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Microsomal protein mediates a pH-dependent fusion of liposomes to rat brain microsomes
R Pistolesi1, L Corazzi, G Arienti
1Istituto di Biochimica e Chimica Medica, Perugia, Italy.
Abstract:
The fusion between rat brain microsomes and liposomes is investigated by measuring the release of octadecylrhodamine B (R18) fluorescence self-quenching. In the experimental conditions used in this work, the method allows a rapid and quantitative evaluation of the mixing of microsome and liposome lipid phases. The decrease of pH below 7 produces an extensive fusion between microsomes and acidic phospholipid liposomes. Microsomal protein is necessary for fusion, which is inactivated by exposure of microsomes to pronase. Therefore, H(+)-induced fusion differs from Ca(2+)-induced fusion since the latter does not require microsomal protein. The pretreatment of microsomes with trinitrobenzenesulfonic acid (TNBS) in nonpenetrating conditions does not affect the extent of fusion. On the other hand, N-ethoxycarbonyl-2-ethoxy-1,2-dihydroquinoline (EEDQ), a reagent able to react with carboxyl groups, causes an extensive inactivation of fusion. Therefore, the H(+)-induced fusion described here depends on some microsomal protein and may have physiological significance because it occurs at pH values present in the living cell. H(+)-dependent fusion can be also considered as a means to enrich membranes in some selected lipid.
Insights
Low pH triggers extensive fusion between rat brain microsomes and liposomes, requiring specific proteins. This H(+)-induced membrane fusion may have physiological relevance in living cells.
Area of Science:
- Biochemistry
- Cell Biology
- Membrane Biophysics
Background:
- Membrane fusion is a critical process in cellular function.
- Understanding the mechanisms of membrane fusion is essential for various biological processes.
- Investigating fusion between cellular membranes and artificial liposomes provides insights into membrane dynamics.
Purpose of the Study:
- To investigate the fusion between rat brain microsomes and liposomes.
- To characterize the conditions and requirements for pH-induced membrane fusion.
- To determine the role of microsomal proteins in H(+)-induced fusion.
Main Methods:
- Utilized octadecylrhodamine B (R18) fluorescence self-quenching to measure lipid phase mixing.
- Assessed fusion extent under varying pH conditions.
- Investigated the effect of protein inactivation (pronase, EEDQ) and chemical modification (TNBS) on microsomes.
Main Results:
- A decrease in pH below 7 induced extensive fusion between microsomes and acidic liposomes.
- Microsomal proteins were found to be essential for H(+)-induced fusion, as evidenced by inactivation with pronase and EEDQ.
- H(+)-induced fusion differs from Ca(2+)-induced fusion, which does not require microsomal proteins.
Conclusions:
- H(+)-induced fusion of rat brain microsomes and liposomes is a protein-dependent process.
- This fusion mechanism occurs at physiological pH values, suggesting potential in vivo relevance.
- The described H(+)-dependent fusion can be a method for selective lipid enrichment in membranes.