RNA in model lipid membranes - interactions in bulk and at surfaces

Sammanfattning: The work presented in the thesis “RNA in model lipid membranes- interactions in bulk and at surfaces” aims to characterize the interaction between RNA and model membranes with different properties as well as to determine if ssRNA associate with model membranes in different ways. Several techniques have been used to characterize these systems, including quartz crystal microbalance with dissipation monitoring (QCM-D), Langmuir film balance in combination with neutron reflectometry and fluorescence microscopy as well as differential scanning calorimetry (DSC) and isothermal titration calorimetry (ITC). Understanding RNA-lipid interactions is important for many research areas, for example for understanding self-assembly in natural systems as well as for biotechnological development and for the development of new delivery systems for RNA based drugs. Association of RNA to model membranes was indicated in the Langmuir monolayer studies and confirmed with neutron reflectometry and QCM-D. The Langmuir monolayer experiments indicated that ssRNA was present at the interface of both zwitterionic and cationic fluid monolayers and with neutron reflectometry we were able to confirm that the ssRNA associated with the monolayer. From the neutron reflectometry studies it was also concluded that ssRNA penetrates into fluid monolayers. With QCM-D we confirmed that ssRNA associates to supported bilayers and that the orientation of the adsorbed ssRNA depends on the lipid phase behaviour. The utilization of model membranes as scaffolds for assembly of nucleic acids has also been studied, using QCM-D and null ellipsometry. The results show that it is possible to perform base pairing of simple RNA oligonucleotides as well as RNA assemblies with well-defined 3D shapes at the bilayer scaffold. From the calorimetric studies of RNA-lipid interactions in bulk solution, it was shown that RNA induces increased Tm as well as splitting of the transition peak in mixed lipid membranes, indicating segregation. We have been able to shown that the model membrane phase behaviour is essential for the association of ssRNA. The interpretation of these results is that the presence of hydrophobic moieties at the surface of fluid membranes is of significant importance for the association of ssRNA indicating that hydrophobic interaction between the exposed RNA bases and the lipid hydrocarbon region occurs. Our studies also showed that the bilayer is suitable to us as scaffolds for building both simple and more complex RNA constructs.