Eukaryotic protein production in designed storage organelles.
BACKGROUND
Protein bodies (PBs) are natural endoplasmic reticulum (ER) or vacuole plant-derived organelles that stably accumulate large amounts of storage proteins in seeds. The proline-rich N-terminal domain derived from the maize storage protein gamma zein (Zera) is sufficient to induce PBs in non-seed tissues of Arabidopsis and tobacco. This Zera property opens up new routes for high-level accumulation of recombinant proteins by fusion of Zera with proteins of interest. In this work we extend the advantageous properties of plant seed PBs to recombinant protein production in useful non-plant eukaryotic hosts including cultured fungal, mammalian and insect cells.


RESULTS
Various Zera fusions with fluorescent and therapeutic proteins accumulate in induced PB-like organelles in all eukaryotic systems tested: tobacco leaves, Trichoderma reesei, several mammalian cultured cells and Sf9 insect cells. This accumulation in membranous organelles insulates both recombinant protein and host from undesirable activities of either. Recombinant protein encapsulation in these PBs facilitates stable accumulation of proteins in a protected sub-cellular compartment which results in an enhancement of protein production without affecting the viability and development of stably transformed hosts. The induced PBs also retain the high-density properties of native seed PBs which facilitate the recovery and purification of the recombinant proteins they contain.


CONCLUSIONS
The Zera sequence provides an efficient and universal means to produce recombinant proteins by accumulation in ER-derived organelles. The remarkable cross-kingdom conservation of PB formation and their biophysical properties should have broad application in the manufacture of non-secreted recombinant proteins and suggests the existence of universal ER pathways for protein insulation.
Torrent
Margarita
M
Departament de Genètica Molecular, Consorci CSIC-IRTA, Jordi Girona, Barcelona, Spain. mtqgmd@cid.csic.es
Llompart
Blanca
B
Lasserre-Ramassamy
Sabine
S
Llop-Tous
Immaculada
I
Bastida
Miriam
M
Marzabal
Pau
P
Westerholm-Parvinen
Ann
A
Saloheimo
Markku
M
Heifetz
Peter B
PB
Ludevid
M Dolors
MD
D000818:Animals; D001709:Biotechnology / Q000379:methods; D016466:CHO Cells; D002460:Cell Line; D006224:Cricetinae; D003412:Cricetulus; D005057:Eukaryotic Cells / Q000378:metabolism*; D005786:Gene Expression Regulation; D015388:Organelles / Q000378:metabolism*; D010940:Plant Proteins / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism*; D014026:Nicotiana / Q000235:genetics / Q000378:metabolism; D014242:Trichoderma / Q000235:genetics / Q000378:metabolism; D003313:Zea mays / Q000235:genetics / Q000378:metabolism; D015028:Zein / Q000235:genetics / Q000378:metabolism*
D010940:Plant Proteins; D011994:Recombinant Proteins; D015028:Zein