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Identification of Putative RuBisCo Activase (TaRca1)-The Catalytic Chaperone Regulating Carbon Assimilatory Pathway in Wheat (Triticum aestivum) under the Heat Stress.

Kumar RR, Goswami S, Singh K, Dubey K, Singh S, Sharma R, Verma N, Kala YK, Rai GK, Grover M, Mishra DC, Singh B, Pathak H, Chinnusamy V, Rai A, Praveen S - Front Plant Sci (2016)

Bottom Line: HS causes decrease in the RuBisCo activity; maximum reduction was observed during pollination stage in thermosusceptible cvs. as validated through immunoblotting.We observed uniform carbon distribution in different tissues of thermotolerant cvs., as compared to thermosusceptible.There is, however, need to manipulate the thermal stability of TaRca1 enzyme through protein engineering for sustaining the photosynthetic rate under HS-a novel approach toward development of "climate-smart" crop.

View Article: PubMed Central - PubMed

Affiliation: Division of Biochemistry, Indian Agricultural Research Institute New Delhi, India.

ABSTRACT
RuBisCo activase (Rca) is a catalytic chaperone involved in modulating the activity of RuBisCo (key enzyme of photosynthetic pathway). Here, we identified eight novel transcripts from wheat through data mining predicted to be Rca and cloned a transcript of 1.4 kb from cv. HD2985, named as TaRca1 (GenBank acc. no. KC776912). Single copy number of TaRca1 was observed in wheat genome. Expression analysis in diverse wheat genotypes (HD2985, Halna, PBW621, and HD2329) showed very high relative expression of TaRca1 in Halna under control and HS-treated, as compared to other cultivars at different stages of growth. TaRca1 protein was predicted to be chloroplast-localized with numerous potential phosphorylation sites. Northern blot analysis showed maximum accumulation of TaRca1 transcript in thermotolerant cv. during mealy-ripe stage, as compared to thermosusceptible. Decrease in the photosynthetic parameters was observed in all the cultivars, except PBW621 in response to HS. We observed significant increase in the Rca activity in all the cultivars under HS at different stages of growth. HS causes decrease in the RuBisCo activity; maximum reduction was observed during pollination stage in thermosusceptible cvs. as validated through immunoblotting. We observed uniform carbon distribution in different tissues of thermotolerant cvs., as compared to thermosusceptible. Similarly, tolerance level of leaf was observed maximum in Halna having high Rca activity under HS. A positive correlation was observed between the transcript and activity of TaRca1 in HS-treated Halna. Similarly, TaRca1 enzyme showed positive correlation with the activity of RuBisCo. There is, however, need to manipulate the thermal stability of TaRca1 enzyme through protein engineering for sustaining the photosynthetic rate under HS-a novel approach toward development of "climate-smart" crop.

No MeSH data available.


Related in: MedlinePlus

Southern blot analysis to identify the copy number of cloned RuBisCo activase (TaRca1) gene from wheat cv. HD2985; EcoRI, HinDIII, and BamHI enzymes were used for the restriction; 12 μg genomic DNA was used per sample; 0.8% agarose gel was used for the electrophoretic separation of restricted product; Biotin-labeled probe was used for the hybridization.
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Figure 2: Southern blot analysis to identify the copy number of cloned RuBisCo activase (TaRca1) gene from wheat cv. HD2985; EcoRI, HinDIII, and BamHI enzymes were used for the restriction; 12 μg genomic DNA was used per sample; 0.8% agarose gel was used for the electrophoretic separation of restricted product; Biotin-labeled probe was used for the hybridization.

Mentions: To find out the copy number of the cloned TaRca1 gene from wheat, southern blot analysis was carried out using the genomic DNA isolated from wheat cv. HD2985. We observed single prominent blot in each lane (restricted with EcoRI, HinDIII, and BamHI; Figure 2). EcoRI restricted lane showed band of ~3.2 kb, whereas BamHI and HinDIII showed bands of ~2.8 and ~4.0 kb. This result suggests that cloned TaRca1 has single copy in Triticum aestivum.


Identification of Putative RuBisCo Activase (TaRca1)-The Catalytic Chaperone Regulating Carbon Assimilatory Pathway in Wheat (Triticum aestivum) under the Heat Stress.

Kumar RR, Goswami S, Singh K, Dubey K, Singh S, Sharma R, Verma N, Kala YK, Rai GK, Grover M, Mishra DC, Singh B, Pathak H, Chinnusamy V, Rai A, Praveen S - Front Plant Sci (2016)

Southern blot analysis to identify the copy number of cloned RuBisCo activase (TaRca1) gene from wheat cv. HD2985; EcoRI, HinDIII, and BamHI enzymes were used for the restriction; 12 μg genomic DNA was used per sample; 0.8% agarose gel was used for the electrophoretic separation of restricted product; Biotin-labeled probe was used for the hybridization.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4940427&req=5

Figure 2: Southern blot analysis to identify the copy number of cloned RuBisCo activase (TaRca1) gene from wheat cv. HD2985; EcoRI, HinDIII, and BamHI enzymes were used for the restriction; 12 μg genomic DNA was used per sample; 0.8% agarose gel was used for the electrophoretic separation of restricted product; Biotin-labeled probe was used for the hybridization.
Mentions: To find out the copy number of the cloned TaRca1 gene from wheat, southern blot analysis was carried out using the genomic DNA isolated from wheat cv. HD2985. We observed single prominent blot in each lane (restricted with EcoRI, HinDIII, and BamHI; Figure 2). EcoRI restricted lane showed band of ~3.2 kb, whereas BamHI and HinDIII showed bands of ~2.8 and ~4.0 kb. This result suggests that cloned TaRca1 has single copy in Triticum aestivum.

Bottom Line: HS causes decrease in the RuBisCo activity; maximum reduction was observed during pollination stage in thermosusceptible cvs. as validated through immunoblotting.We observed uniform carbon distribution in different tissues of thermotolerant cvs., as compared to thermosusceptible.There is, however, need to manipulate the thermal stability of TaRca1 enzyme through protein engineering for sustaining the photosynthetic rate under HS-a novel approach toward development of "climate-smart" crop.

View Article: PubMed Central - PubMed

Affiliation: Division of Biochemistry, Indian Agricultural Research Institute New Delhi, India.

ABSTRACT
RuBisCo activase (Rca) is a catalytic chaperone involved in modulating the activity of RuBisCo (key enzyme of photosynthetic pathway). Here, we identified eight novel transcripts from wheat through data mining predicted to be Rca and cloned a transcript of 1.4 kb from cv. HD2985, named as TaRca1 (GenBank acc. no. KC776912). Single copy number of TaRca1 was observed in wheat genome. Expression analysis in diverse wheat genotypes (HD2985, Halna, PBW621, and HD2329) showed very high relative expression of TaRca1 in Halna under control and HS-treated, as compared to other cultivars at different stages of growth. TaRca1 protein was predicted to be chloroplast-localized with numerous potential phosphorylation sites. Northern blot analysis showed maximum accumulation of TaRca1 transcript in thermotolerant cv. during mealy-ripe stage, as compared to thermosusceptible. Decrease in the photosynthetic parameters was observed in all the cultivars, except PBW621 in response to HS. We observed significant increase in the Rca activity in all the cultivars under HS at different stages of growth. HS causes decrease in the RuBisCo activity; maximum reduction was observed during pollination stage in thermosusceptible cvs. as validated through immunoblotting. We observed uniform carbon distribution in different tissues of thermotolerant cvs., as compared to thermosusceptible. Similarly, tolerance level of leaf was observed maximum in Halna having high Rca activity under HS. A positive correlation was observed between the transcript and activity of TaRca1 in HS-treated Halna. Similarly, TaRca1 enzyme showed positive correlation with the activity of RuBisCo. There is, however, need to manipulate the thermal stability of TaRca1 enzyme through protein engineering for sustaining the photosynthetic rate under HS-a novel approach toward development of "climate-smart" crop.

No MeSH data available.


Related in: MedlinePlus