Limits...
Inactivation of the UGPase1 gene causes genic male sterility and endosperm chalkiness in rice (Oryza sativa L.).

Woo MO, Ham TH, Ji HS, Choi MS, Jiang W, Chu SH, Piao R, Chin JH, Kim JA, Park BS, Seo HS, Jwa NS, McCouch S, Koh HJ - Plant J. (2008)

Bottom Line: A rice genic male-sterility gene ms-h is recessive and has a pleiotropic effect on the chalky endosperm.After fine mapping, nucleotide sequencing analysis of the ms-h gene revealed a single nucleotide substitution at the 3'-splice junction of the 14th intron of the UDP-glucose pyrophosphorylase 1 (UGPase1; EC2.7.7.9) gene, which causes the expression of two mature transcripts with abnormal sizes caused by the aberrant splicing.In addition, both phenotypes were co-segregated with the UGPase1 transgene in segregating T(1) plants, which demonstrates that UGPase1 has functional roles in both male sterility and the development of a chalky endosperm.

View Article: PubMed Central - PubMed

Affiliation: Department of Plant Science and Research Institute of Agriculture and Life Sciences, Seoul National University, Seoul 151-921, Korea.

ABSTRACT
A rice genic male-sterility gene ms-h is recessive and has a pleiotropic effect on the chalky endosperm. After fine mapping, nucleotide sequencing analysis of the ms-h gene revealed a single nucleotide substitution at the 3'-splice junction of the 14th intron of the UDP-glucose pyrophosphorylase 1 (UGPase1; EC2.7.7.9) gene, which causes the expression of two mature transcripts with abnormal sizes caused by the aberrant splicing. An in vitro functional assay showed that both proteins encoded by the two abnormal transcripts have no UGPase activity. The suppression of UGPase by the introduction of a UGPase1-RNAi construct in wild-type plants nearly eliminated seed set because of the male defect, with developmental retardation similar to the ms-h mutant phenotype, whereas overexpression of UGPase1 in ms-h mutant plants restored male fertility and the transformants produced T(1) seeds that segregated into normal and chalky endosperms. In addition, both phenotypes were co-segregated with the UGPase1 transgene in segregating T(1) plants, which demonstrates that UGPase1 has functional roles in both male sterility and the development of a chalky endosperm. Our results suggest that UGPase1 plays a key role in pollen development as well as seed carbohydrate metabolism.

Show MeSH

Related in: MedlinePlus

Schematic diagram of the UGPase1 gene and derived cleaved amplified polymorphic sequence (dCAPS) marker analysis.The mutation within the UGPase1 gene in the ms-h mutant.dCAPS marker development for detection of the 1-bp substitution at the 3′-splice junction of the 14th intron. The dCAPS marker using a mismatch primer selectively generates a restriction site (SpeI) in the mutant, but not in the wild-type parent. Lowercase letters of sequences indicate the 14th intron and uppercase letters indicate the 15th exon.dCAPS marker genotype of F2 plants from Hwacheong ms-h mutant × Hwacheong, classified by phenotype. After digestion with restriction enzyme SpeI, a single, 196-bp PCR product was observed in fertile homozygotes (genotype 1), whereas in the male-sterile homozygotes (genotype 3), a shorter, 169-bp PCR product was observed, resulting from the generation of a new SpeI recognition site resulting from the single nucleotide substitution. In fertile heterozygotes (genotype 2), both fragments are observed. Genotype 1, Ms-h/Ms-h; genotype 2, Ms-h/ms-h; genotype 3, ms-h/ms-h.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2327258&req=5

fig02: Schematic diagram of the UGPase1 gene and derived cleaved amplified polymorphic sequence (dCAPS) marker analysis.The mutation within the UGPase1 gene in the ms-h mutant.dCAPS marker development for detection of the 1-bp substitution at the 3′-splice junction of the 14th intron. The dCAPS marker using a mismatch primer selectively generates a restriction site (SpeI) in the mutant, but not in the wild-type parent. Lowercase letters of sequences indicate the 14th intron and uppercase letters indicate the 15th exon.dCAPS marker genotype of F2 plants from Hwacheong ms-h mutant × Hwacheong, classified by phenotype. After digestion with restriction enzyme SpeI, a single, 196-bp PCR product was observed in fertile homozygotes (genotype 1), whereas in the male-sterile homozygotes (genotype 3), a shorter, 169-bp PCR product was observed, resulting from the generation of a new SpeI recognition site resulting from the single nucleotide substitution. In fertile heterozygotes (genotype 2), both fragments are observed. Genotype 1, Ms-h/Ms-h; genotype 2, Ms-h/ms-h; genotype 3, ms-h/ms-h.

Mentions: Eleven candidate genes were identified in the 60-kb target interval based on in silico genome annotation (http://rgp.dna.affrc.go.jp; http://www.tigr.org/tdb/e2k1/osa1; Figure 1d). To identify the best candidate for the ms-h gene among these genes, we sequenced all 11 gene candidates in the ms-h mutant and in the wild-type (wt), Hwacheong, and compared them with the corresponding sequences in the publicly available genome sequence for cv. Nipponbare. This comparison identified a point mutation in the UGPase1 gene that distinguished the ms-h mutant from both Hwacheong and Nipponbare. The critical polymorphism was a single nucleotide substitution of Guanine to Adenine, corresponding to the final nucleotide at the 3′-splice junction of the 14th intron of the UGPase1 gene (Figure 2a).


Inactivation of the UGPase1 gene causes genic male sterility and endosperm chalkiness in rice (Oryza sativa L.).

Woo MO, Ham TH, Ji HS, Choi MS, Jiang W, Chu SH, Piao R, Chin JH, Kim JA, Park BS, Seo HS, Jwa NS, McCouch S, Koh HJ - Plant J. (2008)

Schematic diagram of the UGPase1 gene and derived cleaved amplified polymorphic sequence (dCAPS) marker analysis.The mutation within the UGPase1 gene in the ms-h mutant.dCAPS marker development for detection of the 1-bp substitution at the 3′-splice junction of the 14th intron. The dCAPS marker using a mismatch primer selectively generates a restriction site (SpeI) in the mutant, but not in the wild-type parent. Lowercase letters of sequences indicate the 14th intron and uppercase letters indicate the 15th exon.dCAPS marker genotype of F2 plants from Hwacheong ms-h mutant × Hwacheong, classified by phenotype. After digestion with restriction enzyme SpeI, a single, 196-bp PCR product was observed in fertile homozygotes (genotype 1), whereas in the male-sterile homozygotes (genotype 3), a shorter, 169-bp PCR product was observed, resulting from the generation of a new SpeI recognition site resulting from the single nucleotide substitution. In fertile heterozygotes (genotype 2), both fragments are observed. Genotype 1, Ms-h/Ms-h; genotype 2, Ms-h/ms-h; genotype 3, ms-h/ms-h.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2327258&req=5

fig02: Schematic diagram of the UGPase1 gene and derived cleaved amplified polymorphic sequence (dCAPS) marker analysis.The mutation within the UGPase1 gene in the ms-h mutant.dCAPS marker development for detection of the 1-bp substitution at the 3′-splice junction of the 14th intron. The dCAPS marker using a mismatch primer selectively generates a restriction site (SpeI) in the mutant, but not in the wild-type parent. Lowercase letters of sequences indicate the 14th intron and uppercase letters indicate the 15th exon.dCAPS marker genotype of F2 plants from Hwacheong ms-h mutant × Hwacheong, classified by phenotype. After digestion with restriction enzyme SpeI, a single, 196-bp PCR product was observed in fertile homozygotes (genotype 1), whereas in the male-sterile homozygotes (genotype 3), a shorter, 169-bp PCR product was observed, resulting from the generation of a new SpeI recognition site resulting from the single nucleotide substitution. In fertile heterozygotes (genotype 2), both fragments are observed. Genotype 1, Ms-h/Ms-h; genotype 2, Ms-h/ms-h; genotype 3, ms-h/ms-h.
Mentions: Eleven candidate genes were identified in the 60-kb target interval based on in silico genome annotation (http://rgp.dna.affrc.go.jp; http://www.tigr.org/tdb/e2k1/osa1; Figure 1d). To identify the best candidate for the ms-h gene among these genes, we sequenced all 11 gene candidates in the ms-h mutant and in the wild-type (wt), Hwacheong, and compared them with the corresponding sequences in the publicly available genome sequence for cv. Nipponbare. This comparison identified a point mutation in the UGPase1 gene that distinguished the ms-h mutant from both Hwacheong and Nipponbare. The critical polymorphism was a single nucleotide substitution of Guanine to Adenine, corresponding to the final nucleotide at the 3′-splice junction of the 14th intron of the UGPase1 gene (Figure 2a).

Bottom Line: A rice genic male-sterility gene ms-h is recessive and has a pleiotropic effect on the chalky endosperm.After fine mapping, nucleotide sequencing analysis of the ms-h gene revealed a single nucleotide substitution at the 3'-splice junction of the 14th intron of the UDP-glucose pyrophosphorylase 1 (UGPase1; EC2.7.7.9) gene, which causes the expression of two mature transcripts with abnormal sizes caused by the aberrant splicing.In addition, both phenotypes were co-segregated with the UGPase1 transgene in segregating T(1) plants, which demonstrates that UGPase1 has functional roles in both male sterility and the development of a chalky endosperm.

View Article: PubMed Central - PubMed

Affiliation: Department of Plant Science and Research Institute of Agriculture and Life Sciences, Seoul National University, Seoul 151-921, Korea.

ABSTRACT
A rice genic male-sterility gene ms-h is recessive and has a pleiotropic effect on the chalky endosperm. After fine mapping, nucleotide sequencing analysis of the ms-h gene revealed a single nucleotide substitution at the 3'-splice junction of the 14th intron of the UDP-glucose pyrophosphorylase 1 (UGPase1; EC2.7.7.9) gene, which causes the expression of two mature transcripts with abnormal sizes caused by the aberrant splicing. An in vitro functional assay showed that both proteins encoded by the two abnormal transcripts have no UGPase activity. The suppression of UGPase by the introduction of a UGPase1-RNAi construct in wild-type plants nearly eliminated seed set because of the male defect, with developmental retardation similar to the ms-h mutant phenotype, whereas overexpression of UGPase1 in ms-h mutant plants restored male fertility and the transformants produced T(1) seeds that segregated into normal and chalky endosperms. In addition, both phenotypes were co-segregated with the UGPase1 transgene in segregating T(1) plants, which demonstrates that UGPase1 has functional roles in both male sterility and the development of a chalky endosperm. Our results suggest that UGPase1 plays a key role in pollen development as well as seed carbohydrate metabolism.

Show MeSH
Related in: MedlinePlus