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Overexpression of two PsnAP1 genes from Populus simonii × P. nigra causes early flowering in transgenic tobacco and Arabidopsis.

Zheng T, Li S, Zang L, Dai L, Yang C, Qu GZ - PLoS ONE (2014)

Bottom Line: Overexpression of PsnAP1-1 and PsnAP1-2 in tobacco under the control of a CaMV 35S promoter significantly enhanced early flowering.These transgenic plants also showed much earlier stem initiation and higher rates of photosynthesis than did wild-type tobacco. qRT-PCR analysis further indicated that overexpression of PsnAP1-1 and PsnAP1-2 resulted in up-regulation of genes related to flowering, such as NtMADS4, NtMADS5 and NtMADS11.Overexpression of PsnAP1-1 and PsnAP1-2 in Arabidopsis also induced early flowering, but did not complement the ap1-10 floral morphology to any noticeable extent.

View Article: PubMed Central - PubMed

Affiliation: State Key Laboratory of Tree Genetics and Breeding (Northeast Forestry University), Harbin, China.

ABSTRACT
In Arabidopsis, AP1 is a floral meristem identity gene and plays an important role in floral organ development. In this study, PsnAP1-1 and PsnAP1-2 were isolated from the male reproductive buds of poplar (Populus simonii × P. nigra), which are the orthologs of AP1 in Arabidopsis, by sequence analysis. Northern blot and qRT-PCR analysis showed that PsnAP1-1 and PsnAP1-2 exhibited high expression level in early inflorescence development of poplar. Subcellular localization showed the PsnAP1-1 and PsnAP1-2 proteins are localized in the nucleus. Overexpression of PsnAP1-1 and PsnAP1-2 in tobacco under the control of a CaMV 35S promoter significantly enhanced early flowering. These transgenic plants also showed much earlier stem initiation and higher rates of photosynthesis than did wild-type tobacco. qRT-PCR analysis further indicated that overexpression of PsnAP1-1 and PsnAP1-2 resulted in up-regulation of genes related to flowering, such as NtMADS4, NtMADS5 and NtMADS11. Overexpression of PsnAP1-1 and PsnAP1-2 in Arabidopsis also induced early flowering, but did not complement the ap1-10 floral morphology to any noticeable extent. This study indicates that PsnAP1-1 and PsnAP1-2 play a role in floral transition of poplar.

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Microscopic observation of tobacco.(A) The seedlings from 25-day-old wild-type and transgenic tobacco plants. (B, C, D) The histological observation of apical shoots from 25-day-old wild-type (B), 35S::PsnAP1-1 (C), and 35S::PsnAP1-2 (D) tobacco seedlings. (E, F, G) Hand-cut sections of 25-day-old wild-type (E), 35S::PsnAP1-1 (F), and 35S::PsnAP1-2 (G) tobacco stems were treated with phloroglucinol-HCl for lignin staining. (H, I, J) Histological observation of 25-day-old wild-type (H), 35S::PsnAP1-1 (I), and 35S::PsnAP1-2 (J) tobacco stems were treated with Safranin T and toluidine blue staining. WT wild-type, PsnAP1-1 35S::PsnAP1-1 line, PsnAP1-2 35S::PsnAP1-2 line.
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pone-0111725-g008: Microscopic observation of tobacco.(A) The seedlings from 25-day-old wild-type and transgenic tobacco plants. (B, C, D) The histological observation of apical shoots from 25-day-old wild-type (B), 35S::PsnAP1-1 (C), and 35S::PsnAP1-2 (D) tobacco seedlings. (E, F, G) Hand-cut sections of 25-day-old wild-type (E), 35S::PsnAP1-1 (F), and 35S::PsnAP1-2 (G) tobacco stems were treated with phloroglucinol-HCl for lignin staining. (H, I, J) Histological observation of 25-day-old wild-type (H), 35S::PsnAP1-1 (I), and 35S::PsnAP1-2 (J) tobacco stems were treated with Safranin T and toluidine blue staining. WT wild-type, PsnAP1-1 35S::PsnAP1-1 line, PsnAP1-2 35S::PsnAP1-2 line.

Mentions: In transgenic lines, pleiotropic phenotypes other than early flowering induction were also induced by the overexpression of PsnAP1-1 and PsnAP1-2. As shown in Fig. 8A, stem initiation took place much earlier in transgenic plants (25 d seedlings) than that in wild type. Histological analysis confirmed that flower transition occurred earlier in the shoot apical meristem of transgenic tobacco than that in wild type. The shoot apex of control plants was still in vegetative stage 25 d after germination under long-day conditions, and no inflorescence meristems were present (Fig. 8B), whereas, in transgenic plants, the inflorescence meristem was already formed at this stage (Fig. 8C, D). We next used phloroglucinol-HCl staining to estimate lignification, as phloroglucinol-HCl reacts with coniferaldehyde groups in lignin, and the area and color intensity roughly reflects total lignin content [21]. The stems of transgenic tobacco plants were much more intensely stained than those of wild-type plants (Fig. 8E, F, G). Cell wall lignification is a complex process. It occurs exclusively in higher plants. Its main function is to strengthen the plant vascular body. Safranine T was also used to stain plant tissue sections for lignification of the cell walls. The results of Safranine T staining also indicated that the stems of transgenic tobacco plants were slightly more intensely stained than those of wild type (Fig. 8H, I, J). These staining results suggested that overexpression of PsnAP1-1 and PsnAP1-2 may accelerate lignification of cell wall in transgenics.


Overexpression of two PsnAP1 genes from Populus simonii × P. nigra causes early flowering in transgenic tobacco and Arabidopsis.

Zheng T, Li S, Zang L, Dai L, Yang C, Qu GZ - PLoS ONE (2014)

Microscopic observation of tobacco.(A) The seedlings from 25-day-old wild-type and transgenic tobacco plants. (B, C, D) The histological observation of apical shoots from 25-day-old wild-type (B), 35S::PsnAP1-1 (C), and 35S::PsnAP1-2 (D) tobacco seedlings. (E, F, G) Hand-cut sections of 25-day-old wild-type (E), 35S::PsnAP1-1 (F), and 35S::PsnAP1-2 (G) tobacco stems were treated with phloroglucinol-HCl for lignin staining. (H, I, J) Histological observation of 25-day-old wild-type (H), 35S::PsnAP1-1 (I), and 35S::PsnAP1-2 (J) tobacco stems were treated with Safranin T and toluidine blue staining. WT wild-type, PsnAP1-1 35S::PsnAP1-1 line, PsnAP1-2 35S::PsnAP1-2 line.
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pone-0111725-g008: Microscopic observation of tobacco.(A) The seedlings from 25-day-old wild-type and transgenic tobacco plants. (B, C, D) The histological observation of apical shoots from 25-day-old wild-type (B), 35S::PsnAP1-1 (C), and 35S::PsnAP1-2 (D) tobacco seedlings. (E, F, G) Hand-cut sections of 25-day-old wild-type (E), 35S::PsnAP1-1 (F), and 35S::PsnAP1-2 (G) tobacco stems were treated with phloroglucinol-HCl for lignin staining. (H, I, J) Histological observation of 25-day-old wild-type (H), 35S::PsnAP1-1 (I), and 35S::PsnAP1-2 (J) tobacco stems were treated with Safranin T and toluidine blue staining. WT wild-type, PsnAP1-1 35S::PsnAP1-1 line, PsnAP1-2 35S::PsnAP1-2 line.
Mentions: In transgenic lines, pleiotropic phenotypes other than early flowering induction were also induced by the overexpression of PsnAP1-1 and PsnAP1-2. As shown in Fig. 8A, stem initiation took place much earlier in transgenic plants (25 d seedlings) than that in wild type. Histological analysis confirmed that flower transition occurred earlier in the shoot apical meristem of transgenic tobacco than that in wild type. The shoot apex of control plants was still in vegetative stage 25 d after germination under long-day conditions, and no inflorescence meristems were present (Fig. 8B), whereas, in transgenic plants, the inflorescence meristem was already formed at this stage (Fig. 8C, D). We next used phloroglucinol-HCl staining to estimate lignification, as phloroglucinol-HCl reacts with coniferaldehyde groups in lignin, and the area and color intensity roughly reflects total lignin content [21]. The stems of transgenic tobacco plants were much more intensely stained than those of wild-type plants (Fig. 8E, F, G). Cell wall lignification is a complex process. It occurs exclusively in higher plants. Its main function is to strengthen the plant vascular body. Safranine T was also used to stain plant tissue sections for lignification of the cell walls. The results of Safranine T staining also indicated that the stems of transgenic tobacco plants were slightly more intensely stained than those of wild type (Fig. 8H, I, J). These staining results suggested that overexpression of PsnAP1-1 and PsnAP1-2 may accelerate lignification of cell wall in transgenics.

Bottom Line: Overexpression of PsnAP1-1 and PsnAP1-2 in tobacco under the control of a CaMV 35S promoter significantly enhanced early flowering.These transgenic plants also showed much earlier stem initiation and higher rates of photosynthesis than did wild-type tobacco. qRT-PCR analysis further indicated that overexpression of PsnAP1-1 and PsnAP1-2 resulted in up-regulation of genes related to flowering, such as NtMADS4, NtMADS5 and NtMADS11.Overexpression of PsnAP1-1 and PsnAP1-2 in Arabidopsis also induced early flowering, but did not complement the ap1-10 floral morphology to any noticeable extent.

View Article: PubMed Central - PubMed

Affiliation: State Key Laboratory of Tree Genetics and Breeding (Northeast Forestry University), Harbin, China.

ABSTRACT
In Arabidopsis, AP1 is a floral meristem identity gene and plays an important role in floral organ development. In this study, PsnAP1-1 and PsnAP1-2 were isolated from the male reproductive buds of poplar (Populus simonii × P. nigra), which are the orthologs of AP1 in Arabidopsis, by sequence analysis. Northern blot and qRT-PCR analysis showed that PsnAP1-1 and PsnAP1-2 exhibited high expression level in early inflorescence development of poplar. Subcellular localization showed the PsnAP1-1 and PsnAP1-2 proteins are localized in the nucleus. Overexpression of PsnAP1-1 and PsnAP1-2 in tobacco under the control of a CaMV 35S promoter significantly enhanced early flowering. These transgenic plants also showed much earlier stem initiation and higher rates of photosynthesis than did wild-type tobacco. qRT-PCR analysis further indicated that overexpression of PsnAP1-1 and PsnAP1-2 resulted in up-regulation of genes related to flowering, such as NtMADS4, NtMADS5 and NtMADS11. Overexpression of PsnAP1-1 and PsnAP1-2 in Arabidopsis also induced early flowering, but did not complement the ap1-10 floral morphology to any noticeable extent. This study indicates that PsnAP1-1 and PsnAP1-2 play a role in floral transition of poplar.

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