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BcMF26a and BcMF26b Are Duplicated Polygalacturonase Genes with Divergent Expression Patterns and Functions in Pollen Development and Pollen Tube Formation in Brassica campestris.

Lyu M, Yu Y, Jiang J, Song L, Liang Y, Ma Z, Xiong X, Cao J - PLoS ONE (2015)

Bottom Line: We found that these two genes were orthologous genes of At4g33440, and they originated from a chromosomal segmental duplication.Promoter activity analysis showed that, at reproductive development stages, BcMF26b promoter was active in tapetum, pollen grains, and pistils, whereas BcMF26a promoter was only active in pistils.In the subcellular localization experiment, BcMF26a and BcMF26b proteins could be localized to the cell wall.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Cell & Molecular Biology, Institute of Vegetable Science, Zhejiang University, Hangzhou, China; Zhejiang Provincial Key Laboratory of Horticultural Plant Integrative Biology, Zhejiang University, Hangzhou, China.

ABSTRACT
Polygalacturonase (PG) is one of the cell wall hydrolytic enzymes involving in pectin degradation. A comparison of two highly conserved duplicated PG genes, namely, Brassica campestris Male Fertility 26a (BcMF26a) and BcMF26b, revealed the different features of their expression patterns and functions. We found that these two genes were orthologous genes of At4g33440, and they originated from a chromosomal segmental duplication. Although structurally similar, their regulatory and intron sequences largely diverged. QRT-PCR analysis showed that the expression level of BcMF26b was higher than that of BcMF26a in almost all the tested organs and tissues in Brassica campestris. Promoter activity analysis showed that, at reproductive development stages, BcMF26b promoter was active in tapetum, pollen grains, and pistils, whereas BcMF26a promoter was only active in pistils. In the subcellular localization experiment, BcMF26a and BcMF26b proteins could be localized to the cell wall. When the two genes were co-inhibited, pollen intine was formed abnormally and pollen tubes could not grow or stretch. Moreover, the knockout mutants of At4g33440 delayed the growth of pollen tubes. Therefore, BcMF26a/b can participate in the construction of pollen wall by modulating intine information and BcMF26b may play a major role in co-inhibiting transformed plants.

No MeSH data available.


Related in: MedlinePlus

Viability percentage and irregular percentage of pollen grains in bcmf26a/b and control lines.Nearly half of pollen grains were nonviable in the bcmf26a/b plants, whereas the percentage was only 1.01% in the control. Approximately 38.9%–71.1% of the pollen grains were irregular in the bcmf26a/b plants, whereas ~3.75% was irregular in the control. Standard errors are shown.
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pone.0131173.g008: Viability percentage and irregular percentage of pollen grains in bcmf26a/b and control lines.Nearly half of pollen grains were nonviable in the bcmf26a/b plants, whereas the percentage was only 1.01% in the control. Approximately 38.9%–71.1% of the pollen grains were irregular in the bcmf26a/b plants, whereas ~3.75% was irregular in the control. Standard errors are shown.

Mentions: The transgenic lines, bcmf26a/b-3, bcmf26a/b-4, and bcmf26a/b-5, which represent the average inhibitory level, were used for further analysis. The bcmf26a/b plants (S8G and S8I Fig) did not show defective phenotype at the vegetative development stage and the reproductive development stage compared with the control plants (S8H and S8J Fig). Meanwhile, the phenotypes of the floral organs in bcmf26a/b plants at the flowering stage were also normal (S8K, S8M, S8O, S8Q, and S8S Fig) compared with the control (S8L, S8N, S8P, S8R, and S8T Fig). Alexander staining was used to analyze pollen viability (Fig 7A and 7B). The statistical results showed that 37.5% to 47.8% of the bcmf26a/b pollen grains were nonviable, whereas this value was only ~2.1% for the control plants (Fig 8). Meanwhile, DAPI staining showed that the nonviable pollen grains of bcmf26a/b plants contained neither vegetative nuclei nor reproductive nuclei (Fig 7E and 7F). By contrast, the pollen grains from the control plants contained normal vegetative nuclei and reproductive nucleus (Fig 7C and 7D). SEM examination was used to detect the surface phenotype of pollen grains. The results revealed that mature pollen grains of the control plants were uniformly spheroid with finely reticulate ornamentation (Fig 7G and 7H). On the contrary, most of the pollen grains in the bcmf26a/b lines formed irregular clumps and exhibited abnormal reticulate ornamentation and germinal furrows (Fig 7I to 7N). Statistically, 38.9% to 71.1% of bcmf26a/b pollen grains exhibited irregular shapes, but this percentage was only ~3.75% for the control plants (Fig 8).


BcMF26a and BcMF26b Are Duplicated Polygalacturonase Genes with Divergent Expression Patterns and Functions in Pollen Development and Pollen Tube Formation in Brassica campestris.

Lyu M, Yu Y, Jiang J, Song L, Liang Y, Ma Z, Xiong X, Cao J - PLoS ONE (2015)

Viability percentage and irregular percentage of pollen grains in bcmf26a/b and control lines.Nearly half of pollen grains were nonviable in the bcmf26a/b plants, whereas the percentage was only 1.01% in the control. Approximately 38.9%–71.1% of the pollen grains were irregular in the bcmf26a/b plants, whereas ~3.75% was irregular in the control. Standard errors are shown.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4495986&req=5

pone.0131173.g008: Viability percentage and irregular percentage of pollen grains in bcmf26a/b and control lines.Nearly half of pollen grains were nonviable in the bcmf26a/b plants, whereas the percentage was only 1.01% in the control. Approximately 38.9%–71.1% of the pollen grains were irregular in the bcmf26a/b plants, whereas ~3.75% was irregular in the control. Standard errors are shown.
Mentions: The transgenic lines, bcmf26a/b-3, bcmf26a/b-4, and bcmf26a/b-5, which represent the average inhibitory level, were used for further analysis. The bcmf26a/b plants (S8G and S8I Fig) did not show defective phenotype at the vegetative development stage and the reproductive development stage compared with the control plants (S8H and S8J Fig). Meanwhile, the phenotypes of the floral organs in bcmf26a/b plants at the flowering stage were also normal (S8K, S8M, S8O, S8Q, and S8S Fig) compared with the control (S8L, S8N, S8P, S8R, and S8T Fig). Alexander staining was used to analyze pollen viability (Fig 7A and 7B). The statistical results showed that 37.5% to 47.8% of the bcmf26a/b pollen grains were nonviable, whereas this value was only ~2.1% for the control plants (Fig 8). Meanwhile, DAPI staining showed that the nonviable pollen grains of bcmf26a/b plants contained neither vegetative nuclei nor reproductive nuclei (Fig 7E and 7F). By contrast, the pollen grains from the control plants contained normal vegetative nuclei and reproductive nucleus (Fig 7C and 7D). SEM examination was used to detect the surface phenotype of pollen grains. The results revealed that mature pollen grains of the control plants were uniformly spheroid with finely reticulate ornamentation (Fig 7G and 7H). On the contrary, most of the pollen grains in the bcmf26a/b lines formed irregular clumps and exhibited abnormal reticulate ornamentation and germinal furrows (Fig 7I to 7N). Statistically, 38.9% to 71.1% of bcmf26a/b pollen grains exhibited irregular shapes, but this percentage was only ~3.75% for the control plants (Fig 8).

Bottom Line: We found that these two genes were orthologous genes of At4g33440, and they originated from a chromosomal segmental duplication.Promoter activity analysis showed that, at reproductive development stages, BcMF26b promoter was active in tapetum, pollen grains, and pistils, whereas BcMF26a promoter was only active in pistils.In the subcellular localization experiment, BcMF26a and BcMF26b proteins could be localized to the cell wall.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Cell & Molecular Biology, Institute of Vegetable Science, Zhejiang University, Hangzhou, China; Zhejiang Provincial Key Laboratory of Horticultural Plant Integrative Biology, Zhejiang University, Hangzhou, China.

ABSTRACT
Polygalacturonase (PG) is one of the cell wall hydrolytic enzymes involving in pectin degradation. A comparison of two highly conserved duplicated PG genes, namely, Brassica campestris Male Fertility 26a (BcMF26a) and BcMF26b, revealed the different features of their expression patterns and functions. We found that these two genes were orthologous genes of At4g33440, and they originated from a chromosomal segmental duplication. Although structurally similar, their regulatory and intron sequences largely diverged. QRT-PCR analysis showed that the expression level of BcMF26b was higher than that of BcMF26a in almost all the tested organs and tissues in Brassica campestris. Promoter activity analysis showed that, at reproductive development stages, BcMF26b promoter was active in tapetum, pollen grains, and pistils, whereas BcMF26a promoter was only active in pistils. In the subcellular localization experiment, BcMF26a and BcMF26b proteins could be localized to the cell wall. When the two genes were co-inhibited, pollen intine was formed abnormally and pollen tubes could not grow or stretch. Moreover, the knockout mutants of At4g33440 delayed the growth of pollen tubes. Therefore, BcMF26a/b can participate in the construction of pollen wall by modulating intine information and BcMF26b may play a major role in co-inhibiting transformed plants.

No MeSH data available.


Related in: MedlinePlus