Os06g0211200

OsAREB1,an ABRE-binding protein responding to ABA and glucosemay, may function as a positive regulator in drought/heat stresses response,
but a negative regulator in flowering time in Arabidopsis.

Annotated Information

Function

Firstly, overexpression of OsAREB1 alters seedling sensitivity to ABA and glucose and OsAREB1 might have a crucial role in these two signaling pathways. Roots of transgenic plants were hypersensitive to ABA. Also,transgenic seeds were hypersensitive to glucose in germination period.

Secondly, 35S-OsAREB1 plants enhanced the resistance to drought and heat.Transgenic seeds can hold more water to stand against drought condition and up-regulate stress-related genes, such as RD29A, RD29B.

Thirdly, OsAREB1 delay the flowering time.by down-regulating the expression of flowering-related genes, such as FT, SOC1, LFYand AP1.

In other work ,A number of transcription factors (TFs) regulate stress-responsive gene expression. OsDREB1s and OsDREB2s were identified as abiotic-stress responsive TFs that belong to the AP2/ERF family. Similar to Arabidopsis, these DREB regulons were most likely not involved in the abscisic acid (ABA) pathway. OsAREBs such as OsAREB1 were identified as key components in ABAdependent transcriptional networks in rice. The abscisic acid (ABA) responsive element (ABRE) binding protein (AREB)/ABRE binding factor (ABF) regulon functions in ABA-dependent gene expression under osmotic stress conditions

Also,in Arabidopsis, bZIP-type transcription factors AREBs/ABFs bind an abscisic acid (ABA)-responsive cis-acting element named ABRE and transactivate downstream gene expression inArabidopsis. Because AREB1 overexpression could not induce downstream gene expression, activation of AREB1 requires ABA-dependent posttranscriptional modification. We confirmed that ABA activated 42-kDa kinase activity, which, in turn, phosphorylated Ser/Thr residues of R-XX-S/T sites in the conserved regions of AREB1. Amino acid substitutions of R-X-X-S/T sites to Ala suppressed transactivation activity, and multiple substitution of these sites resulted in almost complete suppression of transactivation activity in transient assays. In contrast, substitution of the Ser/Thr residues to Asp resulted in high transactivation activity without exogenous ABA application. A phosphorylated, transcriptionally active form was achieved by substitution of Ser/Thr in all conserved R-X-X-S/T sites to Asp. Transgenic plants overexpressing the phosphorylated active form of AREB1 expressed many ABA-inducible genes, such as RD29B, without ABA treatment. These results indicate that the ABA-dependent multisite phosphorylation of AREB1 regulates its own activation in plants.

The phytohormone abscisic acid (ABA) plays important roles in seed maturation and dormancy and is also involved in the adaptation of vegetative tissues to abiotic environmental stresses, such as drought and high salinity. ABA promotes stomatal closure in guard cells and regulates the expression of many genes, the products of which may function in dehydration tolerance in both vegetative tissues and seeds. Many ABA-inducible genes contain a conserved element named ABA-responsive element (ABRE) (PyACGTGG/TC) in their promoter regions. The ABRE functions as a cis-acting element and is involved in ABA-responsive gene expression.Each AREB protein contained a single bZIP-type DNA-binding domain, and expression ofAREB1andAREB2was up-regulated by ABA, drought, and high-salinity stresses, shown to function as trans-acting activators by using transient expression in protoplasts

Expression

Expression patterns of the OsAREB1 gene under various environmental stresses and hormones were analyzed by RT-PCR. OsAREB1 gene was induced within 1 or 2 h under 100 μM ABA and 15% PEG 6,000 treatments, and maintained the expression level for at least 8 hours. It’s expression was induced by heat within 1 h, and rapidly reached the top expression level within 2 h, then declined to initial level. OsAREB1 was not induced by KT, MeJA, NaCl and cold .These results indicated that OsAREB1was induced by exogenous ABA, water stress and heat. This result was consistent with the report of Lu et al..

Evolution

Please input evolution information here.

Extending Knowledge

Binding activity

OsAREB1 has ABRE-binding activity in yeast Blast result indicated that OsAREB1 belongs to ABF subfamily. Most members of this subfamily can bind to the ABRE cis-element with a core sequence ACGTGCC. Yeast one-hybrid system was used to determine the DNA-binding activity of OsAREB1 with ABRE element. The entire coding region of OsAREB1 was fused to the GAL4 transcription active domain (TA). The construct was transformed into yeast (EGY48) harboring ABRE sequence fused upstream of a lacZreporter gene, and the growth status of transformants was observed. Yeast cells harboring pPC86 and G222 could grow on SD medium lacking Trp, while cells only with G222 could not grow on the selection medium (Fig. 1A). The colony-lift filter assay suggested that OsAREB1 can bind to the ABRE cis-element. Shown as Fig. 1B, when the colony grew on X-gal containing plate, only cells with pPC86-OsAREB1 and G222 turned blue, cells only with G222 or with both G222 and pPC86 did not turn blue. This result indicated that only OsAREB1 can bind to the ABRE cis-element and then active the expression of lacZ gene. Further, quantificational analysis for β-galactosidase activity was performed. Compared to the negative control, the relative β-galactosidase activity of the transformants was about four (Fig. 1C), which revealed there’s a distinct enhancement for β-galactosidase activity.

AREB regulon

Abscisic Acid acts as a crucial signal molecule in abiotic stress responses (Fujita et al. 2011). The ABA content is increased by abiotic stresses, and leads to expression of numerous genes. Application of exogenous ABA also stimulates a myriad of genes. ABRE was identified as a cis-acting element conserved in promoter regions of ABA-inducible genes.ArabidopsiscDNAs that encode bZIP-type TFs were screened as ABRE-binding proteins (Yamaguchi-Shinozaki & Shinozaki 2006). Among these genes,AREB1/ABF2, AREB2/ABF4,andABF3were reported to be induced by ABA and osmotic stress in vegetative tissues (Fujita et al. 2011,). Evidence indicates that activation of AREB1 needs ABA-dependent posttranscriptional modification. The ABA-activated SnRK2 protein kinases phosphorylate the AREB1 protein (Furihata et al. 2006). TransgenicArabidopsisplants overexpressing the phosphorylated active form of AREB1 showed enhanced expression of a number of ABA-inducible genes (Furihata et al. 2006). The ABA-activated phosphorylation of AREB/ABFs was completely impaired in the SnRK2 triple mutant, srk2d srk2e srk2i (Fujii et al. 2009,; Fujii & Zhu 2009). The down-regulated genes in the srk2d srk2e srk2i andareb1 areb2 abf3triple mutants largely overlapped in ABA-dependent expression, which supports the view that SRK2D/ E/I regulate AREBs in ABA signaling in response to osmotic stress. (Fujita et al. 2009). The rice TRANSCRIPTION FACTOR RESPONSIBLE FOR ABA REGULATION1 (TRAB1) shows high homology toArabidopsisAREB2/ABF4. Expression ofTRAB1 was up-regulated by ABA treatment (Hobo et al. 1999,). TRAB1 is phosphorylated rapidly in response to ABA treatment (Kagaya et al. 2002).

activation mechanisms of AREB1

Kanget al. (17) reported that overexpression of ABF3 and ABF4/AREB2 resulted in ABA-hypersensitive phenotypes in germination and seedling growth stages in Arabidopsis. These transgenic plants also showed improvement of drought stress tolerance, suggesting that AREB/ABF proteins are involved in ABA response and stress tolerance in plants. However, AREB1 and AREB2 require ABA for their maximum activation, as shown by their low transactivation abilities in protoplasts prepared from the ABA-deficient aba2 mutant (7). We have shown that the ABA-responsive 42-kDa kinase activities phosphorylate conserved regions of AREBs, suggesting that ABA-dependent phosphorylation may be involved in activation of the AREB subfamily proteins (7). Phosphorylation/dephosphorylation-regulated events were reported to play important roles in ABA signaling; SNF1-related protein kinase homologues, ABA-activated protein kinase (AAPK) inVicia faba(18), and SRK2E/OST1 inArabidopsis(19, 20) modulate ABA-dependent stomatal closure.ABI1andABI2,of which dominant-negative mutation causes ABA-insensitive mutantsabi1andabi2, encode a type 2C protein phosphatase (4, 5, 21). Because null mutations of ABI1 and ABI2 resulted in ABA hypersensitivity, ABI1 and ABI2 negatively regulate ABAdependent responses (22). Here, we report that the ABA-activated 42-kDa kinase activity phosphorylates Ser/Thr residues in the conserved R-X-X-S/Tsites of AREB1. Amino acid substitution of the Ser/Thr residues to Ala and Asp resulted in suppression and high transactivation activity, respectively. A phosphorylated active form of AREB1 was obtained by substitution of Ser/Thr to Asp in all conserved R-X-XS/T sites. We show that transgenic plants overexpressing the phosphorylated active form AREB1 express not only ABA inducible genes, such as RD29B, but also seed-specific gene without ABA treatment. We also discuss the activation mechanism of AREB1 by ABA-dependent phosphorylation in plants.

Fig. 2 shows the properties of AREB1 protein fragment phosphorylation.

Fig. 2. Properties of AREB1 protein fragment phosphorylation. (A) Effects of protein kinase inhibitor staurosporine on phosphorylation of the recombinant AREB1a polypeptide. Staurosporine (20 and 100 nM) was added to reaction mixture. (B) Effects of BAPTA on phosphorylation of the recombinant AREB1b polypeptide. Na-BAPTA (5 mM final concentration) was added to reaction buffer and equilibrated for 30 min before addition of radiolabeled ATP. In the far right lane, additional kinase activity appeared near 60 kDa (see Resultsfor details). (C) Stress-dependent phosphorylation of the recombinant AREB1b polypeptide. Protein extracts prepared from T87 cells treated for 30 min with 50/M ABA (Ab), 0.5 M NaCl (Na), and 0.8 M mannitol (Os, high osmolality) and at low temperature (Lt, 4°C) or untreated (Ct) were used for in-gel kinase activity assay. The recombinant AREB1b polypeptide was used as a substrate. (D) ABA-activated SnRK2-type protein kinases phosphorylate the recombinant AREB1b polypeptide. Protein extracts prepared from transgenic T87 cells overexpressing each SnRK2-GFP fusion protein under control of the CaMV 35Spromoter were used for in-gel kinase activity assay. Phosphorylated bands derived from SnRK2-GFP fusion proteins were indicated by circles. Arrowheads indicate the position of 42 kDa in A–D.

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Labs working on this gene

1 Biotechnology Research Institute, Shanghai Academy of Agricultural Sciences, Shanghai, 2 College of Life Science and Technology, Yangzhou University, Jiangsu, PR China 3

• Biological Resources Division, Japan International Research Center for Agricultural Sciences, Tsukuba, Ibaraki 305-8686, Japan;

† Laboratory of Plant Molecular Biology, RIKEN Tsukuba Institute, Tsukuba, Ibaraki 305-0074, Japan;¶

Graduate School of Agricultural and Life Sciences, University of Tokyo, Tokyo 113-8657, Japan; ‡ RIKEN Plant Science Center, Yokohama, Kanagawa 203-0045, Japan; and Core Research for Evolutional Science and Technology, Japan Science and Technology Agency, Kawaguchi, Saitama 332-0012, Japan

References

Lu, G. J., Gao, C. X., Zheng, X. N. and Han, B. (2009) Identification of OsbZIP72 as a positive regulator of ABA response and drought tolerance in rice. Planta 229, 605-615.

Daisuke Todaka 1, Kazuo Nakashima1, Kazuo Shinozaki2and Kazuko Yamaguchi-Shinozaki1,3*(2012) Toward understanding transcriptional regulatorynetworks in abiotic stress responses and tolerance in rice Rice 2012,5:6

Takashi Furihata*, Kyonoshin Maruyama*, Yasunari Fujita*, Taishi Umezawa†‡, Riichiro Yoshida†, Kazuo Shinozaki†‡§,and Kazuko Yamaguchi-Shinozaki*§¶(2005) Abscisic acid-dependent multisite phosphorylation regulates the activity of a transcription activator AREB1.PNAS February 7, 2006 vol. 103 no. 6

Jin XF1, Xiong AS, Peng RH, Liu JG, Gao F, Chen JM, Yao QH. (2010) OsAREB1, an ABRE-binding protein responding to ABA and glucose, has multiple functions in Arabidopsis.BMB Rep 43(1):34-9.

Structured Information

Gene Name	Os06g0211200
Description	Similar to Abscisic acid responsive elements-binding factor (ABA-responsive element binding protein 1) (AREB1)
Version	NM_001063653.1 GI:115467037 GeneID:4340462
Length	4829 bp
Definition	Oryza sativa Japonica Group Os06g0211200, complete gene.
Source	Oryza sativa Japonica Group ORGANISM Oryza sativa Japonica Group Eukaryota; Viridiplantae; Streptophyta; Embryophyta; Tracheophyta; Spermatophyta; Magnoliophyta; Liliopsida; Poales; Poaceae; BEP clade; Ehrhartoideae; Oryzeae; Oryza.
Chromosome	Chromosome 6
Location	Chromosome 6:5676158..5680986
Sequence Coding Region	5676361..5677152,5677846..5677917,5678514..5678543,5680499..5680575,5680665..5680668
Expression	GEO Profiles:Os06g0211200
Genome Context	<gbrowseImage1> name=NC_008399:5676158..5680986 source=RiceChromosome06 preset=GeneLocation </gbrowseImage1>
Gene Structure	<gbrowseImage2> name=NC_008399:5676158..5680986 source=RiceChromosome06 preset=GeneLocation </gbrowseImage2>
Coding Sequence	<cdnaseq>atggagttgccggcggatgggagcgcgctggcgaggcaggggtcgatctactcgctgacgttcgacgagttccagagcgcgctgggaagcgccgagaaggatttcgggtcgatgaacatggatgagctgctgcgcaacatctggacggcggaggagtcgcaggccatagcgccggcggcggcggctgcttcggcggcggcggtggttggggacgcgcagcagcagcagcagccgatccagaggcaggggtcgctgacgctgccacgcacgctgagccagaagacggtggacgaggtgtggcgcgacatcatgggcttgggcggcagcgacgacgaagaccccgcggcggcggcggctgcggcggcgcccgcgcagcggcagccgacgctgggggagatgacgctggaggagttcctggtgcgggccggcgtcgtgcgggaggacatggggcagaccatcgtgctgccgccgcaggcgcaggcgttgttccccgggagcaatgtggtcgccccggccatgcagctcgccaacgggatgctgcctggtgtcgtcggcgtcgcccccggcgccgccgccgcgatgacggtggcggcgccggccacgccggtggtgctgaacgggctggggaaggtggagggcggggatctctcgtcgctctcgccggtgccttacccattcgacaccgcgctcagggtgaggaagggccctaccgtcgagaaggtggtggagaggcggcagaggcggatgatcaagaacagggagtccgctgctaggtctcgcgcgcggaagcaggcttatataatggagttggaagctgaggtggcaaaactgaaggaacagaaggctgaattgcagaaaaagcaggtggaaatgatacagaagcaaaatgatgaggtcatggagagaatcactcagcaacttggaccaaaggcaaagagattttgcctccgacgaacactgactggtccatgctga</cdnaseq>
Protein Sequence	<aaseq>MELPADGSALARQGSIYSLTFDEFQSALGSAEKDFGSMNMDELL RNIWTAEESQAIAPAAAAASAAAVVGDAQQQQQPIQRQGSLTLPRTLSQKTVDEVWRD IMGLGGSDDEDPAAAAAAAAPAQRQPTLGEMTLEEFLVRAGVVREDMGQTIVLPPQAQ ALFPGSNVVAPAMQLANGMLPGVVGVAPGAAAAMTVAAPATPVVLNGLGKVEGGDLSS LSPVPYPFDTALRVRKGPTVEKVVERRQRRMIKNRESAARSRARKQAYIMELEAEVAK LKEQKAELQKKQVEMIQKQNDEVMERITQQLGPKAKRFCLRRTLTGPC</aaseq>
Gene Sequence	<dnaseqindica>204..995#1689..1760#2357..2386#4342..4418#4508..4511#gattaaacctgatttccccttgctaattcgggccatcgcatcactcccccaactaatcacactcctctcttctccgcttcctcttctcgtatatttataaccccacttcccttttcttcctctttcttctcatcttggtttcttcctagtttcgggagaggattttagtgagggatttgaaggattttgaggtgggagaggagatggagttgccggcggatgggagcgcgctggcgaggcaggggtcgatctactcgctgacgttcgacgagttccagagcgcgctgggaagcgccgagaaggatttcgggtcgatgaacatggatgagctgctgcgcaacatctggacggcggaggagtcgcaggccatagcgccggcggcggcggctgcttcggcggcggcggtggttggggacgcgcagcagcagcagcagccgatccagaggcaggggtcgctgacgctgccacgcacgctgagccagaagacggtggacgaggtgtggcgcgacatcatgggcttgggcggcagcgacgacgaagaccccgcggcggcggcggctgcggcggcgcccgcgcagcggcagccgacgctgggggagatgacgctggaggagttcctggtgcgggccggcgtcgtgcgggaggacatggggcagaccatcgtgctgccgccgcaggcgcaggcgttgttccccgggagcaatgtggtcgccccggccatgcagctcgccaacgggatgctgcctggtgtcgtcggcgtcgcccccggcgccgccgccgcgatgacggtggcggcgccggccacgccggtggtgctgaacgggctggggaaggtggagggcggggatctctcgtcgctctcgccggtgccttacccattcgacaccgcgctcagggtgaggaagggccctaccgtcgagaaggtggtggagaggcggcagaggcggatgatcaagaacagggagtccgctgctaggtctcgcgcgcggaagcaggtgaagcctctcttctcttcaactgcactaggatgtaggaatacgtagcaatcttatgtccatttgcttgattaattagttctgaaaattcgatggtgcctatatttggtatgcctctcgataatgctgtactttattcacatgatgtgatccccccacttctaattcagcttgtagatgttaatttatgcctaatagccactgcaaatagcaagttagcgtcgttaaatattgttcaaccacagaggataagaatctaaagtgaataagccatggttcaagttgatgctctcagattcagagagatgatgagtggctttgttcatttcgggacactggctgagcggtgtttttttttttttttggatcgactattgctgtgatagggatagcatgctcaccatgaagcttggaaggacaattgacaagaccaattcgagaaatagtaacttccgttgatcttttctttaaaaaaaaaacttgtgggggtaataggtcttttttatgtgcaagttgtgctgccatgatgcactcatacttctataaaagctagtatatacttttattgttctaacatagcaaggtcaagtctttctgatactctgattgacagagaatagttctagacagttatggttgtgttgcgaaacacaattttttaacattaaattttggctttctgatatacaacaggcttatataatggagttggaagctgaggtggcaaaactgaaggaacagaaggctgaattgcagaaaaagcaggtatacctgctgtcatataaaattgctttgatccatgcatactctttatttttttctgtttctcttaccaattcctgtaatcagttagtagtccttagataacctcttgactttggataattcctatgtttcttgcctcgattgtcatatttgtttggggatttgggcttaccaatggctgttgttttaatctacccccagcaatgcttgttgctggtattgcaatatgtaggtgaccacaaatagcattcaaatgtccttgttctatgtatttcctgtggaatttatctgagttcaaatctttaatggttgttggaggtttgcacttaaaaggtgtatccctttttaatttgaacattgatggggttacattaattttgtattactgtcctgcaaacttgtattgaaatcctatgccatctggtatcttcttgttcacacattttccatgtgcctactattgttatgccagtatgtcattgtatcatgattttgtaattgaataacttaacaaaagcaccaaaccttttcttcttccaaattcctgacaaaaaatccatgaagctttattcacttattatgcttctaatttgcaggtggaaatgatacagaagcaaaatgatgaggtaatgaccttacagttggtagtaaataccatggttctgaattcgcactatgtttgtcctcattgatgtggaagttgtctatacctctttttgatcataccatactttcttcttttttaaataaaaaacaagagtaagttcatttccacaaacttgctgattaagggatctttggatctgggtgaattttttggggcatttgcaagtttggactaaagttctgatgacttgaatccagataccctctatatccaaacagacccaaattggggcaaaacaacaagctaactctatcgaaccaattcgtttttcttgctgtgagcctttcctaaaactggcagacgccttagaatctccatggctgaagtttgtcaaagtgcacaacaacagtagcattaggtcatttattagagctgcatgacattcaaattctttttaatgagctgcacaacaacatgcacgatttcccaattctggctgtgaggtgtgtttgttattgcattttttgtggtcatgtcagaaaatatcttactacatatggtttaaattatgtttcagaaatcccttattctttaaccaactgccatcataaatgtatatgctggtgttggccatatatttcatgtcactgacactctggcctcttatcccttctcattttcattcatgtatcatgttatctatgtagggtcattgtgttacatccatcttcagttttttaacactttagtgccatccacttgtggttttttttttcataaaataaatttgaagaaactaacagagatatgcaatactggaactcctgccattgcaatagaaaattagcacatcatgattttcagctatcacagtgtggattactgtgatgtgttttttgttaaaaaaaaatgtacactactttgtcctacactcgtccctgctgtcaaataattgagtttgccatgtacagagtaaatgtccagatgtgcaccattctgaatatggtatcccattttgggcatgcatgcttgtaacaccaaaattctgttggagaacatactacccaataagctcaaggtttcattttcaaaatctagttatctgtaattcccatgtcattatagttaaagtatttaattttctaccttctgttttatttttaactaatcagttcaattttttaaccaaagaatgatactcttttaccagggagatctgaagtgacttattggacatacaatgctaaaacaacaaactaaaacaacaaaatggacattgggtaggtttgtctgggtggtatgctcaacccagtctgatctggagtattaaaaagaatcctgactctaaatttagtccaccgctactatagacgctatagtgagcctgtccagcattcaaattaaggacattagttgtgttcgtttctaatggttgggaaccttccccctctagcatgtaaaacggagcaacgatttagcacacgatcaattaagtattagctaaaaaaaacttgaaaaaatggattaatataattttttaaaccaacttttctatagaaagtttttctagaaaacacattgtttagcagttttggaagcgtgcgtgcggaaaacaagtgagtagaggtgggaaagtgtagggaagatgtcatgttggtttgaattttgaaagtctgcaaactcttactaatgttacataaatagttttggttgctactctgcggtttcaagcagtaaacctctgcctcaatcattctgaagtttaagcatttctttgattataatcagtacagttagaaggccttaactgggtgcatttttatgttccgacttctgacgatctcactgaaatctgacaagtgttttctagctgaagatagtacattttactagatgctttgcaatgttgagaaagttctcttcatagattcttctttccctaacaggagttttatctataaatggattttgcaggtcatggagagaatcactcagcaacttggaccaaaggcaaagagattttgcctccgacgaacactgactggtccatggtaagttgatcaagtttgcacagcattgaccaaagttaagatcgttggcttccttggatgtgaccaatcgccgcgtgtatatatatcagctgaagccagagtctccggtttcgccgtggcgctcagcttcagagttgcttctctccttgttggtgaatggtgatggctcagtctcttggacggtcgaatgctggcgtcgattactcactaggtttagctgcgagatcgttgcgtgagcaaaggcatactatctaatctgtttaactccttatttagggaaatctggcatggtgaaaacggggcatgccatctgtgtttgttgtttttgtgcagctgttgcatctgctctgtatgttgctgttgcgttgacatgtcatcccgtttacagttcagtgattctgttctgtaccc</dnaseqindica>
External Link(s)	NCBI Gene:Os06g0211200, RefSeq:Os06g0211200