SeeDev-binary@ldeleger:SeeDev-binary-19719479-4 / 1634-1651
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Functional dissections of the BCCP2 and PKp-β1 promoters have led to the identification of enhancing regions of 54 and 79 bp, respectively, that are essential to direct the activity of these promoters in oil-accumulating tissues of the embryo. Comparison of these enhancer elements using PLACE reveals the presence of shared nucleotide sequences that may constitute putative recognition sequences bound by the transcriptional machinery triggering fatty acid biosynthesis. The hexanucleotide AACCCA that can be observed on the minus strand of each enhancer element has previously been described as the core of the SEF3-recognition sequence (Allen et al., 1989). This DNA binding factor was identified in nuclear extracts from developing soybean seeds, and was initially presented as a positive regulator of transcription of the β-conglycinin, α-subunit gene. SEF3-like activities were reported in seeds of other oleaginous species, such as tobacco and sunflower (Lessard et al., 1991). In soybean, SEF3 was shown to bind specifically to a site composed of two AACCCA elements separated by 27 bp (Allen et al., 1989). Nevertheless, complementary studies then established that mutations abolishing the protein binding of this SEF3 factor in vitro did not affect the in vivo promoter activity in a significant manner (Fujiwara and Beachy, 1994). Similarly, in this study, we show that mutations in the AACCCA element of the BCCP2 promoter (m2; Figure 5b) do not impact on promoter activity (Figure 6). These results strongly suggest that isolated core SEF3 recognition sequences do not constitute essential cis-regulatory elements of the lipogenic pathway.
By carefully analysing the results of the functional dissections carried out on the enhancer regions of the BCCP2 and PKp-β1 promoters, it is possible to identify a DNA sequence highly conserved between the two promoters that is important for their activity (Figure 5).
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