Immobilization of bioactive fibroblast growth factor-2 into cubic proteinous microcrystals (Bombyx mori cypovirus polyhedra) that are insoluble in a physiological cellular environment.
The supramolecular architecture of the extracellular matrix and the disposition of its specific accessory molecules give rise to variable heterotopic signaling cues for single cells. Here we have described the successful occlusion of human fibroblast growth factor-2 (FGF-2) into the cubic inclusion bodies (FGF-2 polyhedra) of the Bombyx mori cytoplasmic polyhedrosis virus (BmCPV). The polyhedra are proteinous cubic crystals of several microns in size that are insoluble in the extracellular milieu. Purified FGF-2 polyhedra were found to stimulate proliferation and phosphorylation of p44/p42 mitogen-activated protein kinase in cultured fibroblasts. Moreover, cellular responses were blocked by a synthetic inhibitor of the FGF signaling pathway, SU5402, suggesting that FGF-2 polyhedra indeed act through FGF receptors. Furthermore, FGF-2 polyhedra retained potent growth stimulatory properties even after desiccation. We have demonstrated that BmCPV polyhedra microcrystals that occlude extracellular signaling proteins are a novel and versatile tool that can be employed to analyze cellular behavior at the single cell level.
Mori
Hajime
H
Insect Biomedical Research Center, Kyoto Institute of Technology, Matsugasaki, Sakyo-ku, Kyoto 606-8585, Japan.
Shukunami
Chisa
C
Furuyama
Akiko
A
Notsu
Hiroyuki
H
Nishizaki
Yuriko
Y
Hiraki
Yuji
Y
D000818:Animals; D001483:Base Sequence; D015153:Blotting, Western; D012831:Bombyx / Q000821:virology*; D002460:Cell Line; D003460:Crystallization; D017077:Culture Media, Conditioned; D017931:DNA Primers; D004797:Enzyme-Linked Immunosorbent Assay; D016222:Fibroblast Growth Factor 2 / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000378:metabolism; D051379:Mice; D041681:NIH 3T3 Cells; D017924:Nucleopolyhedroviruses / Q000737:chemistry*; D015398:Signal Transduction; D012995:Solubility; D018411:Spodoptera
D017077:Culture Media, Conditioned; D017931:DNA Primers; D016222:Fibroblast Growth Factor 2