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Histone acetylation accompanied with promoter sequences displaying differential expression profiles of B-class MADS-box genes for phalaenopsis floral morphogenesis.

Hsu CC, Wu PS, Chen TC, Yu CW, Tsai WC, Wu K, Wu WL, Chen WH, Chen HH - PLoS ONE (2014)

Bottom Line: The amplified promoter sequences of PeMADS2∼6 could drive beta-glucuronidase (GUS) gene expression in all floral organs, similar to their expression at the floral primordia stage.The promoter sequence of PeMADS4, exclusively expressed in lip and column, showed a 1.6∼3-fold higher expression in lip/column than in sepal/petal.Furthermore, we noted a 4.9-fold increase in histone acetylation (H3K9K14ac) in the translation start region of PeMADS4 in lip as compared in petal.

View Article: PubMed Central - PubMed

Affiliation: Department of Life Sciences, National Cheng Kung University, Tainan, Taiwan.

ABSTRACT
Five B-class MADS-box genes, including four APETALA3 (AP3)-like PeMADS2∼5 and one PISTILLATA (PI)-like PeMADS6, specify the spectacular flower morphology in orchids. The PI-like PeMADS6 ubiquitously expresses in all floral organs. The four AP3-like genes, resulted from two duplication events, express ubiquitously at floral primordia and early floral organ stages, but show distinct expression profiles at late floral organ primordia and floral bud stages. Here, we isolated the upstream sequences of PeMADS2∼6 and studied the regulatory mechanism for their distinct gene expression. Phylogenetic footprinting analysis of the 1.3-kb upstream sequences of AP3-like PeMADS2∼5 showed that their promoter regions have sufficiently diverged and contributed to their subfunctionalization. The amplified promoter sequences of PeMADS2∼6 could drive beta-glucuronidase (GUS) gene expression in all floral organs, similar to their expression at the floral primordia stage. The promoter sequence of PeMADS4, exclusively expressed in lip and column, showed a 1.6∼3-fold higher expression in lip/column than in sepal/petal. Furthermore, we noted a 4.9-fold increase in histone acetylation (H3K9K14ac) in the translation start region of PeMADS4 in lip as compared in petal. All these results suggest that the regulation via the upstream sequences and increased H3K9K14ac level may act synergistically to display distinct expression profiles of the AP3-like genes at late floral organ primordia stage for Phalaenopsis floral morphogenesis.

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Promoter sequences of PeMADS2∼6 and the putative CArG boxes.Length of promoter sequences of PeMADS2, PeMADS3, and PeMADS6 were 3,249, 1,293 and 1,514 bp, respectively. The promoter sequence of the PeMADS4 was extended from −422 bp to −3,303 bp (horizontal line box), and the original fragment of −2,121 to −840 bp of PeMADS5 promoter was replaced by a 1,227-bp fragment (diagonal line box). The rhombus and the triangles indicate the putative CArG boxes predicted by consensus CC(A/T)6GG and C(A/T)8G motifs, respectively. “+1” means the transcription start site and ATG was the translation start site. The lengths of the promoters are show from the transcription start site to the upstream sequences.
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pone-0106033-g001: Promoter sequences of PeMADS2∼6 and the putative CArG boxes.Length of promoter sequences of PeMADS2, PeMADS3, and PeMADS6 were 3,249, 1,293 and 1,514 bp, respectively. The promoter sequence of the PeMADS4 was extended from −422 bp to −3,303 bp (horizontal line box), and the original fragment of −2,121 to −840 bp of PeMADS5 promoter was replaced by a 1,227-bp fragment (diagonal line box). The rhombus and the triangles indicate the putative CArG boxes predicted by consensus CC(A/T)6GG and C(A/T)8G motifs, respectively. “+1” means the transcription start site and ATG was the translation start site. The lengths of the promoters are show from the transcription start site to the upstream sequences.

Mentions: The five PeMADS promoter regions were cloned from genomic DNA by use of GenomeWalker. The PeMADS2∼6 promoter fragments obtained were 3,249, 1,293, 422, 2,121, and 1,514 bp, respectively (Fig. 1). However, the 2,121-bp promoter sequence of PeMADS5 could not be amplified from the genomic DNA of P. equestris by using PCR, and both PeMADS3 and PeMADS4 with the promoter fragments <1.5 kb could not be extended. We then used BAC clones of P. equestris for promoter identification of PeMADS3∼5 by Southern blot hybridization with probes from the coding sequences of PeMADS3∼5 (Table 1). BAC DNA was isolated and confirmed by restriction enzyme digestion. The resolved bands corresponding to the hybridized signals were recovered and sequenced. After assembly, the PeMADS3 promoter remained at 1,293 bp. For the PeMADS4 promoter, a 4.8-kb DNA fragment from BAC clones was recovered and sequenced, which extended its upstream sequence to 3,303 bp (Fig. 1, horizontal line box). For PeMADS5, the promoter fragment was replaced with a 2,062-bp fragment (Fig. 1).


Histone acetylation accompanied with promoter sequences displaying differential expression profiles of B-class MADS-box genes for phalaenopsis floral morphogenesis.

Hsu CC, Wu PS, Chen TC, Yu CW, Tsai WC, Wu K, Wu WL, Chen WH, Chen HH - PLoS ONE (2014)

Promoter sequences of PeMADS2∼6 and the putative CArG boxes.Length of promoter sequences of PeMADS2, PeMADS3, and PeMADS6 were 3,249, 1,293 and 1,514 bp, respectively. The promoter sequence of the PeMADS4 was extended from −422 bp to −3,303 bp (horizontal line box), and the original fragment of −2,121 to −840 bp of PeMADS5 promoter was replaced by a 1,227-bp fragment (diagonal line box). The rhombus and the triangles indicate the putative CArG boxes predicted by consensus CC(A/T)6GG and C(A/T)8G motifs, respectively. “+1” means the transcription start site and ATG was the translation start site. The lengths of the promoters are show from the transcription start site to the upstream sequences.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4263434&req=5

pone-0106033-g001: Promoter sequences of PeMADS2∼6 and the putative CArG boxes.Length of promoter sequences of PeMADS2, PeMADS3, and PeMADS6 were 3,249, 1,293 and 1,514 bp, respectively. The promoter sequence of the PeMADS4 was extended from −422 bp to −3,303 bp (horizontal line box), and the original fragment of −2,121 to −840 bp of PeMADS5 promoter was replaced by a 1,227-bp fragment (diagonal line box). The rhombus and the triangles indicate the putative CArG boxes predicted by consensus CC(A/T)6GG and C(A/T)8G motifs, respectively. “+1” means the transcription start site and ATG was the translation start site. The lengths of the promoters are show from the transcription start site to the upstream sequences.
Mentions: The five PeMADS promoter regions were cloned from genomic DNA by use of GenomeWalker. The PeMADS2∼6 promoter fragments obtained were 3,249, 1,293, 422, 2,121, and 1,514 bp, respectively (Fig. 1). However, the 2,121-bp promoter sequence of PeMADS5 could not be amplified from the genomic DNA of P. equestris by using PCR, and both PeMADS3 and PeMADS4 with the promoter fragments <1.5 kb could not be extended. We then used BAC clones of P. equestris for promoter identification of PeMADS3∼5 by Southern blot hybridization with probes from the coding sequences of PeMADS3∼5 (Table 1). BAC DNA was isolated and confirmed by restriction enzyme digestion. The resolved bands corresponding to the hybridized signals were recovered and sequenced. After assembly, the PeMADS3 promoter remained at 1,293 bp. For the PeMADS4 promoter, a 4.8-kb DNA fragment from BAC clones was recovered and sequenced, which extended its upstream sequence to 3,303 bp (Fig. 1, horizontal line box). For PeMADS5, the promoter fragment was replaced with a 2,062-bp fragment (Fig. 1).

Bottom Line: The amplified promoter sequences of PeMADS2∼6 could drive beta-glucuronidase (GUS) gene expression in all floral organs, similar to their expression at the floral primordia stage.The promoter sequence of PeMADS4, exclusively expressed in lip and column, showed a 1.6∼3-fold higher expression in lip/column than in sepal/petal.Furthermore, we noted a 4.9-fold increase in histone acetylation (H3K9K14ac) in the translation start region of PeMADS4 in lip as compared in petal.

View Article: PubMed Central - PubMed

Affiliation: Department of Life Sciences, National Cheng Kung University, Tainan, Taiwan.

ABSTRACT
Five B-class MADS-box genes, including four APETALA3 (AP3)-like PeMADS2∼5 and one PISTILLATA (PI)-like PeMADS6, specify the spectacular flower morphology in orchids. The PI-like PeMADS6 ubiquitously expresses in all floral organs. The four AP3-like genes, resulted from two duplication events, express ubiquitously at floral primordia and early floral organ stages, but show distinct expression profiles at late floral organ primordia and floral bud stages. Here, we isolated the upstream sequences of PeMADS2∼6 and studied the regulatory mechanism for their distinct gene expression. Phylogenetic footprinting analysis of the 1.3-kb upstream sequences of AP3-like PeMADS2∼5 showed that their promoter regions have sufficiently diverged and contributed to their subfunctionalization. The amplified promoter sequences of PeMADS2∼6 could drive beta-glucuronidase (GUS) gene expression in all floral organs, similar to their expression at the floral primordia stage. The promoter sequence of PeMADS4, exclusively expressed in lip and column, showed a 1.6∼3-fold higher expression in lip/column than in sepal/petal. Furthermore, we noted a 4.9-fold increase in histone acetylation (H3K9K14ac) in the translation start region of PeMADS4 in lip as compared in petal. All these results suggest that the regulation via the upstream sequences and increased H3K9K14ac level may act synergistically to display distinct expression profiles of the AP3-like genes at late floral organ primordia stage for Phalaenopsis floral morphogenesis.

Show MeSH