OVEREXPRESSION OF DELTA-EMBP, A TRUNCATED DOMINANT-NEGATIVE VERSION OF THE WHEAT G-BOX BINDING-PROTEIN EMBP-1, ALTERS VEGETATIVE DEVELOPMENT IN TRANSGENIC TOBACCO

As a first step toward elucidating the in vivo function of plant bZIP proteins and their related G-box cis elements, we have introduced a do minant negative inhibitor of G-box-dependent transcriptional activatio n into tobacco plants by transforming them with a truncated EmBP-1 gen e (Delta EmBP) containing the DNA binding and dimerization domains und er the control of the CaMV 35S promoter. Five independent lines of tra nsgenic plants expressing Delta EmBP were identified, as demonstrated by immunodetection of the transgenic protein in leaf extracts, and the ability of the protein to bind a target G-box DNA sequence. The trans genic plants exhibited an abnormal phenotype characterized by intervei nal chlorosis, growth inhibition and weakening of stems and petioles, the severity of which positively correlated with Delta EmBP expression and G-box DNA binding capability. Furthermore, development of chloros is and growth inhibition was dependent on growth irradiance. Low light promoted the development of interveinal chlorosis and growth inhibiti on in the transgenic plants, whereas high light conditions led to near -complete amelioration of the abnormal phenotype. Transgenic plants un der both light regimes showed signs of impaired stem and petiole funct ion which was not observed in wild-type tobacco. RbcS gene expression was not significantly altered by Delta EmBP expression, suggesting tha t down-regulation of this gene was not responsible for the altered phe notype. The results suggest that G-box elements specific for the EmBP- 1 class of bZIP proteins have an important developmental function in v egetative plant tissues, and that the trans-dominant negative mutant a pproach is a useful tool for continued in viva functional analysis of bZIP transcription factors and their corresponding cis elements in pla nts.