thermal cell killing and, therefore, could have potential in treatment of several diseases.EThOS - Electronic Theses Online ServiceGBUnited Kingdo