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Identification of Multiple Stress Responsive Genes by Sequencing a Normalized cDNA Library from Sea-Land Cotton (Gossypium barbadense L.).

Zhou B, Zhang L, Ullah A, Jin X, Yang X, Zhang X - PLoS ONE (2016)

Bottom Line: The blastx results demonstrated 2,746 unigenes showing significant similarity to known genes, 74 uniESTs displaying significant similarity to genes of predicted proteins, and 315 uniESTs remain uncharacterized.Annotation results showed that a huge number of genes respond to stress in our study, such as MYB-related, C2H2, FAR1, bHLH, bZIP, MADS, and mTERF.These results will improve our knowledge of stress tolerance in cotton.

View Article: PubMed Central - PubMed

Affiliation: National Key Laboratory of Crop Genetic Improvement, Huazhong Agricultural University, Wuhan, Hubei 430070, P. R. China.

ABSTRACT

Background: Plants often face multiple stresses including drought, extreme temperature, salinity, nutrition deficiency and biotic stresses during growth and development. All the stresses result in a series of physiological and metabolic reactions and then generate reversible inhibition of metabolism and growth and can cause seriously irreversible damage, even death. At each stage of cotton growth, environmental stress conditions pose devastating threats to plant growth and development, especially yield and quality. Due to the complex stress conditions and unclear molecular mechanisms of stress response, there is an urgent need to explore the mechanisms of cotton response against abiotic stresses.

Methodology and principal findings: A normalized cDNA library was constructed using Gossypium barbadense Hai-7124 treated with different stress conditions (heat, cold, salt, drought, potassium and phosphorus deficit and Verticillium dahliae infection). Random sequencing of this library generated 6,047 high-quality expressed sequence tags (ESTs). The ESTs were clustered and assembled into 3,135 uniESTs, composed of 2,497 contigs and 638 singletons. The blastx results demonstrated 2,746 unigenes showing significant similarity to known genes, 74 uniESTs displaying significant similarity to genes of predicted proteins, and 315 uniESTs remain uncharacterized. Functional classification unveiled the abundance of uniESTs in binding, catalytic activity, and structural molecule activity. Annotations of the uniESTs by the plant transcription factor database (PlantTFDB) and Plant Stress Protein Database (PSPDB) disclosed that transcription factors and stress-related genes were enriched in the current library. The expression of some transcription factors and specific stress-related genes were verified by RT-PCR under various stress conditions.

Conclusions/significance: Annotation results showed that a huge number of genes respond to stress in our study, such as MYB-related, C2H2, FAR1, bHLH, bZIP, MADS, and mTERF. These results will improve our knowledge of stress tolerance in cotton. In addition, they are also helpful in discovering candidate genes related to stress tolerance. The publicly available ESTs from G. barbadense are a valuable genomic resource that will facilitate further molecular study and breeding of stress-tolerant cotton.

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Related in: MedlinePlus

RT-PCR analysis of selected genes under different abiotic stress environments.0, 1, 2, 4, and 8 refer to samples from different time points (0 h, 1 h, 2 h, 4 h, and 8 h) after treatment by different stress factors (200 mM ABA as ABA, 15% PEG as PEG, 200 mmol L-1 NaCl as NaCl and 4°C as cold).
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pone.0152927.g007: RT-PCR analysis of selected genes under different abiotic stress environments.0, 1, 2, 4, and 8 refer to samples from different time points (0 h, 1 h, 2 h, 4 h, and 8 h) after treatment by different stress factors (200 mM ABA as ABA, 15% PEG as PEG, 200 mmol L-1 NaCl as NaCl and 4°C as cold).

Mentions: In accordance with previous studies, ABA is a widely studied phytohormone, and its role in ameliorating abiotic stress in plants is well established [39]. There are three major components of ABA signaling: pyrabactin resistance (PYR)/PYR1-like (PYL)/regulatory component of the ABA receptor (RCAR), protein phosphatase 2C (PP2C; a negative regulator) and SNF (sucrose non-fermenting) 1-related protein kinase 2 (SnRK2: a positive regulator). These are gathered as a double negative regulatory system and form a signaling complex known as the ‘ABA signalosome’ [40]. Negative regulation of MAPK (MAPK4 and MAPK6) by A. thaliana PP2C, AP2C1 also potentially links PP2Cs to cold and drought stress responses [41]. The SnRK2 family members are the key regulators of plant responses to multiple abiotic stresses. SnRK2s (40 kDa) are monomeric serine/threonine protein kinases. Major transcription factor families, which are concerned with the regulation of multiple abiotic stress responses, include bZIP, MYB, MYC, NAC, ERF and DREB/CBF (C repeat binding factor). From all the stress-related genes in the cDNA library, 45 genes, including three genes in the ABA pathway, one WRKY gene, one MYB and MYB-related gene, one AP2-EREBP gene, two auxin response factors, one brassinosteroid-regulated family protein, two cytokinin genes, two ethylene-responsive factors, two gibberellin-related genes, one gene responsive to jasmonic acid and salicylic acid, two JAZ genes, one NAC gene, one bHLH gene, one bZIP gene, one FAR1 gene, one LEA family member, 8 redox-related genes, two AMPK family members, two BSK family members, three CDPK family members, one CIPK family member and four SNF1 family members were chosen for validation by RT-PCR. The main results corresponded to the data from the transcription profiles, as depicted in Fig 7. In our results, the expression of DREB2 is up-regulated in cold treatment after 4 h; NAC029 is up-regulated in PEG, NaCl and cold treatments; and TGA6 is down-regulated in PEG and NaCl treatments, but is up-regulated in cold treatment. CIPK11 is up-regulated in ABA treatment, GSH-PX8 is up-regulated in PEG treatment, ABAH1 is up-regulated in ABA and cold treatments, and ABF1 is especially up-regulated in all the four treatments (ABA, PEG, NaCl and cold).


Identification of Multiple Stress Responsive Genes by Sequencing a Normalized cDNA Library from Sea-Land Cotton (Gossypium barbadense L.).

Zhou B, Zhang L, Ullah A, Jin X, Yang X, Zhang X - PLoS ONE (2016)

RT-PCR analysis of selected genes under different abiotic stress environments.0, 1, 2, 4, and 8 refer to samples from different time points (0 h, 1 h, 2 h, 4 h, and 8 h) after treatment by different stress factors (200 mM ABA as ABA, 15% PEG as PEG, 200 mmol L-1 NaCl as NaCl and 4°C as cold).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4816313&req=5

pone.0152927.g007: RT-PCR analysis of selected genes under different abiotic stress environments.0, 1, 2, 4, and 8 refer to samples from different time points (0 h, 1 h, 2 h, 4 h, and 8 h) after treatment by different stress factors (200 mM ABA as ABA, 15% PEG as PEG, 200 mmol L-1 NaCl as NaCl and 4°C as cold).
Mentions: In accordance with previous studies, ABA is a widely studied phytohormone, and its role in ameliorating abiotic stress in plants is well established [39]. There are three major components of ABA signaling: pyrabactin resistance (PYR)/PYR1-like (PYL)/regulatory component of the ABA receptor (RCAR), protein phosphatase 2C (PP2C; a negative regulator) and SNF (sucrose non-fermenting) 1-related protein kinase 2 (SnRK2: a positive regulator). These are gathered as a double negative regulatory system and form a signaling complex known as the ‘ABA signalosome’ [40]. Negative regulation of MAPK (MAPK4 and MAPK6) by A. thaliana PP2C, AP2C1 also potentially links PP2Cs to cold and drought stress responses [41]. The SnRK2 family members are the key regulators of plant responses to multiple abiotic stresses. SnRK2s (40 kDa) are monomeric serine/threonine protein kinases. Major transcription factor families, which are concerned with the regulation of multiple abiotic stress responses, include bZIP, MYB, MYC, NAC, ERF and DREB/CBF (C repeat binding factor). From all the stress-related genes in the cDNA library, 45 genes, including three genes in the ABA pathway, one WRKY gene, one MYB and MYB-related gene, one AP2-EREBP gene, two auxin response factors, one brassinosteroid-regulated family protein, two cytokinin genes, two ethylene-responsive factors, two gibberellin-related genes, one gene responsive to jasmonic acid and salicylic acid, two JAZ genes, one NAC gene, one bHLH gene, one bZIP gene, one FAR1 gene, one LEA family member, 8 redox-related genes, two AMPK family members, two BSK family members, three CDPK family members, one CIPK family member and four SNF1 family members were chosen for validation by RT-PCR. The main results corresponded to the data from the transcription profiles, as depicted in Fig 7. In our results, the expression of DREB2 is up-regulated in cold treatment after 4 h; NAC029 is up-regulated in PEG, NaCl and cold treatments; and TGA6 is down-regulated in PEG and NaCl treatments, but is up-regulated in cold treatment. CIPK11 is up-regulated in ABA treatment, GSH-PX8 is up-regulated in PEG treatment, ABAH1 is up-regulated in ABA and cold treatments, and ABF1 is especially up-regulated in all the four treatments (ABA, PEG, NaCl and cold).

Bottom Line: The blastx results demonstrated 2,746 unigenes showing significant similarity to known genes, 74 uniESTs displaying significant similarity to genes of predicted proteins, and 315 uniESTs remain uncharacterized.Annotation results showed that a huge number of genes respond to stress in our study, such as MYB-related, C2H2, FAR1, bHLH, bZIP, MADS, and mTERF.These results will improve our knowledge of stress tolerance in cotton.

View Article: PubMed Central - PubMed

Affiliation: National Key Laboratory of Crop Genetic Improvement, Huazhong Agricultural University, Wuhan, Hubei 430070, P. R. China.

ABSTRACT

Background: Plants often face multiple stresses including drought, extreme temperature, salinity, nutrition deficiency and biotic stresses during growth and development. All the stresses result in a series of physiological and metabolic reactions and then generate reversible inhibition of metabolism and growth and can cause seriously irreversible damage, even death. At each stage of cotton growth, environmental stress conditions pose devastating threats to plant growth and development, especially yield and quality. Due to the complex stress conditions and unclear molecular mechanisms of stress response, there is an urgent need to explore the mechanisms of cotton response against abiotic stresses.

Methodology and principal findings: A normalized cDNA library was constructed using Gossypium barbadense Hai-7124 treated with different stress conditions (heat, cold, salt, drought, potassium and phosphorus deficit and Verticillium dahliae infection). Random sequencing of this library generated 6,047 high-quality expressed sequence tags (ESTs). The ESTs were clustered and assembled into 3,135 uniESTs, composed of 2,497 contigs and 638 singletons. The blastx results demonstrated 2,746 unigenes showing significant similarity to known genes, 74 uniESTs displaying significant similarity to genes of predicted proteins, and 315 uniESTs remain uncharacterized. Functional classification unveiled the abundance of uniESTs in binding, catalytic activity, and structural molecule activity. Annotations of the uniESTs by the plant transcription factor database (PlantTFDB) and Plant Stress Protein Database (PSPDB) disclosed that transcription factors and stress-related genes were enriched in the current library. The expression of some transcription factors and specific stress-related genes were verified by RT-PCR under various stress conditions.

Conclusions/significance: Annotation results showed that a huge number of genes respond to stress in our study, such as MYB-related, C2H2, FAR1, bHLH, bZIP, MADS, and mTERF. These results will improve our knowledge of stress tolerance in cotton. In addition, they are also helpful in discovering candidate genes related to stress tolerance. The publicly available ESTs from G. barbadense are a valuable genomic resource that will facilitate further molecular study and breeding of stress-tolerant cotton.

Show MeSH
Related in: MedlinePlus