# Gene transcriptions/Boxes/Pyrimidines

"Functional analyses of a number of hydrolase gene promoters, induced by gibberellin (GA) in aleurone cells following germination, have identified a GA-responsive complex as a tripartite element containing a pyrimidine box motif 5′-CCTTTT-3′."
"The pyrimidine box is another promoter element that is observed in cereal GA-responsive promoters examined thus far (Huang et al., 1990). Mutations on the pyrimidine box caused the reduction of the GA induction with a lesser magnitude than those of GARE in the GA-treated aleurone (Gubler and Jacobsen, 1992; Rogers and Rogers, 1992). The pyrimidine box alone could not confer the hormone responsiveness to the minimal promoter of the cauliflower mosaic virus 35S (CaMV35S), suggesting accessory roles of the pyrimidine box on the transcription response to GA (Skriver et al., 1991)."
"[A]leurone proteins that recognized the pyrimidine box sequence [are] from barley (BPBF; Mena et al., 2002) and rice (Oryza sativa; OsDOF3; Washio, 2001)."
"A member of Dof proteins, prolamine box-binding factors PBF that were originally identified to be transcription factors regulating the expression of genes for stored proteins in developing seeds (Vicente-Carbajosa et al., 1997; Mena et al., 1998), BPBF and OsDOF3 are likely to be a pyrimidine box-binding protein in the germinated aleurone."
"OsDOF3, binding the pyrimidine box, affected the DNA binding of GAMYB to GARE".
"Functional promoter analysis using transgenic rice seeds has confirmed that the 5'-regulatory region extending from -232 (nucleotide position relative to the site for transcription initiation) to +31 is sufficient for hormonal regulation by GA (Itoh et al., 1995), along which are found the distal (Pyr-1, -312) and proximal pyrimidine boxes (Pyr-2, -214), GARE (-148), and three potential sites for the Dof binding (D-1, -191; D-2, -109; D-3, -78; [on the diagram in the GA responsive complexes section above]; Huang et al., 1990)."
"The upstream portion from the RAmy1A promoter (-380 to +27) had a stimulatory effect on the GA-induced expression of the reporter gene in transfected aleurone cells."
"The drastic loss of the GA induction associated with a mutation on GARE (M4) further verifies the importance of this motif in the GA-regulated expression of genes. Other mutations of the proximal pyrimidine box (M2) and one site for the Dof binding (M5) also reduced the GA-induced activities to 48% and 54% of the wild-type sequence, respectively, but the effects were not as marked as seen when GARE was mutated."
"Sequence analysis shows that the OsDof3 cDNA encodes a 371-amino acid polypeptide related to the PBF factors of cereal plants. The predicted amino acid sequence of OsDOF3 aligns well with those of maize (Zea mays; Vicente-Carbajosa et al., 1997), wheat (Triticum aestivum), and barley PBF proteins (Mena et al., 1998; [...]). The N-terminal sequence of OsDOF3 contains four Cys residues reminiscent of the Dof zinc finger and shows around 80% sequence identities with PBFs, whereas the C-terminal parts are divergent showing several insertions and deletions."
"Pentanucleotide sequence from the pyrimidine box (CTTTT; Huang et al., 1990) and D-3 (AAAAG) is capable of matching a favored substrate selected by in vitro DNA binding of maize Dof proteins (CTTTT or AAAAG; Yanagisawa and Schmidt, 1999). These results indicate preferential binding of OsDOF3 to two pyrimidine box and D-3 motifs in the RAmy1A promoter context."

## Gibberellic acid responsive complexes

"Although this GARC [GA responsive complex] may not always be tripartite, most often it includes three sequence motifs, the TAACAAA box or GA responsive element (GARE), the pyrimidine box CCTTTT, and the TATCCAC box (Skriver et al., 1991;Gubler and Jacobsen, 1992; Rogers et al., 1994)."
"Some other cis-acting elements, such as pyrimidine boxes (GGTTTT) and TAT boxes (TATCCAT), are usually present in the vicinity of the GARE sequence of genes regulated by GA in cereal aleurone cells (Gubler and Jacobsen 1992; Cercos et al. 1999; Tsuji et al. 2006)."
"Given the general observations that a couple of the promoter motifs, GARE and the pyrimidine box, always exist in a close distance in the reported GA-responsive promoters (Huang et al., 1990), the synergistic effect on RAmy1A promoter activation are likely to be mediated through protein-protein interaction."

## P-boxes

The "complementary strand of the pyrimidine box element (5′-CTTTT-3′) in GA-induced hydrolase gene promoters was identical to the core sequence (5′-AAAAG-3′) recognized by PBF in prolamin gene promoters (P-box: 5′-T/AAAAG-3′; Vicente-Carbajosa et al., 1997; Mena et al., 1998; Yanagisawa and Schmidt, 1999)".

## Sampling of A1BG promoters

For the Basic programs (starting with SuccessablesPyr.bas) written to compare nucleotide sequences with the sequences on either the template strand (-), or coding strand (+), of the DNA, in the negative direction (-), or the positive direction (+), including extending the number of nts from 958 to 4445, the programs are, are looking for, and found:
* negative strand in the negative direction (from ZSCAN22 to A1BG) is SuccessablesPyr--.bas, looking for 3'-CCTTTT-5', 3, 3'-CCTTTT-5', 2459, 3'-CCTTTT-5', 2927, 3'-CCTTTT-5', 2968,
* negative strand in the positive direction (from ZNF497 to A1BG) is SuccessablesPyr-+.bas, looking for 3'-CCTTTT-5', 1, 3'-CCTTTT-5', 135,
* positive strand in the negative direction is SuccessablesPyr+-.bas, looking for 3'-CCTTTT-5', 0,
* positive strand in the positive direction is SuccessablesPyr++.bas, looking for 3'-CCTTTT-5', 1, 3'-CCTTTT-5', 291,
* complement, negative strand, negative direction is SuccessablesPyrc--.bas, looking for 3'-GGAAAA-5', 0,
* complement, negative strand, positive direction is SuccessablesPyrc-+.bas, looking for 3'-GGAAAA-5', 1, 3'-GGAAAA-5', 291,
* complement, positive strand, negative direction is SuccessablesPyrc+-.bas, looking for 3'-GGAAAA-5', 3, 3'-GGAAAA-5', 2459, 3'-GGAAAA-5', 2927, 3'-GGAAAA-5', 2968,
* complement, positive strand, positive direction is SuccessablesPyrc++.bas, looking for 3'-GGAAAA-5', 1, 3'-GGAAAA-5', 135,
* inverse complement, negative strand, negative direction is SuccessablesPyrci--.bas, looking for 3'-AAAAGG-5', 0,
* inverse complement, negative strand, positive direction is SuccessablesPyrci-+.bas, looking for 3'-AAAAGG-5', 0,
* inverse complement, positive strand, negative direction is SuccessablesPyrci+-.bas, looking for 3'-AAAAGG-5', 4, 3'-AAAAGG-5', 105, 3'-AAAAGG-5', 1107, 3'-AAAAGG-5', 3345, 3'-AAAAGG-5', 3441,
* inverse complement, positive strand, positive direction is SuccessablesPyrci++.bas, looking for 3'-AAAAGG-5', 0,
* inverse, negative strand, negative direction, is SuccessablesPyri--.bas, looking for 3'-TTTTCC-5', 4, 3'-TTTTCC-5', 105, 3'-TTTTCC-5', 1107, 3'-TTTTCC-5', 3345, 3'-TTTTCC-5', 3441,
* inverse, negative strand, positive direction, is SuccessablesPyri-+.bas, looking for 3'-TTTTCC-5', 0,
* inverse, positive strand, negative direction, is SuccessablesPyri+-.bas, looking for 3'-TTTTCC-5', 0,
* inverse, positive strand, positive direction, is SuccessablesPyri++.bas, looking for 3'-TTTTCC-5', 0.