NADPH-protochlorophyllide oxidoreductase in cucumber is encoded by a single gene and its expression is transcriptionally enhanced by illumination

NADPH-protochlorophyllide oxidoreductase (POR) catalyzes the photoreduction of protochlorophyllide a in the chlorophyll biosynthetic pathway. Recently, two POR genes, the expressions of which were differently regulated by light, were isolated in barley and Arabidopsis, and it is suggested that in angiosperms, the chlorophyll synthesis is controlled by the differential action of the two distinct POR isozymes. Meanwhile, we have isolated POR cDNA from cucumber, and reported that its expression was enhanced by illumination. In this study, we analyzed the gene expression and organization of cucumber POR in more detail. Northern blot analysis with 5'- or 3'-noncoding gene-specific and full length probes and RNase protection assay revealed that cucumber POR mRNA is transcribed from a single gene. Run-on assay revealed that the transcript level of isolated nuclei from illuminated cotyledons was 6-fold that of dark control, indicating that the light-enhanced expression of cucumber POR is caused by transcriptional activation. Using degenerated primers and the genomic DNA of cucumber as template, PCR showed the presence of only one type of fragment encoding cucumber POR. The PCR product was hybridized with the genomic Southern blot from cucumber and only one band was detected under low stringency conditions. From these results, we conclude that there is a single POR gene in cucumber and PORs are organized by different gene families among higher plants, and in cucumber the single POR may play the roles of two POR genes, which are present in other higher plants such as Arabidopsis and barley.