title: Recombinant protein yield in rice seed is enhanced by specific suppression of endogenous seed proteins at the same deposit site.
abstract: Human IL-10 (hIL-10) is a therapeutic treatment candidate for inflammatory allergy and autoimmune diseases. Rice seed-produced IL-10 can be effectively delivered directly to gut-associated lymphoreticular tissue (GALT) via bio-encapsulation. Previously, the codon-optimized hIL-10 gene was expressed in transgenic rice with the signal peptide and endoplasmic reticulum (ER) retention signal (KDEL) at its 5' and 3' ends, respectively, under the control of the endosperm-specific glutelin GluB-1 promoter. The resulting purified hIL-10 was biologically active. In this study, the yield of hIL-10 in transgenic rice seed was improved. This protein accumulated at the intended deposition sites, which had been made vacant through the selective reduction, via RNA interference, of the endogenous seed storage proteins prolamins or glutelins. Upon suppression of prolamins that were sequestered into ER-derived protein bodies (PB-I), hIL-10 accumulation increased approximately 3-fold as compared to rice seed with no such suppression and reached 219 μg/grain. In contrast, reducing the majority of the glutelins stored in protein-storage vacuoles (PB-II) did not significantly affect the accumulation of hIL-10. Considering that hIL-10 is synthesized in the ER lumen and subsequently buds off in ER-derived granules called IL-10 granules in a manner similar to PB-Is, these results indicate that increases in the available deposition space for the desired recombinant proteins may be crucial for improvements in yield. Furthermore, efficient dimeric intermolecular formation of hIL-10 by inhibiting interaction with Cys-rich prolamins also contributed to the enhanced formation of IL-10 bodies. Higher yield of hIL-10 produced in rice seeds is expected to have broad application in the future.
authors.affiliation: Functional Transgenic Crops Research Unit, National Institute of Agrobiological Sciences, Tsukuba, Ibaraki, Japan.
authors.forename: Lijun
authors.initials: L
authors.lastname: Yang
authors.forename: Sakiko
authors.initials: S
authors.lastname: Hirose
authors.forename: Hideyuki
authors.initials: H
authors.lastname: Takahashi
authors.forename: Taiji
authors.initials: T
authors.lastname: Kawakatsu
authors.forename: Fumio
authors.initials: F
authors.lastname: Takaiwa
mesh_terms: D019281:Dimerization; D056625:Endosperm / Q000378:metabolism; D005818:Genetic Engineering; D005983:Glutens / Q000378:metabolism*; D006801:Humans; D016753:Interleukin-10 / Q000096:biosynthesis* / Q000235:genetics; D018832:Molecular Chaperones / Q000378:metabolism; D012275:Oryza / Q000235:genetics / Q000378:metabolism*; D030821:Plants, Genetically Modified / Q000378:metabolism*; D055315:Prolamins / Q000378:metabolism*; D011994:Recombinant Proteins / Q000096:biosynthesis; D012639:Seeds / Q000378:metabolism*
chemical_list: C508609:IL10 protein, human; D018832:Molecular Chaperones; D055315:Prolamins; D011994:Recombinant Proteins; D016753:Interleukin-10; D005983:Glutens