Polyelectrolyte multilayer-mediated gene delivery for semaphorin signaling pathway control.
The capability of multilayered polyelectrolyte films (MPFs) to control the sequential expression of two genes encoding cell receptors involved in a common cell signalling activity is shown, while achieving a fully functional signal transduction. As a functional model system representative of a cell signalling process that proceeds in a top-down manner, cell collapse induced by semaphorin 3A (Sema3A) was chosen as the target. Polyelectrolyte multilayers were sequentially functionalized with two plasmids encoding Neuropilin-1 (NRP-1) and Plexin-A1 (Plx-A1), respectively, acting as co-receptors for Sema3A. By using hyaluronan and chitosan as structural components for the incorporation of plasmid DNA layers onto precursor films made of poly-allylamine hydrochloride and poly-sodium-4-styrenesulfonate, the polyelectrolyte system is established; this systems is capable of delivering both plasmids to Cos-1 cells in a manner that permits control over the timing and the respective order in which the two plasmid DNA constructs are expressed. Importantly, it was observed that, following Sema3A stimulation, COS-1 cells co-expressing Plx-A1 and NRP-1 display a collapse phenotype, which is determined by the multilayer build-up scheme, and that the expression products of both transgenes embedded in MPFs are temporally functional over several days while acting their role of co-receptors for Sema3A.
Richard
Doriane
D
INSERM UMR 977, 11, rue Humann, 67085 Strasbourg, France.
Nguyen
Isabelle
I
Affolter
Christine
C
Meyer
Florent
F
Schaaf
Pierre
P
Voegel
Jean-Claude
JC
Bagnard
Dominique
D
Ogier
Joelle
J
D000818:Animals; D015153:Blotting, Western; D019556:COS Cells; D002522:Chlorocebus aethiops; D018014:Gene Transfer Techniques; D007150:Immunohistochemistry; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000378:metabolism; D039942:Neuropilin-1 / Q000235:genetics / Q000378:metabolism; D011108:Polymers / Q000138:chemical synthesis / Q000737:chemistry; D039961:Semaphorins / Q000494:pharmacology*; D015398:Signal Transduction / Q000235:genetics; D013451:Sulfonic Acids / Q000138:chemical synthesis / Q000737:chemistry; D014162:Transfection
D009419:Nerve Tissue Proteins; D011108:Polymers; D039961:Semaphorins; D013451:Sulfonic Acids; D039942:Neuropilin-1; C077114:styrenesulfonic acid polymer