Cellular membrane shape plays an important role in cellular signaling events, including endocytosis and exocytosis. This project focused on studying how cellular cations, osmotic pressure, and lipid number asymmetry cause membrane shape deformations in giant unilamellar lipid vesicles (GUVs). GUVs are used in many studies because of their similarity to cells and easy preparation. While sodium and potassium ions are common cellular ions, their effect on membrane shape is unclear. So, we investigated the effects of sodium and potassium ions on phospholipid GUVs by incubating the vesicles in solutions of these cations and imaging with a confocal microscope. These images were analyzed to determine if deformations - specifically internal or external tubules - could be observed. Injection experiments were also performed by injecting a potassium solution near an aspirated GUV to provide more information about potassium. The sodium results showed that it induces positive curvature in GUVs, but the potassium results were inconsistent. The effects of osmolarity imbalances on GUVs were examined by carefully controlling the osmolarity of the GUVs and the incubation solution so that the incubation solution had a higher osmolarity than the vesicles. Analysis of the GUVs indicated that at high differences in osmolarity, small vesicles bud off of GUVs externally, but at lower differences in osmolarity, they form internally. Finally, methyl-b-cyclodextrin (MbCD), which is a molecule that extracts cholesterol from lipid membranes, was studied at low concentrations to determine if it could extract lipids from cholesterol-free membranes. GUVs were incubated in solutions containing varying concentrations of MbCD and imaged. The results suggested that MbCD extracts lipids from the membrane, and lipid extraction increases with the concentration of MbCD. All of these studies yielded interesting results that provide insight into the properties of GUVs and causes of membrane deformation.
Through this research experience, I had a chance to learn about a new topic that is vital to chemistry and biology studies around the world. I worked with new chemicals and techniques, which allowed me to improve upon my laboratory skills. Additionally, I spent a lot of time troubleshooting techniques because of the inconsistent results with potassium ions. Even though the effect of potassium on GUVs remains unclear, we had a chance to work on a different experimental design, and this indicated to us that there may be other unknown factors that cloud the interactions between the GUV membranes and potassium. Therefore, I had the opportunity to understand the amount of work that researchers and PhD students around the world do in their projects in order to achieve successful results. The experience over the summer was very rewarding and gave me a chance to learn about chemistry as well as experience full-time research.
Cellular membrane shape plays an important role in cellular signaling events, including endocytosis and exocytosis. This project focused on studying how cellular cations, osmotic pressure, and lipid number asymmetry cause membrane shape deformations in giant unilamellar lipid vesicles (GUVs). GUVs are used in many studies because of their similarity to cells and easy preparation. While sodium and potassium ions are common cellular ions, their effect on membrane shape is unclear. So, we investigated the effects of sodium and potassium ions on phospholipid GUVs by incubating the vesicles in solutions of these cations and imaging with a confocal microscope. These images were analyzed to determine if deformations - specifically internal or external tubules - could be observed. Injection experiments were also performed by injecting a potassium solution near an aspirated GUV to provide more information about potassium. The sodium results showed that it induces positive curvature in GUVs, but the potassium results were inconsistent. The effects of osmolarity imbalances on GUVs were examined by carefully controlling the osmolarity of the GUVs and the incubation solution so that the incubation solution had a higher osmolarity than the vesicles. Analysis of the GUVs indicated that at high differences in osmolarity, small vesicles bud off of GUVs externally, but at lower differences in osmolarity, they form internally. Finally, methyl-b-cyclodextrin (MbCD), which is a molecule that extracts cholesterol from lipid membranes, was studied at low concentrations to determine if it could extract lipids from cholesterol-free membranes. GUVs were incubated in solutions containing varying concentrations of MbCD and imaged. The results suggested that MbCD extracts lipids from the membrane, and lipid extraction increases with the concentration of MbCD. All of these studies yielded interesting results that provide insight into the properties of GUVs and causes of membrane deformation.
Through this research experience, I had a chance to learn about a new topic that is vital to chemistry and biology studies around the world. I worked with new chemicals and techniques, which allowed me to improve upon my laboratory skills. Additionally, I spent a lot of time troubleshooting techniques because of the inconsistent results with potassium ions. Even though the effect of potassium on GUVs remains unclear, we had a chance to work on a different experimental design, and this indicated to us that there may be other unknown factors that cloud the interactions between the GUV membranes and potassium. Therefore, I had the opportunity to understand the amount of work that researchers and PhD students around the world do in their projects in order to achieve successful results. The experience over the summer was very rewarding and gave me a chance to learn about chemistry as well as experience full-time research.