OVER-EXPRESSION OF A TOBACCO HOMEOBOX GENE, NTH15, DECREASES THE EXPRESSION OF A GIBBERELLIN BIOSYNTHETIC GENE ENCODING GA 20-OXIDASE

Ectopic expression of the homeobox gene, NTH15 (Nicotiana tabacum home obox 15) in transgenic tobacco leads to abnormal leaf and flower morph ology, accompanied by a decrease in the content of the active gibberel lin, GA(1). Quantitative analysis of intermediates in the GA biosynthe tic pathway revealed that the step from GA(19) to GA(20) was blocked i n transgenic tobacco plants overexpressing NTH15. To investigate the r elationship between the expression of NTH15 and genes involved in GA b iosynthesis, we isolated three cDNA clones from tobacco encoding two t ypes of GA 20-oxidase and a 3 beta-hydroxylase. RNA gel blot analysis revealed that the expression of one gene (Ntc12, encoding GA 20-oxidas e), which in wild-type tobacco plants was abundantly expressed in leav es, was strongly suppressed in the transformants. The expression level of Ntc12 decreased with increasing severity of phenotype of transgeni c tobacco leaves. The abnormal leaf morphology was largely overcome by treatment with GA(20) or GA1 but not by GA(19). These data strongly s uggest that overexpression of NTH15 inhibits the expression of Ntc12, resulting in reduced levels of active GA and abnormal leaf morphology in transgenic tobacco plants. In situ hybridization in wild-type tobac co revealed that expression of Ntc12 occurred mainly in the rib merist em, cells surrounding the procambium and in leaf primordia. Expression was not seen in the tunica, corpus and procambium, tissues in which N TH15 was predominantly expressed. The contrasting expression patterns of these genes may reflect their antagonistic functions in the formati on of lateral organs from the shoot apical meristem.