Self-reproduction of supramolecular giant vesicles combined with the amplification of encapsulated DNA.

Wednesday, September 21st 2011

A recent article published in Nature Chemistry is shortly explained and analyzed.


Figure 1.a. published in the paper: Amplification of DNA within a GV. An aqueous dispersion of GVs containing PCR reagents was prepared using a film-swelling method with a buffered solution containing template DNA, primers, fluorescent tag SYBR Green I, deoxynucleoside triphosphates, DNA polymerase and Mg2+
Figure 1.a. published in the paper: Amplification of DNA within a GV. An aqueous dispersion of GVs containing PCR reagents was prepared using a film-swelling method with a buffered solution containing template DNA, primers, fluorescent tag SYBR Green I, deoxynucleoside triphosphates, DNA polymerase and Mg2+

Reference: Nature Chemistry doi: 10.1038/nchem.1127

Title: Self-reproduction of supramolecular giant vesicles combined with the amplification of encapsulated DNA.

Authors: K. Kurihara, M. Tamura, K. Shohda, T. Toyota, K. Suzuki and T. Sugawara

Review by: Anna Venancio-Marques

The authors carry out simultaneously the self-reproduction of giant vesicles and the amplification of DNA encapsulated in these vesicles. They show that the two phenomena are linked and that thanks to electrostatic interactions, DNA amplification triggers self-replication of vesicles.

1. Experimental conditions

- vesicle composition: a hollow giant vesicle (GV) is obtained by mixing an amphiphile molecule V made up of two hydrophobis chains and bearing a benzaldehyde moiety at one end (used in previous studies of myelin-type GV) with POPC (a zwitterionic phospholipid that helps to form hollow vesicles), POPG ( an anionic phospholipid added to reduce surface charge ) and an amphilic acidic catalyst (used to transform membrane precursor molecules into membrane molecules). The vesicles obtained displayed a good resistance to highly ionic medium and to the high temperature associated with PCR reaction. The inner space is sufficient for PCR.
- vesicle preparation: the vesicles are obtained by swelling a film in the presence of DNA (DNAse is added to eliminate the DNA outside vesicles), then the vesicles are extruded to a final size of about 10 micrometers.
- DNA amplification: DNA (1 229 base pair) is amplified by PCR by the action of DNA polymerase with thermal cycles.
- self-reproduction: membrane precursor V* are added to the solution containing vesicles with amplified DNA content.

2. Results

A clear correlation was established between the amplification of DNA with PCR and the self-replication of the vesicles.
The authors explain that polyanionic DNA is embedded in the positively charged membrane, leading to a morphological change of the membrane as V molecules start to accumulate in the vicinity of the embedded DNA. The embedded DNA also helps to retain precursor membrane molecules in the membrane where they are then converted to membrane molecules. In the absence of DNA, the precursor molecule can simply diffuse across the membrane, and would therefore not contribute to the formation of new membrane.
Interestingly, during the process of self-reproduction of vesicles, DNA is partitioned in the daughter vesicles, whereas in prokaryotic cells specialized proteins fulfill that function.

In these experiments, self reproduction is limited by quantity of phospholipids and of nucleotide. As GV divide, the amount of phospholipid in the membrane decreases, changing the surface charge of the membrane. Authors hope to overcome this limitation by using a molecular transportation system. Challenges also include being able to synthesis the catalysts (whether for DNA replication with DNA polymerase, or for membrane formation with the specific amphiphilic acidic catalyst) to ensure that the process can be carried out over many generations.

3. Contribution to synthetic biology

A recent article in Nature Chemistry by Pier Luigi Luisi and Pasquale Stano (Synthetic biology: minimal cell mimicry, published online 23 september 2011) comment on this work. They underline that altougth the two phenomena occur one after the other, rather than at the same time and that the system itself has been optimised with the right membrane precursor V*, the achieved behaviour holds a lot of promises for future work in the field of synthetic biology.

Anna Venancio

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