Enhancement of cell proliferation and motility of mammalian cells grown in co-culture with Pichia pastoris expressing recombinant human FGF-2

Many studies examining the biological function of recombinant proteins and their effects on the physiology of mammalian cells stipulate that the proteins be purified before being used as therapeutic agents. In this study, we explored the possibility of using unpurified recombinant proteins to treat...

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Published in:	Protein expression and purification Vol. 176; p. 105724
Main Authors:	Le, Henry Hieu M., Vang, David, Amer, Nadia, Vue, Tou, Yee, Colwin, Kaou, Hyam, Harrison, Joseph S., Xiao, Nan, Lin-Cereghino, Joan, Lin-Cereghino, Geoff P., Thor, Der
Format:	Journal Article
Language:	English
Published:	United States Elsevier Inc 01-12-2020
Subjects:	Animals Cell motility Cell Proliferation Coculture Techniques FGF-2 Fibroblast Growth Factor 2 - biosynthesis Fibroblast Growth Factor 2 - genetics Gene Expression Humans Mice NIH 3T3 Cells NIH/3T3 Pichia pastoris Recombinant Proteins - biosynthesis Recombinant Proteins - genetics Saccharomycetales - genetics Saccharomycetales - growth & development Cell proliferation FGF-2 Cell motility Pichia pastoris NIH/3T3
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Summary:	Many studies examining the biological function of recombinant proteins and their effects on the physiology of mammalian cells stipulate that the proteins be purified before being used as therapeutic agents. In this study, we explored the possibility of using unpurified recombinant proteins to treat mammalian cells. The recombinant protein was used directly from the expression source and the biological function was compared to purified commercially available, equivalent protein. The model for this purpose was recombinant FGF-2, expressed by Pichia pastoris, which was used to treat the murine fibroblast cell line, NIH/3T3. We generated a P. pastoris strain (yHL11) that constitutively secreted a biologically active recombinant FGF-2 protein containing an N-terminal c-myc epitope (Myc-FGF-2). Myc-FGF-2 was then used without purification either a) in the form of conditioned mammalian cell culture medium or b) during co-cultures of yHL11 with NIH/3T3 to induce higher proliferation and motility of NIH/3T3 cells. The effects of Myc-FGF-2 on cell physiology were comparable to commercially available FGF-2. To our knowledge, this is the first time the physiology of cultured mammalian cells had been successfully altered with a recombinant protein secreted by P. pastoris while the two species shared the same medium and culture conditions. Our data demonstrated the biological activity of unpurified recombinant FGF-2 on NIH/3T3 cells and provided a foundation for directly using unpurified recombinant proteins expressed by P. pastoris with mammalian cells, potentially as wound-healing therapeutics. •P. pastoris strain yHL11 was generated to constitutively secrete recombinant FGF-2.•P. pastoris secreted FGF-2 using mammalian cell culture medium and conditions.•NIH/3T3 proliferation increased during co-culture with P. pastoris yHL11.•Unpurified FGF-2 secreted by P. pastoris had similar effects as purified FGF-2.•Co-culture enabled P. pastoris to deliver proteins to mammalian cells.
Bibliography:	ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23
ISSN:	1046-5928 1096-0279
DOI:	10.1016/j.pep.2020.105724