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GrainGenes Reference Report: PNA-98-11438

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Reference	PNA-98-11438
Title	Expression of the REB transcriptional activator in rice grains improves the yield of recombinant proteins whose genes are controlled by a Reb-responsive promoter
Journal	Proceedings of the National Academy of Sciences, USA
Year	2001
Volume	98
Pages	11438-11443
Author	Yang D Wu L Hwang YS Chen L Huang N
Abstract	The gene encoding the rice transcription factor, REB (rice endosperm bZIP) was cloned from a bacterial artificial chromosome library of rice. The cloned 6,227-bp-long Reb gene is composed of six exons and five introns and is flanked by a 1.2-kb 5' promoter and a 1.2-kb 3' terminator region. The function of the Reb gene was explored by a transient assay by using a rice immature endosperm system. The effector constructs containing the native gene or fusion genes linking Reb to the rice actin (Act) or globulin (Glb) gene promoters and the reporter gene construct Glb-beta-glucuronidase (GUS) were used in this study. When these effector constructs were cotransferred with the reporter uidA gene encoding GUS under the control of the Glb promoter into immature rice endosperm cells, the Glb promoter was activated. The transient GUS expression was 2.0 to 2.5-fold higher with the effector construct than without. When the upstream activation sequence containing the GCCACGT(A/C)AG motifs of the Glb promoter was deleted, the activation by REB was abolished. On the other hand, a gain-of-function experiment showed that inserting the upstream activation sequence into the glutelin-1 (Gt1) promoter made it responsive to activation by REB. When cotransformed with Reb gene, mature transgenic rice grains containing the human lysozyme gene driven by the Glb promoter produced 3.7-fold more lysozyme. Accumulation of recombinant lysozyme in mature seed ranged from 30.57 to 279.61 microgram(.)mg-1 total soluble protein in individual transformants from 30 independent transformation events. Thus, our results show that REB is not only a transcriptional activator, it can also be used to increase the expression of recombinant protein in transgenic rice grains
Keyword	[ Hide all but 1 of 47 ] accumulation activation assay bacterial beta glucuronidase cell chromosome endosperm exons expression function fusion gene gene expression gene promoter globulins grain gus gus expression human increase introns libraries library lysozyme motif nucleotide sequences oryza sativa promoter protein recombinant protein region regulatory genes reporter reporter genes rice seed seeds sequence system transcription transformation transgenic transgenic plants transient uida yield

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