Structure of two Solanum bulbocastanum polyubiquitin genes and expression of their promoters in transgenic potatoes

Two polyubiquitin genes, bul409 and bul427, were isolated from a Solanum bulbocastanum BAC library. The bul409 and bul427 genes encode hexameric and heptameric polyproteins, respectively. bul427 exhibits a number of features suggestive of a pseudogene: (1) The last ubiquitin monomer of bul427 is interrupted by a frame shift mutation. (2) The coding sequence is flanked 3' by mitochondrial and chloroplast sequences and 5' by a protein kinase pseudogene. However, characterization of cDNAs amplified using bul427-based primers demonstrated that bul427 is transcriptionally active. Chimeric transgenes encoding beta-glucuronidase (GUS) translationally fused to the first ubiquitin-coding units of bul409, a truncated version 409s, and bul427 were constructed and introduced into potato. In transgenic potato lines both S. bulbocastanum promoters were weakly transcribed in tubers and efficiently transcribed in leaves. In leaves, bul409s was wound-induced, while bul427 was not. In tubers both promoters were wound-induced. In unwounded leaves and tubers, the steady state mRNA levels from both bul promoters were lower than the steady state mRNA levels from the Cauliflower Mosaic Virus 35S promoter. However, in the leaves and tubers of many of the transgenic lines the GUS activity was significantly higher in the bul lines compared to the 35S lines. The apparent inconsistency of higher enzymatic activity correlated with lower steady state levels of mRNA demonstrates the enhanced protein expression observed with ubiquitin fusion proteins.