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Serine proteases-like genes in the asian rice gall midge show differential expression in compatible and incompatible interactions with rice.

Sinha DK, Lakshmi M, Anuradha G, Rahman SJ, Siddiq EA, Bentur JS, Nair S - Int J Mol Sci (2011)

Bottom Line: Quantitative real time PCR analysis revealed that both the genes were significantly upregulated in larvae feeding on resistant cultivar than in those feeding on susceptible cultivar.These results provide an opportunity to understand the gut physiology of the insect under compatible or incompatible interactions with the host.Phylogenetic analysis grouped these genes in the clade containing proteases of phytophagous insects away from hematophagous insects.

View Article: PubMed Central - PubMed

Affiliation: Plant Molecular Biology Group, International Centre for Genetic Engineering and Biotechnology, Aruna Asaf Ali Marg, New Delhi 110067, India; E-Mail: deepak22sinha@yahoo.co.in.

ABSTRACT
The Asian rice gall midge, Orseolia oryzae (Wood-Mason), is a serious pest of rice. Investigations into the gall midge-rice interaction will unveil the underlying molecular mechanisms which, in turn, can be used as a tool to assist in developing suitable integrated pest management strategies. The insect gut is known to be involved in various physiological and biological processes including digestion, detoxification and interaction with the host. We have cloned and identified two genes, OoprotI and OoprotII, homologous to serine proteases with the conserved His(87), Asp(136) and Ser(241) residues. OoProtI shared 52.26% identity with mosquito-type trypsin from Hessian fly whereas OoProtII showed 52.49% identity to complement component activated C1s from the Hessian fly. Quantitative real time PCR analysis revealed that both the genes were significantly upregulated in larvae feeding on resistant cultivar than in those feeding on susceptible cultivar. These results provide an opportunity to understand the gut physiology of the insect under compatible or incompatible interactions with the host. Phylogenetic analysis grouped these genes in the clade containing proteases of phytophagous insects away from hematophagous insects.

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Related in: MedlinePlus

Transcript levels of OoprotI (A) and OoprotII (B) of the rice gall midge feeding on compatible host Jaya (white bars) and incompatible host RP2068 (black bars) 24, 48, 72 and 96 h after hatching. Two biological replicates and two technical replicates were taken for the study. Asterisk (*) indicates no detectable amplification. Error bars represent mean ±SD.
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f3-ijms-12-02842: Transcript levels of OoprotI (A) and OoprotII (B) of the rice gall midge feeding on compatible host Jaya (white bars) and incompatible host RP2068 (black bars) 24, 48, 72 and 96 h after hatching. Two biological replicates and two technical replicates were taken for the study. Asterisk (*) indicates no detectable amplification. Error bars represent mean ±SD.

Mentions: Based on the quantitative Real Time (RT) PCR we observed that both OoProtI (Figure 3A) and OoProtII (Figure 3B) expressed in the larvae examined at 48, 72 and 96 h post hatching, independent of the fact that they were feeding on the resistant (RP2068) or the susceptible (Jaya) plants with highest expression observed at 96 h. However, the expression level of OoProtI was three-fold higher at 96 h when feeding on a resistant cultivar compared with larvae feeding on the susceptible cultivar. In the case of the expression pattern of OoProtII, we observed a similar pattern. Here too the expression of OoProtII was the highest at 96 h and the expression of this transcript was approximately one and a half times higher at 96 h in the larvae feeding on the resistant cultivar as compared to larvae on the susceptible cultivar. It should be noted that in the case of OoProtI, we did not observe any amplification in 24 h post-hatch larvae even after repeating the PCR amplifications on several batches of larvae from this stage. Owing to this reason, amplification at 48 h time point was used as calibrator for OoProtI but for the case of OoProtII 24 h post-hatch time point was used as the calibrator. It may be noted that trypsins hydrolyze peptide bonds involving amino acids with positively charged side chains, such as arginine and lysine, whereas chymotrypsins cleave peptide bonds on the carboxyl terminus of aromatic amino acids (tryptophan, tyrosine and phenylalanine) [19]. It has also been reported that functional diversity of encoded enzymes and differential ability to hydrolyze ingested proteins may account for the regulatory dexterity evident from differential gene expression of multiple, sequence divergent midgut proteases [20]. The observation that expression levels of both OoProtI and OoProtII were the highest at 96 h and when feeding on the resistant cultivar, could be due to the response of the larvae to the possible presence of PI and/or other toxins. The PI could be inducing their up-regulation and/or the de-novo synthesis promoting changes in key amino acids that help these two proteases resist the inhibitors [21]. However, this needs to be confirmed. Although both OoProtI and II were overexpressed in larvae feeding on the resistant cultivar, the larvae eventually succumb.


Serine proteases-like genes in the asian rice gall midge show differential expression in compatible and incompatible interactions with rice.

Sinha DK, Lakshmi M, Anuradha G, Rahman SJ, Siddiq EA, Bentur JS, Nair S - Int J Mol Sci (2011)

Transcript levels of OoprotI (A) and OoprotII (B) of the rice gall midge feeding on compatible host Jaya (white bars) and incompatible host RP2068 (black bars) 24, 48, 72 and 96 h after hatching. Two biological replicates and two technical replicates were taken for the study. Asterisk (*) indicates no detectable amplification. Error bars represent mean ±SD.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC3116160&req=5

f3-ijms-12-02842: Transcript levels of OoprotI (A) and OoprotII (B) of the rice gall midge feeding on compatible host Jaya (white bars) and incompatible host RP2068 (black bars) 24, 48, 72 and 96 h after hatching. Two biological replicates and two technical replicates were taken for the study. Asterisk (*) indicates no detectable amplification. Error bars represent mean ±SD.
Mentions: Based on the quantitative Real Time (RT) PCR we observed that both OoProtI (Figure 3A) and OoProtII (Figure 3B) expressed in the larvae examined at 48, 72 and 96 h post hatching, independent of the fact that they were feeding on the resistant (RP2068) or the susceptible (Jaya) plants with highest expression observed at 96 h. However, the expression level of OoProtI was three-fold higher at 96 h when feeding on a resistant cultivar compared with larvae feeding on the susceptible cultivar. In the case of the expression pattern of OoProtII, we observed a similar pattern. Here too the expression of OoProtII was the highest at 96 h and the expression of this transcript was approximately one and a half times higher at 96 h in the larvae feeding on the resistant cultivar as compared to larvae on the susceptible cultivar. It should be noted that in the case of OoProtI, we did not observe any amplification in 24 h post-hatch larvae even after repeating the PCR amplifications on several batches of larvae from this stage. Owing to this reason, amplification at 48 h time point was used as calibrator for OoProtI but for the case of OoProtII 24 h post-hatch time point was used as the calibrator. It may be noted that trypsins hydrolyze peptide bonds involving amino acids with positively charged side chains, such as arginine and lysine, whereas chymotrypsins cleave peptide bonds on the carboxyl terminus of aromatic amino acids (tryptophan, tyrosine and phenylalanine) [19]. It has also been reported that functional diversity of encoded enzymes and differential ability to hydrolyze ingested proteins may account for the regulatory dexterity evident from differential gene expression of multiple, sequence divergent midgut proteases [20]. The observation that expression levels of both OoProtI and OoProtII were the highest at 96 h and when feeding on the resistant cultivar, could be due to the response of the larvae to the possible presence of PI and/or other toxins. The PI could be inducing their up-regulation and/or the de-novo synthesis promoting changes in key amino acids that help these two proteases resist the inhibitors [21]. However, this needs to be confirmed. Although both OoProtI and II were overexpressed in larvae feeding on the resistant cultivar, the larvae eventually succumb.

Bottom Line: Quantitative real time PCR analysis revealed that both the genes were significantly upregulated in larvae feeding on resistant cultivar than in those feeding on susceptible cultivar.These results provide an opportunity to understand the gut physiology of the insect under compatible or incompatible interactions with the host.Phylogenetic analysis grouped these genes in the clade containing proteases of phytophagous insects away from hematophagous insects.

View Article: PubMed Central - PubMed

Affiliation: Plant Molecular Biology Group, International Centre for Genetic Engineering and Biotechnology, Aruna Asaf Ali Marg, New Delhi 110067, India; E-Mail: deepak22sinha@yahoo.co.in.

ABSTRACT
The Asian rice gall midge, Orseolia oryzae (Wood-Mason), is a serious pest of rice. Investigations into the gall midge-rice interaction will unveil the underlying molecular mechanisms which, in turn, can be used as a tool to assist in developing suitable integrated pest management strategies. The insect gut is known to be involved in various physiological and biological processes including digestion, detoxification and interaction with the host. We have cloned and identified two genes, OoprotI and OoprotII, homologous to serine proteases with the conserved His(87), Asp(136) and Ser(241) residues. OoProtI shared 52.26% identity with mosquito-type trypsin from Hessian fly whereas OoProtII showed 52.49% identity to complement component activated C1s from the Hessian fly. Quantitative real time PCR analysis revealed that both the genes were significantly upregulated in larvae feeding on resistant cultivar than in those feeding on susceptible cultivar. These results provide an opportunity to understand the gut physiology of the insect under compatible or incompatible interactions with the host. Phylogenetic analysis grouped these genes in the clade containing proteases of phytophagous insects away from hematophagous insects.

Show MeSH
Related in: MedlinePlus