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Multicopper oxidase-1 is required for iron homeostasis in Malpighian tubules of Helicoverpa armigera.

Liu X, Sun C, Liu X, Yin X, Wang B, Du M, An S - Sci Rep (2015)

Bottom Line: HaMCO1 was also found to be highly abundant in Malpighian tubules.HaMCO1 knockdown caused a significant decrease in iron accumulation and also significantly reduced transferrin and ferritin transcript expression.Therefore, HaMCO1 is coordinately regulated by 20E and JH and is required for iron homeostasis in Malpighian tubules.

View Article: PubMed Central - PubMed

Affiliation: State key Laboratory of Wheat and Maize Crop Science/College of Plant Protection, Henan Agricultural University, Zhengzhou 450002 P.R. China.

ABSTRACT
Multicopper oxidases (MCOs) are enzymes that contain 10 conserved histidine residues and 1 cysteine residue. MCO1 has been extensively investigated in the midgut because this MCO is implicated in ascorbate oxidation, iron homeostasis and immune responses. However, information regarding the action of MCO1 in Malpighian tubules is limited. In this study, Helicoverpa armigera was used as a model to investigate the function of MCO1 in Malpighian tubules. Sequence analysis results revealed that HaMCO1 exhibits typical MCO characteristics, with 10 histidine and 1 cysteine residues for copper ion binding. HaMCO1 was also found to be highly abundant in Malpighian tubules. Temporal expression patterns indicated that HaMCO1 is mainly expressed during larval molting stages. Hormone treatments [the molting hormone 20-hydroxyecdysone (20E) and juvenile hormone (JH)] revealed that 20E inhibits HaMCO1 transcript expression via its heterodimer receptor, which consists of ecdysone receptor (EcR) and ultraspiracle (USP), and that JH counteracts the action of 20E to activate HaMCO1 transcript expression via its intracellular receptor methoprene-tolerant (Met). HaMCO1 knockdown caused a significant decrease in iron accumulation and also significantly reduced transferrin and ferritin transcript expression. Therefore, HaMCO1 is coordinately regulated by 20E and JH and is required for iron homeostasis in Malpighian tubules.

No MeSH data available.


Effect of HaMCO1 knockdown on iron accumulation and the expression of transferrin and ferritin transcripts.(A) qPCR analysis of HaMCO1 dsRNAi efficiency on the HaMCO1 transcript level. The 18S rRNA was used as the housekeeping gene for normalization in all qPCR analyses. The data represent the mean ± SD of three biological replicates. The significance of comparisons are marked with ***(p < 0.001), as determined by Student’s t-test. (B) The effect of HaMCO1 knockdown on iron accumulation. (B-1) Larval Malpighian tubules from the control (EGFP dsRNA) and treatment (HaMCO1 dsRNA) were analyzed. (B-2) The relative blue color density of the control (EGFP dsRNA) and treatment (HaMCO1 dsRNA). The relative blue color density was measured using Quantity One software. The data represent the mean ± SD of three biological replicates. Statistically significant differences were assessed by Student’s t-test (***p < 0.001). (C) The effect of HaMCO1 knockdown on the expression of transferrin and ferritin transcripts. qPCR analysis of the effects of HaMCO1 dsRNA injection on transferrin and ferritin transcript levels. The 18S rRNA was used as the housekeeping gene for normalization in all qPCR analyses. The data represent the mean ± SD of three biological replicates. The significance of comparisons are marked with ***(p < 0.001), as determined by Student’s t-test.
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f8: Effect of HaMCO1 knockdown on iron accumulation and the expression of transferrin and ferritin transcripts.(A) qPCR analysis of HaMCO1 dsRNAi efficiency on the HaMCO1 transcript level. The 18S rRNA was used as the housekeeping gene for normalization in all qPCR analyses. The data represent the mean ± SD of three biological replicates. The significance of comparisons are marked with ***(p < 0.001), as determined by Student’s t-test. (B) The effect of HaMCO1 knockdown on iron accumulation. (B-1) Larval Malpighian tubules from the control (EGFP dsRNA) and treatment (HaMCO1 dsRNA) were analyzed. (B-2) The relative blue color density of the control (EGFP dsRNA) and treatment (HaMCO1 dsRNA). The relative blue color density was measured using Quantity One software. The data represent the mean ± SD of three biological replicates. Statistically significant differences were assessed by Student’s t-test (***p < 0.001). (C) The effect of HaMCO1 knockdown on the expression of transferrin and ferritin transcripts. qPCR analysis of the effects of HaMCO1 dsRNA injection on transferrin and ferritin transcript levels. The 18S rRNA was used as the housekeeping gene for normalization in all qPCR analyses. The data represent the mean ± SD of three biological replicates. The significance of comparisons are marked with ***(p < 0.001), as determined by Student’s t-test.

Mentions: HaMCO1 is most abundantly expressed in Malpighian tubules, an important tissue that regulates the balance of water and ions. Accordingly, the effect of HaMCO1 knockdown on iron homeostasis was assessed. The results revealed that HaMCO1 knockdown caused a significant reduction in iron accumulation in Malpighian tubules compared with the EGFP controls (Fig. 8A,B).


Multicopper oxidase-1 is required for iron homeostasis in Malpighian tubules of Helicoverpa armigera.

Liu X, Sun C, Liu X, Yin X, Wang B, Du M, An S - Sci Rep (2015)

Effect of HaMCO1 knockdown on iron accumulation and the expression of transferrin and ferritin transcripts.(A) qPCR analysis of HaMCO1 dsRNAi efficiency on the HaMCO1 transcript level. The 18S rRNA was used as the housekeeping gene for normalization in all qPCR analyses. The data represent the mean ± SD of three biological replicates. The significance of comparisons are marked with ***(p < 0.001), as determined by Student’s t-test. (B) The effect of HaMCO1 knockdown on iron accumulation. (B-1) Larval Malpighian tubules from the control (EGFP dsRNA) and treatment (HaMCO1 dsRNA) were analyzed. (B-2) The relative blue color density of the control (EGFP dsRNA) and treatment (HaMCO1 dsRNA). The relative blue color density was measured using Quantity One software. The data represent the mean ± SD of three biological replicates. Statistically significant differences were assessed by Student’s t-test (***p < 0.001). (C) The effect of HaMCO1 knockdown on the expression of transferrin and ferritin transcripts. qPCR analysis of the effects of HaMCO1 dsRNA injection on transferrin and ferritin transcript levels. The 18S rRNA was used as the housekeeping gene for normalization in all qPCR analyses. The data represent the mean ± SD of three biological replicates. The significance of comparisons are marked with ***(p < 0.001), as determined by Student’s t-test.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4593997&req=5

f8: Effect of HaMCO1 knockdown on iron accumulation and the expression of transferrin and ferritin transcripts.(A) qPCR analysis of HaMCO1 dsRNAi efficiency on the HaMCO1 transcript level. The 18S rRNA was used as the housekeeping gene for normalization in all qPCR analyses. The data represent the mean ± SD of three biological replicates. The significance of comparisons are marked with ***(p < 0.001), as determined by Student’s t-test. (B) The effect of HaMCO1 knockdown on iron accumulation. (B-1) Larval Malpighian tubules from the control (EGFP dsRNA) and treatment (HaMCO1 dsRNA) were analyzed. (B-2) The relative blue color density of the control (EGFP dsRNA) and treatment (HaMCO1 dsRNA). The relative blue color density was measured using Quantity One software. The data represent the mean ± SD of three biological replicates. Statistically significant differences were assessed by Student’s t-test (***p < 0.001). (C) The effect of HaMCO1 knockdown on the expression of transferrin and ferritin transcripts. qPCR analysis of the effects of HaMCO1 dsRNA injection on transferrin and ferritin transcript levels. The 18S rRNA was used as the housekeeping gene for normalization in all qPCR analyses. The data represent the mean ± SD of three biological replicates. The significance of comparisons are marked with ***(p < 0.001), as determined by Student’s t-test.
Mentions: HaMCO1 is most abundantly expressed in Malpighian tubules, an important tissue that regulates the balance of water and ions. Accordingly, the effect of HaMCO1 knockdown on iron homeostasis was assessed. The results revealed that HaMCO1 knockdown caused a significant reduction in iron accumulation in Malpighian tubules compared with the EGFP controls (Fig. 8A,B).

Bottom Line: HaMCO1 was also found to be highly abundant in Malpighian tubules.HaMCO1 knockdown caused a significant decrease in iron accumulation and also significantly reduced transferrin and ferritin transcript expression.Therefore, HaMCO1 is coordinately regulated by 20E and JH and is required for iron homeostasis in Malpighian tubules.

View Article: PubMed Central - PubMed

Affiliation: State key Laboratory of Wheat and Maize Crop Science/College of Plant Protection, Henan Agricultural University, Zhengzhou 450002 P.R. China.

ABSTRACT
Multicopper oxidases (MCOs) are enzymes that contain 10 conserved histidine residues and 1 cysteine residue. MCO1 has been extensively investigated in the midgut because this MCO is implicated in ascorbate oxidation, iron homeostasis and immune responses. However, information regarding the action of MCO1 in Malpighian tubules is limited. In this study, Helicoverpa armigera was used as a model to investigate the function of MCO1 in Malpighian tubules. Sequence analysis results revealed that HaMCO1 exhibits typical MCO characteristics, with 10 histidine and 1 cysteine residues for copper ion binding. HaMCO1 was also found to be highly abundant in Malpighian tubules. Temporal expression patterns indicated that HaMCO1 is mainly expressed during larval molting stages. Hormone treatments [the molting hormone 20-hydroxyecdysone (20E) and juvenile hormone (JH)] revealed that 20E inhibits HaMCO1 transcript expression via its heterodimer receptor, which consists of ecdysone receptor (EcR) and ultraspiracle (USP), and that JH counteracts the action of 20E to activate HaMCO1 transcript expression via its intracellular receptor methoprene-tolerant (Met). HaMCO1 knockdown caused a significant decrease in iron accumulation and also significantly reduced transferrin and ferritin transcript expression. Therefore, HaMCO1 is coordinately regulated by 20E and JH and is required for iron homeostasis in Malpighian tubules.

No MeSH data available.