DNA-induced endocytosis upon local microinjection to giant unilamellar cationic vesicles

We suggest a novel approach for direct optical microscopy observation of DN A interaction with the bilayers of giant cationic liposomes. Giant unilamel lar vesicles, about 100 mu m in diameter, made of phosphatidylcholines and up to 33 mol% of the natural bioactive cationic amphiphile sphingosine, wer e obtained by electro formation. "Short" DNAs (oligonucleotide 21b and calf thymus 250 bp) were locally injected by micropipette to a part of the gian t unilamellar vesicle (GUV) membrane. DNAs were injected native, as well as marked with a fluorescent dye. The resulting membrane topology transformat ions were monitored in phase contrast, while DNA distribution was followed in fluorescence. We observed DNA-induced endocytosis due to the DNA/lipid m embrane local interactions and complex formation. A characteristic minimum concentration (C-endo) of D-erythro-sphingosine (Sph(+)) in the GUV membran e was necessary for the endocytic phenomenon to occur. Below C-endo, only l ateral adhesions between neighboring vesicles were observed upon DNA local addition. C-endo depends on the type of zwitterionic (phosphocholine) lipid used, being about 10 mol% for DPhPC/Sph(+) GUVs and about 20 mol% for SOPC /Sph(+) or eggPC/Sph(+) GUVs. The characteristic sizes and shapes of the re sulting endosomes depend on the kind of DNA, and initial GUV membrane tensi on. When the fluorescent DNA marker dye was injected after the DNA/lipid lo cal interaction and complex formation, no fluorescence was detected. This o bservation could be explained if one assumes that the DNA is protected by l ipids in the DNA/lipid complex, thereby inaccessible for the dye molecules. We suggest a possible mechanism for DNA/lipid membrane interaction involvi ng DNA encapsulation within an inverted micelle included in the lipid membr ane. Our model observations could help in under standing events associated with the interaction of DNA with biological membranes, as well as cationic liposomes/DNA complex formation in gene transfer processes.