DNA-induced endocytosis upon local microinjection to giant unilamellar cationic vesicles |
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Authors: | M I Angelova Nadejda Hristova Iana Tsoneva |
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Institution: | (1) Institute of Biophysics, Bulgarian Academy of Sciences, Acad. G. Bonchev Str., Bl. #21, BG-1113 Sofia, Bulgaria e-mail: mig@obzor.bio21.bas.bg, BG |
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Abstract: | We suggest a novel approach for direct optical microscopy observation of DNA interaction with the bilayers of giant cationic
liposomes. Giant unilamellar vesicles, about 100 μm in diameter, made of phosphatidylcholines and up to 33 mol% of the natural
bioactive cationic amphiphile sphingosine, were obtained by electroformation. “Short” DNAs (oligonucleotide 21b and calf thymus
250 bp) were locally injected by micropipette to a part of the giant unilamellar vesicle (GUV) membrane. DNAs were injected
native, as well as marked with a fluorescent dye. The resulting membrane topology transformations were monitored in phase
contrast, while DNA distribution was followed in fluorescence. We observed DNA-induced endocytosis due to the DNA/lipid membrane
local interactions and complex formation. A characteristic minimum concentration (C
endo) of d-erythro-sphingosine (Sph+) in the GUV membrane was necessary for the endocytic phenomenon to occur. Below C
endo, only lateral 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 resulting endosomes depend on the kind of DNA, and initial GUV membrane
tension. When the fluorescent DNA marker dye was injected after the DNA/lipid local interaction and complex formation, no
fluorescence was detected. This observation could be explained if one assumes that the DNA is protected by lipids in the DNA/lipid
complex, thereby inaccessible for the dye molecules. We suggest a possible mechanism for DNA/lipid membrane interaction involving
DNA encapsulation within an inverted micelle included in the lipid membrane. Our model observations could help in understanding
events associated with the interaction of DNA with biological membranes, as well as cationic liposomes/DNA complex formation
in gene transfer processes.
Received: 18 April 1998 / Revised version: 6 August 1998 / Accepted: 7 August 1998 |
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Keywords: | DNA interactions Cationic giant unilamellar vesicle Microinjection Sphingosine Endocytosis |
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