Surface Display of Human GM-CSF in Methylotrophic Yeasts

Human granulocyte-macrophage colony-stimulating factor (GM-CSF) is an important therapeutic cytokine. Methylotrophic yeasts such as Komagataella phaffii and Ogataea parapolymorpha are attractive hosts for recombinant protein production. In this study, these yeasts were engineered to produce GM-CSF in both secreted and cell-surface-anchored forms. Secreted GM-CSF accumulated to tens of milligrams per liter in culture supernatants following induction. Fluorescent antibody labeling confirmed that yeast strains expressing surface-displayed GM-CSF exhibited markedly increased fluorescence compared to parental strains. The highest signal was observed in K. phaffii and further validated by immunofluorescence microscopy. Functional assays demonstrated that K. phaffii cells displaying GM-CSF stimulated TF-1 cell proliferation 1.41-fold compared to control yeast lacking GM-CSF. These results confirm both the successful surface display and preserved biological activity of the cytokine. This work demonstrates the feasibility of engineering K. phaffii to present active human GM-CSF on the cell surface. Surface display was efficient and yielded biologically functional cytokine, as shown by fluorescence-based quantification and TF-1 proliferation assays. The study highlights yeast surface display as a promising platform for delivering therapeutic cytokines without requiring purification steps.