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Subfunctionalization of sigma factors during the evolution of land plants based on mutant analysis of liverwort (Marchantia polymorpha L.) MpSIG1.

Ueda M, Takami T, Peng L, Ishizaki K, Kohchi T, Shikanai T, Nishimura Y - Genome Biol Evol (2013)

Bottom Line: The mutant did not show any visible phenotypes, implying that MpSIG1 function is redundant with that of other sigma factors.The transcript levels of genes clustered in the petL, psaA, psbB, psbK, and psbE operons of liverwort were lower than those in the wild type, a result similar to that in the SIG1 defective mutant in rice (Oryza sativa).Our study shows the ancestral function of SIG1 and the process of functional partitioning (subfunctionalization) of sigma factors during the evolution of land plants.

View Article: PubMed Central - PubMed

Affiliation: Department of Botany, Graduate School of Science, Kyoto University, Japan.

ABSTRACT
Sigma factor is a subunit of plastid-encoded RNA polymerase that regulates the transcription of plastid-encoded genes by recognizing a set of promoters. Sigma factors have increased in copy number and have diversified during the evolution of land plants, but details of this process remain unknown. Liverworts represent the basal group of embryophytes and are expected to retain the ancestral features of land plants. In liverwort (Marchantia polymorpha L.), we isolated and characterized a T-DNA-tagged mutant (Mpsig1) of sigma factor 1 (MpSIG1). The mutant did not show any visible phenotypes, implying that MpSIG1 function is redundant with that of other sigma factors. However, quantitative reverse-transcription polymerase chain reaction and RNA gel blot analysis revealed that genes related to photosynthesis were downregulated, resulting in the minor reduction of some protein complexes. The transcript levels of genes clustered in the petL, psaA, psbB, psbK, and psbE operons of liverwort were lower than those in the wild type, a result similar to that in the SIG1 defective mutant in rice (Oryza sativa). Overexpression analysis revealed primitive functional divergence between the SIG1 and SIG2 proteins in bryophytes, whereas these proteins still retain functional redundancy. We also discovered that the predominant sigma factor for ndhF mRNA expression has been diversified in liverwort, Arabidopsis (Arabidopsis thaliana), and rice. Our study shows the ancestral function of SIG1 and the process of functional partitioning (subfunctionalization) of sigma factors during the evolution of land plants.

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Plastid transcript levels in the Ossig1 mutant. (A) The exon–intron structure of OsSIG1. The black boxes, white boxes, and horizontal lines indicate exons, untranslated regions, and introns, respectively. The black arrows show the positions of the primers designed for genotyping. The position of a tos17 insertion in the OsSIG1 locus of line NE8184 is shown with a triangle. The black arrows and the triangle are not to scale. (B) Genotyping in the Ossig1 mutant. The P4 and P5 (P4/P5 in the panel) primer pair detects WT plants, and the P4 and P6 (P4/P6 in the panel) primer pair detects the tos17 insertion. The primer sequences are shown in supplementary table S2, Supplementary Material online. (C) Expression analysis of OsSIG1 by RT-PCR in plants. A miss-spliced RT-PCR product was detected in the Ossig1 mutant as previously reported (Tozawa et al. 2007). (D) The abundance of mRNAs was measured in the Ossig1 mutant using qRT-PCR.
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evt137-F6: Plastid transcript levels in the Ossig1 mutant. (A) The exon–intron structure of OsSIG1. The black boxes, white boxes, and horizontal lines indicate exons, untranslated regions, and introns, respectively. The black arrows show the positions of the primers designed for genotyping. The position of a tos17 insertion in the OsSIG1 locus of line NE8184 is shown with a triangle. The black arrows and the triangle are not to scale. (B) Genotyping in the Ossig1 mutant. The P4 and P5 (P4/P5 in the panel) primer pair detects WT plants, and the P4 and P6 (P4/P6 in the panel) primer pair detects the tos17 insertion. The primer sequences are shown in supplementary table S2, Supplementary Material online. (C) Expression analysis of OsSIG1 by RT-PCR in plants. A miss-spliced RT-PCR product was detected in the Ossig1 mutant as previously reported (Tozawa et al. 2007). (D) The abundance of mRNAs was measured in the Ossig1 mutant using qRT-PCR.

Mentions: Total DNA was isolated from thalli using conventional methods and diluted in 200 µl of Tris–EDTA (ethylenediaminetetraacetic acid) buffer (10 mM Tris–HCl [pH 8.0], 1 mM EDTA [pH 8.0]). To confirm the position of the T-DNA insertion in the Mpsig1 mutant and the success of the complementation, 1 µl from the extract was used in genomic PCR with primers P1, P2, and P3. All of the primers used in this article are shown in supplementary tables S2–S5, Supplementary Material online. Each primer position and the primer combinations for the genotyping of both types are shown in figure 1A. To confirm the position of the tos17 insertion in the Ossig1 mutant (NE8184), 1 µl from the extract was used for genomic PCR with primers P4, P5, and P6. Each primer position and the primer combinations for the genotyping of both types are shown in figure 6A. The PCRs were conducted using KOD-FX DNA polymerase (TOYOBO, Osaka, Japan).Fig. 1.—


Subfunctionalization of sigma factors during the evolution of land plants based on mutant analysis of liverwort (Marchantia polymorpha L.) MpSIG1.

Ueda M, Takami T, Peng L, Ishizaki K, Kohchi T, Shikanai T, Nishimura Y - Genome Biol Evol (2013)

Plastid transcript levels in the Ossig1 mutant. (A) The exon–intron structure of OsSIG1. The black boxes, white boxes, and horizontal lines indicate exons, untranslated regions, and introns, respectively. The black arrows show the positions of the primers designed for genotyping. The position of a tos17 insertion in the OsSIG1 locus of line NE8184 is shown with a triangle. The black arrows and the triangle are not to scale. (B) Genotyping in the Ossig1 mutant. The P4 and P5 (P4/P5 in the panel) primer pair detects WT plants, and the P4 and P6 (P4/P6 in the panel) primer pair detects the tos17 insertion. The primer sequences are shown in supplementary table S2, Supplementary Material online. (C) Expression analysis of OsSIG1 by RT-PCR in plants. A miss-spliced RT-PCR product was detected in the Ossig1 mutant as previously reported (Tozawa et al. 2007). (D) The abundance of mRNAs was measured in the Ossig1 mutant using qRT-PCR.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3814195&req=5

evt137-F6: Plastid transcript levels in the Ossig1 mutant. (A) The exon–intron structure of OsSIG1. The black boxes, white boxes, and horizontal lines indicate exons, untranslated regions, and introns, respectively. The black arrows show the positions of the primers designed for genotyping. The position of a tos17 insertion in the OsSIG1 locus of line NE8184 is shown with a triangle. The black arrows and the triangle are not to scale. (B) Genotyping in the Ossig1 mutant. The P4 and P5 (P4/P5 in the panel) primer pair detects WT plants, and the P4 and P6 (P4/P6 in the panel) primer pair detects the tos17 insertion. The primer sequences are shown in supplementary table S2, Supplementary Material online. (C) Expression analysis of OsSIG1 by RT-PCR in plants. A miss-spliced RT-PCR product was detected in the Ossig1 mutant as previously reported (Tozawa et al. 2007). (D) The abundance of mRNAs was measured in the Ossig1 mutant using qRT-PCR.
Mentions: Total DNA was isolated from thalli using conventional methods and diluted in 200 µl of Tris–EDTA (ethylenediaminetetraacetic acid) buffer (10 mM Tris–HCl [pH 8.0], 1 mM EDTA [pH 8.0]). To confirm the position of the T-DNA insertion in the Mpsig1 mutant and the success of the complementation, 1 µl from the extract was used in genomic PCR with primers P1, P2, and P3. All of the primers used in this article are shown in supplementary tables S2–S5, Supplementary Material online. Each primer position and the primer combinations for the genotyping of both types are shown in figure 1A. To confirm the position of the tos17 insertion in the Ossig1 mutant (NE8184), 1 µl from the extract was used for genomic PCR with primers P4, P5, and P6. Each primer position and the primer combinations for the genotyping of both types are shown in figure 6A. The PCRs were conducted using KOD-FX DNA polymerase (TOYOBO, Osaka, Japan).Fig. 1.—

Bottom Line: The mutant did not show any visible phenotypes, implying that MpSIG1 function is redundant with that of other sigma factors.The transcript levels of genes clustered in the petL, psaA, psbB, psbK, and psbE operons of liverwort were lower than those in the wild type, a result similar to that in the SIG1 defective mutant in rice (Oryza sativa).Our study shows the ancestral function of SIG1 and the process of functional partitioning (subfunctionalization) of sigma factors during the evolution of land plants.

View Article: PubMed Central - PubMed

Affiliation: Department of Botany, Graduate School of Science, Kyoto University, Japan.

ABSTRACT
Sigma factor is a subunit of plastid-encoded RNA polymerase that regulates the transcription of plastid-encoded genes by recognizing a set of promoters. Sigma factors have increased in copy number and have diversified during the evolution of land plants, but details of this process remain unknown. Liverworts represent the basal group of embryophytes and are expected to retain the ancestral features of land plants. In liverwort (Marchantia polymorpha L.), we isolated and characterized a T-DNA-tagged mutant (Mpsig1) of sigma factor 1 (MpSIG1). The mutant did not show any visible phenotypes, implying that MpSIG1 function is redundant with that of other sigma factors. However, quantitative reverse-transcription polymerase chain reaction and RNA gel blot analysis revealed that genes related to photosynthesis were downregulated, resulting in the minor reduction of some protein complexes. The transcript levels of genes clustered in the petL, psaA, psbB, psbK, and psbE operons of liverwort were lower than those in the wild type, a result similar to that in the SIG1 defective mutant in rice (Oryza sativa). Overexpression analysis revealed primitive functional divergence between the SIG1 and SIG2 proteins in bryophytes, whereas these proteins still retain functional redundancy. We also discovered that the predominant sigma factor for ndhF mRNA expression has been diversified in liverwort, Arabidopsis (Arabidopsis thaliana), and rice. Our study shows the ancestral function of SIG1 and the process of functional partitioning (subfunctionalization) of sigma factors during the evolution of land plants.

Show MeSH
Related in: MedlinePlus