Independent Functional Expression of Populus trichocarpa Cellulose Synthase Isoforms CesA1 and CesA8 in Escherichia coli

Abstract Cellulose synthase (CesA) drives cellulose biosynthesis in plants, yet the isoform-specific roles of CesA subunits in cellulose assembly remain elusive. To dissect their distinct functions, two critical isoforms of PtCesA1 and PtCesA8 from Populus trichocarpa were heterologous expressed in Escherichia coli.Both PtCesA1 and PtCesA8 proteins were expressed correctly in E.coli. Transmission electron microscopy (TEM) revealed both PtCesA1 and PtCesA8 formed disordered needle-like networks more than vector control. These architectures were validated by ATR-FTIR spectroscopy and cellulase hydrolysis, confirming β-1,4-glucan synthesis distinct from bacterial polysaccharides. Immunofluorescence localized PtCesA1 to linear terminal complexes, whereas PtCesA8 showed disorganized distributions, suggesting divergent assembly mechanisms.To our knowledge, this study pioneers the independent prokaryotic expression of plant CesA isoforms in E. coli, circumventing the cooperative subunit interactions required in native systems. By bridging plant and bacterial cellulose synthesis paradigms, we establish a microbial platform for producing structurally tailored cellulose, advancing both fundamental understanding and bio-inspired materials engineering.