Functional display of an alpha2 integrin-specific motif (RKK) on the surface of baculovirus particles.
The use of baculovirus vectors shows promise as a tool for gene delivery into mammalian cells. These insect viruses have been shown to transduce a variety of mammalian cell lines, and gene transfer has also been demonstrated in vivo. In this study, we generated two recombinant baculovirus vectors displaying an integrin-specific motif, RKK, as a part of two different loops of the green fluorescent protein (GFP) fused with the major envelope protein gp64 of Autographa californica M nucleopolyhedrovirus. By enzyme linked immunosorbent assays, these viruses were shown to bind a peptide representing the receptor binding site of an alpha2 integrin, the alpha2I-domain. However, the interaction was not strong enough to overcome binding of wild type gp64 to the unknown cellular receptor(s) on the surface of alpha2 integrin-expressing cells (CHO-alpha2beta1) or enhance the viral uptake. After treatment of these cells with phospholipase C, internalization of all viruses was blocked or decreased significantly. However, one of the RKK displaying viruses, AcGFP(K)gp64, was still able to internalize into CHO-alpha2beta1 cells, although at a lower level as compared to non-treated cells. This may indicate the possible utilization of a PLC independent alternative route via, in this case, the alpha2beta1 integrin.
Riikonen
Reetta
R
University of Jyvaskyla, Dept. of Biological and Environmental Science, PO Box 35, FIN-40351 Jyvaskyla, Finland.
Matilainen
Heli
H
Rajala
Nina
N
Pentikainen
Olli
O
Johnson
Mark
M
Heino
Jyrki
J
Oker-Blom
Christian
C
D020816:Amino Acid Motifs; D000818:Animals; D016367:Baculoviridae / Q000737:chemistry / Q000378:metabolism*; D016466:CHO Cells; D002460:Cell Line; D003001:Cloning, Molecular; D006224:Cricetinae; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D005434:Flow Cytometry; D018014:Gene Transfer Techniques; D049452:Green Fluorescent Proteins / Q000737:chemistry / Q000378:metabolism; D007313:Insecta; D039421:Integrin alpha2 / Q000737:chemistry*; D018613:Microscopy, Confocal; D008856:Microscopy, Fluorescence; D008958:Models, Molecular; D016296:Mutagenesis; D010455:Peptides / Q000737:chemistry; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry; D010738:Type C Phospholipases / Q000737:chemistry; D014760:Viral Fusion Proteins / Q000737:chemistry
D039421:Integrin alpha2; D010455:Peptides; D011994:Recombinant Proteins; D014760:Viral Fusion Proteins; D049452:Green Fluorescent Proteins; D010738:Type C Phospholipases