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Overexpression of the soybean GmERF3 gene, an AP2/ERF type transcription factor for increased tolerances to salt, drought, and diseases in transgenic tobacco.

Zhang G, Chen M, Li L, Xu Z, Chen X, Guo J, Ma Y - J. Exp. Bot. (2009)

Bottom Line: The GmERF3 protein fused to the GAL4 DNA-binding domain to activate transcription of reporter genes in yeast.Furthermore, overexpression of GmERF3 in transgenic tobacco led to higher levels of free proline and soluble carbohydrates compared to wild-type plants under drought conditions.The overall results suggested that GmERF3 as an AP2/ERF transcription factor may play dual roles in response to biotic and abiotic stresses in plants.

View Article: PubMed Central - PubMed

Affiliation: The National Key Facility for Crop Genetic Resources and Genetic Improvement, Institute of Crop Sciences, Chinese Academy of Agricultural Sciences, Beijing, China.

ABSTRACT
A new member of the AP2/ERF transcription factor family, GmERF3, was isolated from soybean. Sequence analysis showed that GmERF3 contained an AP2/ERF domain of 58 amino acids and two putative nuclear localization signal (NLS) domains. It belonged to a group IV protein in the ERF (ethylene response factor) subfamily as typified by a conserved N-terminal motif [MCGGAI(I/L)]. Expression of GmERF3 was induced by treatments with high salinity, drought, abscisic acid (ABA), salicylic acid (SA), jasmonic acid (JA), ethylene (ET), and soybean mosaic virus (SMV), whereas there was no significant GmERF3 mRNA accumulation under cold stress treatment. GmERF3 could bind to the GCC box and DRE/CRT element, and was targeted to the nucleus when transiently expressed in onion epidermal cells. The GmERF3 protein fused to the GAL4 DNA-binding domain to activate transcription of reporter genes in yeast. Ectopic expression of the GmERF3 gene in transgenic tobacco plants induced the expression of some PR genes and enhanced resistance against infection by Ralstonia solanacearum, Alternaria alternata, and tobacco mosaic virus (TMV), and gave tolerance to high salinity and dehydration stresses. Furthermore, overexpression of GmERF3 in transgenic tobacco led to higher levels of free proline and soluble carbohydrates compared to wild-type plants under drought conditions. The overall results suggested that GmERF3 as an AP2/ERF transcription factor may play dual roles in response to biotic and abiotic stresses in plants.

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Deduced amino acid sequences of AP2/ERF-related proteins and phylogenic relationships of selected AP2/ERF domains from AP2/ERF-related proteins. (A) Comparison of deduced amino acid sequences of AP2/ERF-related proteins that have high sequence similarity with GmERF3. Amino acid residues that are conserved in at least three of the six sequences are shaded, whereas amino acids identical in all six proteins are shown in dark grey. The black bar above the sequences represents the putative acidic domain. Double overline represents the highly conserved N-terminal MCGGAII/L motif of unknown function. Dots (·) represent putative nuclear localization signals. An asterisk (*) indicates a conserved DNA-binding domain (AP2/ERF domain). Dashes show gaps in the amino acid sequences introduced to optimize alignment. (B) Phylogenic comparison of the GmERF3 protein and some AP2/ERF-related protein sequences, based on the selected AP2/ERF domain amino acid sequences of those proteins. Alignments were made in Clustal X using the default parameters. Accession numbers for the AP2/ERF proteins used are as follows: RAP2.3, NP188299; OsBIERF1, AAV98700; FsERF1, CAE54591; CaERFLP1, AAS20427; CMeERF2, BAD01556; LeERF2, AAO34704; RAP2.12, NP001077718; RAP2.2, NP850583; RAP2.6L, NP196837; ABR1, NP201280; RAP2.6, NP175008; AtERF10, NP171876; AtERF3, NP175479; OsBIERF2, AAV98701; LeERF3, AAO34705; NtERF5, AAU81956; GmEREBP1, AAM45475; ORCA3, CAB96899; ATERF13, NP182011; ATERF2, NP199533; LeERF1, AAO34703; Tsi1, AAC14323; CRF1, NP192852.
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fig1: Deduced amino acid sequences of AP2/ERF-related proteins and phylogenic relationships of selected AP2/ERF domains from AP2/ERF-related proteins. (A) Comparison of deduced amino acid sequences of AP2/ERF-related proteins that have high sequence similarity with GmERF3. Amino acid residues that are conserved in at least three of the six sequences are shaded, whereas amino acids identical in all six proteins are shown in dark grey. The black bar above the sequences represents the putative acidic domain. Double overline represents the highly conserved N-terminal MCGGAII/L motif of unknown function. Dots (·) represent putative nuclear localization signals. An asterisk (*) indicates a conserved DNA-binding domain (AP2/ERF domain). Dashes show gaps in the amino acid sequences introduced to optimize alignment. (B) Phylogenic comparison of the GmERF3 protein and some AP2/ERF-related protein sequences, based on the selected AP2/ERF domain amino acid sequences of those proteins. Alignments were made in Clustal X using the default parameters. Accession numbers for the AP2/ERF proteins used are as follows: RAP2.3, NP188299; OsBIERF1, AAV98700; FsERF1, CAE54591; CaERFLP1, AAS20427; CMeERF2, BAD01556; LeERF2, AAO34704; RAP2.12, NP001077718; RAP2.2, NP850583; RAP2.6L, NP196837; ABR1, NP201280; RAP2.6, NP175008; AtERF10, NP171876; AtERF3, NP175479; OsBIERF2, AAV98701; LeERF3, AAO34705; NtERF5, AAU81956; GmEREBP1, AAM45475; ORCA3, CAB96899; ATERF13, NP182011; ATERF2, NP199533; LeERF1, AAO34703; Tsi1, AAC14323; CRF1, NP192852.

Mentions: The full-length cDNA sequence of an ERF homologue, GmERF3 gene (GenBank Accession EU681278) was isolated from soybean total RNA by reverse transcription polymerase chain reaction (RT-PCR). Sequence analysis revealed that it contained a 1038 bp open reading frame encoding a polypeptide of 345 amino acids with predicted molecular mass of 38 kDa (pI 4.78). The predicted GmERF3 protein contained a conserved DNA-binding domain (AP2/ERF domain) of 58 amino acids, two putative basic amino acid regions (K32KRK and R94KRK) that potentially acted as NLS, an acidic amino acid-rich region (D37LDVDFEADFRDFKDDSDIDDDDDD) that might act as a transcriptional activation domain in the N-terminal region, and a conserved N-terminal motif of unknown function (MCGGAII) (Fig. 1A). Alignment and phylogenetic tree analysis revealed that GmERF3 is most similar to previously described ERF class B-2 subgroup members (RAP2.3, RAP2.2, and RAP2.12; Sakuma et al., 2002) (Fig. 1B). GmERF3 has 38–59% identity for overall amino acid sequence and 94–99% for the AP2/ERF domain to Fagus sylvatica FsERF1, rice OsBIERF1, Capsicum annuum CaERFLP1, Cucumis melo CMe-ERF2, and tomato LeERF2. Tournier et al. (2003) identified tomato LeERF2, a novel class IV ERF, characterized by an N-terminal signature sequence, MCGGAII/L. This motif was also present in GmERF3 and members of the recently identified Class IV (Tournier et al., 2003; Lee et al., 2004; Cao et al., 2006; Mizuno et al., 2006).


Overexpression of the soybean GmERF3 gene, an AP2/ERF type transcription factor for increased tolerances to salt, drought, and diseases in transgenic tobacco.

Zhang G, Chen M, Li L, Xu Z, Chen X, Guo J, Ma Y - J. Exp. Bot. (2009)

Deduced amino acid sequences of AP2/ERF-related proteins and phylogenic relationships of selected AP2/ERF domains from AP2/ERF-related proteins. (A) Comparison of deduced amino acid sequences of AP2/ERF-related proteins that have high sequence similarity with GmERF3. Amino acid residues that are conserved in at least three of the six sequences are shaded, whereas amino acids identical in all six proteins are shown in dark grey. The black bar above the sequences represents the putative acidic domain. Double overline represents the highly conserved N-terminal MCGGAII/L motif of unknown function. Dots (·) represent putative nuclear localization signals. An asterisk (*) indicates a conserved DNA-binding domain (AP2/ERF domain). Dashes show gaps in the amino acid sequences introduced to optimize alignment. (B) Phylogenic comparison of the GmERF3 protein and some AP2/ERF-related protein sequences, based on the selected AP2/ERF domain amino acid sequences of those proteins. Alignments were made in Clustal X using the default parameters. Accession numbers for the AP2/ERF proteins used are as follows: RAP2.3, NP188299; OsBIERF1, AAV98700; FsERF1, CAE54591; CaERFLP1, AAS20427; CMeERF2, BAD01556; LeERF2, AAO34704; RAP2.12, NP001077718; RAP2.2, NP850583; RAP2.6L, NP196837; ABR1, NP201280; RAP2.6, NP175008; AtERF10, NP171876; AtERF3, NP175479; OsBIERF2, AAV98701; LeERF3, AAO34705; NtERF5, AAU81956; GmEREBP1, AAM45475; ORCA3, CAB96899; ATERF13, NP182011; ATERF2, NP199533; LeERF1, AAO34703; Tsi1, AAC14323; CRF1, NP192852.
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Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC2736888&req=5

fig1: Deduced amino acid sequences of AP2/ERF-related proteins and phylogenic relationships of selected AP2/ERF domains from AP2/ERF-related proteins. (A) Comparison of deduced amino acid sequences of AP2/ERF-related proteins that have high sequence similarity with GmERF3. Amino acid residues that are conserved in at least three of the six sequences are shaded, whereas amino acids identical in all six proteins are shown in dark grey. The black bar above the sequences represents the putative acidic domain. Double overline represents the highly conserved N-terminal MCGGAII/L motif of unknown function. Dots (·) represent putative nuclear localization signals. An asterisk (*) indicates a conserved DNA-binding domain (AP2/ERF domain). Dashes show gaps in the amino acid sequences introduced to optimize alignment. (B) Phylogenic comparison of the GmERF3 protein and some AP2/ERF-related protein sequences, based on the selected AP2/ERF domain amino acid sequences of those proteins. Alignments were made in Clustal X using the default parameters. Accession numbers for the AP2/ERF proteins used are as follows: RAP2.3, NP188299; OsBIERF1, AAV98700; FsERF1, CAE54591; CaERFLP1, AAS20427; CMeERF2, BAD01556; LeERF2, AAO34704; RAP2.12, NP001077718; RAP2.2, NP850583; RAP2.6L, NP196837; ABR1, NP201280; RAP2.6, NP175008; AtERF10, NP171876; AtERF3, NP175479; OsBIERF2, AAV98701; LeERF3, AAO34705; NtERF5, AAU81956; GmEREBP1, AAM45475; ORCA3, CAB96899; ATERF13, NP182011; ATERF2, NP199533; LeERF1, AAO34703; Tsi1, AAC14323; CRF1, NP192852.
Mentions: The full-length cDNA sequence of an ERF homologue, GmERF3 gene (GenBank Accession EU681278) was isolated from soybean total RNA by reverse transcription polymerase chain reaction (RT-PCR). Sequence analysis revealed that it contained a 1038 bp open reading frame encoding a polypeptide of 345 amino acids with predicted molecular mass of 38 kDa (pI 4.78). The predicted GmERF3 protein contained a conserved DNA-binding domain (AP2/ERF domain) of 58 amino acids, two putative basic amino acid regions (K32KRK and R94KRK) that potentially acted as NLS, an acidic amino acid-rich region (D37LDVDFEADFRDFKDDSDIDDDDDD) that might act as a transcriptional activation domain in the N-terminal region, and a conserved N-terminal motif of unknown function (MCGGAII) (Fig. 1A). Alignment and phylogenetic tree analysis revealed that GmERF3 is most similar to previously described ERF class B-2 subgroup members (RAP2.3, RAP2.2, and RAP2.12; Sakuma et al., 2002) (Fig. 1B). GmERF3 has 38–59% identity for overall amino acid sequence and 94–99% for the AP2/ERF domain to Fagus sylvatica FsERF1, rice OsBIERF1, Capsicum annuum CaERFLP1, Cucumis melo CMe-ERF2, and tomato LeERF2. Tournier et al. (2003) identified tomato LeERF2, a novel class IV ERF, characterized by an N-terminal signature sequence, MCGGAII/L. This motif was also present in GmERF3 and members of the recently identified Class IV (Tournier et al., 2003; Lee et al., 2004; Cao et al., 2006; Mizuno et al., 2006).

Bottom Line: The GmERF3 protein fused to the GAL4 DNA-binding domain to activate transcription of reporter genes in yeast.Furthermore, overexpression of GmERF3 in transgenic tobacco led to higher levels of free proline and soluble carbohydrates compared to wild-type plants under drought conditions.The overall results suggested that GmERF3 as an AP2/ERF transcription factor may play dual roles in response to biotic and abiotic stresses in plants.

View Article: PubMed Central - PubMed

Affiliation: The National Key Facility for Crop Genetic Resources and Genetic Improvement, Institute of Crop Sciences, Chinese Academy of Agricultural Sciences, Beijing, China.

ABSTRACT
A new member of the AP2/ERF transcription factor family, GmERF3, was isolated from soybean. Sequence analysis showed that GmERF3 contained an AP2/ERF domain of 58 amino acids and two putative nuclear localization signal (NLS) domains. It belonged to a group IV protein in the ERF (ethylene response factor) subfamily as typified by a conserved N-terminal motif [MCGGAI(I/L)]. Expression of GmERF3 was induced by treatments with high salinity, drought, abscisic acid (ABA), salicylic acid (SA), jasmonic acid (JA), ethylene (ET), and soybean mosaic virus (SMV), whereas there was no significant GmERF3 mRNA accumulation under cold stress treatment. GmERF3 could bind to the GCC box and DRE/CRT element, and was targeted to the nucleus when transiently expressed in onion epidermal cells. The GmERF3 protein fused to the GAL4 DNA-binding domain to activate transcription of reporter genes in yeast. Ectopic expression of the GmERF3 gene in transgenic tobacco plants induced the expression of some PR genes and enhanced resistance against infection by Ralstonia solanacearum, Alternaria alternata, and tobacco mosaic virus (TMV), and gave tolerance to high salinity and dehydration stresses. Furthermore, overexpression of GmERF3 in transgenic tobacco led to higher levels of free proline and soluble carbohydrates compared to wild-type plants under drought conditions. The overall results suggested that GmERF3 as an AP2/ERF transcription factor may play dual roles in response to biotic and abiotic stresses in plants.

Show MeSH
Related in: MedlinePlus