Biologically active human GM-CSF produced in the seeds of transgenic rice plants

Sardana, Ravinder; Dudani, Anil K.; Tackaberry, Eilleen S.; Alli, Zaman; Porter, Suzanne; Rowlandson, Karen; Ganz, Peter; Altosaar, Illimar

doi:10.1007/s11248-006-9062-y

Cited by 42 publications

(22 citation statements)

References 40 publications

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“…However, the expression level is still not high enough to be economically viable. We have successfully obtained a very high expression level (14 lg/seed) of OsrhGM-CSF in rice endosperm, which is the highest expression level reported in any plant system (Sardana et al 2007). To improve the expression level of OsrhGM-CSF in rice endosperm cells, we chose a stronger, endosperm-specific promoter and rice-preferred codons to improve the transcription and translation efficiency, respectively.…”

Section: Discussionmentioning

confidence: 75%

Oral administration of recombinant human granulocyte-macrophage colony stimulating factor expressed in rice endosperm can increase leukocytes in mice

et al. 2008

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Human granulocyte-macrophage colony stimulating factor (hGM-CSF) is used clinically to treat leucopenia typically caused by cancer chemotherapy or radiotherapy. This study used multiple strategies to obtain very high expression levels of OsrhGM-CSF (14 microg/seed) in rice endosperm. Electron micrographs of immunogold-labeled transgenic endosperm showed that rhGM-CSF was not only localized in protein bodies but was also distributed in the apoplast. A biological activity assay indicated that OsrhGM-CSF stimulated the growth of TF-1 cells in vitro. In addition, the transgene was used to effectively treat leucopenia by oral administration of the unprocessed transgenic grains. In cyclophosphamide-induced leucopenic mice, transgenic seeds produced a 27% (t=0.021) gain in leukocytes after 14 days feeding. Even in non-leucopenic mice, leukocyte gain was 37% (t=0.002) more than that of mice fed non-transgenic seeds. This study provides a novel approach to the use of oral unprocessed transgenic OsrhGM-CSF seeds to treat leucopenia.

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Section: Discussionmentioning

confidence: 75%

Oral administration of recombinant human granulocyte-macrophage colony stimulating factor expressed in rice endosperm can increase leukocytes in mice

et al. 2008

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“…Glycosylation at one or more of these sites in the protein would affect its mobility on the gel, explaining the higher molecular weight protein bands observed on the gel. Furthermore, heterogeneous glycosylation has been observed for expressed growth factors in other plant systems like tobacco [16] and rice seeds [17]. In contrast, E. coli produced Flt3 ligand presents a single band in a Coomassie brilliant blue-stained SDS-PAGE (Fig.…”

Section: Resultsmentioning

confidence: 99%

Barley as a green factory for the production of functional Flt3 ligand

Erlendsson¹,

Muench

Hellman

et al. 2010

Biotechnology Journal

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Biologically active recombinant human Flt3 ligand was expressed and isolated from transgenic barley seeds. Its expression is under the control of a tissue specific promoter that confines accumulation of the recombinant protein to the endosperm tissue of the seed. The recombinant Flt3 ligand variant expressed in the seeds contains a HQ-tag for affinity purification on IMAC resin. Using sandwich enzyme-linked immunosorbent assay seventy-three individual barley lines were analyzed for expression. Seventeen lines accumulated threshold amount of the protein in the seeds. The tagged protein was purified from seed extract to near homogeneity using sequential chromatography on IMAC affinity resin and cation exchange resin. The barley expressed recombinant Flt3 ligand migrates as two bands of 20 kDa and 22 kDa in SDS-PAGE which is slightly bigger than the 19 kDa predicted from the sequence. This indicates that post-translational modifications occur in the plant. We show that the recombinant Flt3 ligand protein is a glycoprotein containing α-1,3-fucose and α-1,2-xylose. The HQ-tagged Flt3 ligand variant exhibits comparable biological activity to commercial Flt3 ligand. This is the first report showing expression and accumulation of recombinant human growth factor in barley seeds with a yield of active protein similar to bacterial expression system. The present results demonstrate that plant molecular farming is viable approach for the bioproduction of human derived growth factors.

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“…The relative gB concentrations varied over a range of about 5-fold, reflecting heterogeneous expression patterns that are quite typical for transgenic plant systems [13,17,54]. After rice lines homozygous for gB were developed, 14 second-generation plants again showed a range of gB concentrations.…”

Section: Discussionmentioning

confidence: 99%

Sustained Expression of Human Cytomegalovirus Glycoprotein B (UL55) in the Seeds of Homozygous Rice Plants

et al. 2008

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Production of recombinant subunit vaccines in transgenic plants may be a means of reducing vaccine costs while increasing availability and safety. A plant-derived product found safe and effective for oral administration would provide additional advantages when used as a vaccine. Outstanding issues with the technology include transgene stability through successive generations and consistent bioproduction. We previously reported expression of glycoprotein B (gB) of human cytomegalovirus in seeds of transgenic tobacco. Here the goal was to determine if gB could be similarly expressed in rice, and if so, to examine expression over several plant generations. Results show that immunoreactive gB was successfully expressed in transgenic rice seeds, with sustained expression over three generations. The gB contained several neutralizing epitopes and was stable over 27 months.

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Biologically active human GM-CSF produced in the seeds of transgenic rice plants

Cited by 42 publications

References 40 publications

Oral administration of recombinant human granulocyte-macrophage colony stimulating factor expressed in rice endosperm can increase leukocytes in mice

Oral administration of recombinant human granulocyte-macrophage colony stimulating factor expressed in rice endosperm can increase leukocytes in mice

Barley as a green factory for the production of functional Flt3 ligand

Sustained Expression of Human Cytomegalovirus Glycoprotein B (UL55) in the Seeds of Homozygous Rice Plants

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