Abstract
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.
Highlights
We developed a novel protein expression system that enables us to immobilize foreign proteins on insect virus occlusion bodies of protein crystals, termed polyhedra [3, 4]
The foreign proteins containing the immobilization signal are produced under the control of the polyhedrosis virus; BrdUrd, bromodeoxyuridine; FBS, fetal bovine serum; DME medium, Dulbecco’s modified Eagle’s medium; GFP, green fluorescence protein; PBS, phosphate-buffered saline; BSA, bovine serum albumin; ELISA, enzyme-linked immunosorbent assay; heparan sulfate proteoglycans (HSPGs), heparan sulfate proteoglycan; MAP, mitogen-activated protein; ITS, insulin, transferrin, and selenite
To evaluate the biological activity of fibroblast growth factor-2 (FGF-2), we studied the effects of FGF-2 polyhedra on the proliferation of mouse chondrogenic ATDC5 cells in culture
Summary
We developed a novel protein expression system that enables us to immobilize foreign proteins on insect virus occlusion bodies of protein crystals, termed polyhedra [3, 4]. To further visualize these dimensions, polyhedra containing immobilized enhanced GFP were added to confluent cultures of mouse chondrogenic ATDC5 cells (Fig. 1D) [11, 12]. To evaluate the biological activity of FGF-2, we studied the effects of FGF-2 polyhedra on the proliferation of mouse chondrogenic ATDC5 cells in culture.
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
