Functional characterization of a reproductive tissue specific promoter from Eucalyptus camaldulensis

SWEET proteins are essential for the maintenance of nectar production, as well as seed and pollen development, in plants. A search within the Eucalyptus genome identified 52 putative genes belonging to the SWEET gene family based on sequence similarity. The expression of two of these genes, EcSWEET2 and EcSWEET5, was analyzed in vegetative and reproductive tissues of Eucalyptus camaldulensis. The expression of EcSWEET5 was specific to male reproductive tissues, and transcripts were detected only at certain stages of flower development. Tobacco Rattle Virus (TRV)-mediated suppression of EcSWEET5 resulted in a significant reduction in pollen germination percentage in Nicotiana benthamiana without adverse effect on vegetative growth. A promoter sequence 1 kb upstream of the start codon of EcSWEET5 contained many elements suggestive of pollen specificity of the promoter. This specificity was confirmed in transgenic tobacco lines harboring a GUS gene whose expression was controlled by the EcSWEET5 gene promoter. GUS expression was limited to pollen alone in transgenic tobacco as evidenced by histochemical staining. The expression of a cytotoxic gene, barnase under the control of the EcSWEET5 gene promoter, showed pollen ablation in transgenic tobacco with normal vegetative growth.