MODULATION OF CELLULAR POLYAMINES IN TOBACCO BY TRANSFER AND EXPRESSION OF MOUSE ORNITHINE DECARBOXYLASE CDNA

In an attempt to modulate the metabolism of polyamines in plants, Agro bacterium tumefaciens strains were produced which contained either a f ull-length or a 3'-truncated mouse ornithine decarboxylase (ODC) cDNA under the control of the cauliflower mosaic virus 35S promoter. Plants of Nicotiana tabacum cv. Xanthi were used for transformation with the se two strains of Agrobacterium. Transformations were confirmed by Sou thern hybridization and amplification by polymerase chain reaction. Tw o plants containing the full-length cDNA (ODC-12 and ODC-30) and two c ontaining the truncated cDNA (12701-2 and 12701-31) were selected for further experiments. Northern blot analysis indicated that transcripti on was occurring and western blot analysis detected a polypeptide of c a. 50 kDa that was unique to the plants transformed with truncated ODC cDNA. In order to distinguish between the native and the mouse ODC in the transformed tissues, enzyme activity was assayed at pH optima for the two enzymes, i.e. pH 8.2 and 6.8, respectively. Substantially hig her levels of ODC activity were seen at pH 6.8 (optimum for mouse ODC) in the transformants as compared to the controls. This ODC activity w as inhibited by alpha-difluoromethylornithine and anti-mouse ODC antis era in a manner consistent with that reported for the mouse ODC. Analy sis of cellular polyamines showed significantly elevated levels (4-12- fold) of putrescine in callus derived from transformed plant tissues a s compared to the controls. The modulation of polyamine biosynthesis i n plants by these techniques should allow us to further analyze the ro le of these ubiquitous compounds in plant growth and development.