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Ecdysteroid-dependent expression of the tweedle and peroxidase genes during adult cuticle formation in the honey bee, Apis mellifera.

Soares MP, Silva-Torres FA, Elias-Neto M, Nunes FM, Simões ZL, Bitondi MM - PLoS ONE (2011)

Bottom Line: Gene sequencing and annotation validated the formerly predicted tweedle genes, and revealed a novel gene, Ampxd, in the honey bee genome.The effect of this hormone was confirmed in vivo by tying a ligature between the thorax and abdomen of early pupae to prevent the abdominal integument from coming in contact with ecdysteroids released from the prothoracic gland.Together, the data strongly suggest that these three genes play roles in ecdysteroid-dependent exoskeleton construction and differentiation and also point to a possible role for the two tweedle genes in the formation of the cuticle (peritrophic membrane) that internally lines the gut.

View Article: PubMed Central - PubMed

Affiliation: Departamento de Biologia, Faculdade de Filosofia, Ciências e Letras de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, São Paulo, Brazil.

ABSTRACT
Cuticle renewal is a complex biological process that depends on the cross talk between hormone levels and gene expression. This study characterized the expression of two genes encoding cuticle proteins sharing the four conserved amino acid blocks of the Tweedle family, AmelTwdl1 and AmelTwdl2, and a gene encoding a cuticle peroxidase containing the Animal haem peroxidase domain, Ampxd, in the honey bee. Gene sequencing and annotation validated the formerly predicted tweedle genes, and revealed a novel gene, Ampxd, in the honey bee genome. Expression of these genes was studied in the context of the ecdysteroid-coordinated pupal-to-adult molt, and in different tissues. Higher transcript levels were detected in the integument after the ecdysteroid peak that induces apolysis, coinciding with the synthesis and deposition of the adult exoskeleton and its early differentiation. The effect of this hormone was confirmed in vivo by tying a ligature between the thorax and abdomen of early pupae to prevent the abdominal integument from coming in contact with ecdysteroids released from the prothoracic gland. This procedure impaired the natural increase in transcript levels in the abdominal integument. Both tweedle genes were expressed at higher levels in the empty gut than in the thoracic integument and trachea of pharate adults. In contrast, Ampxd transcripts were found in higher levels in the thoracic integument and trachea than in the gut. Together, the data strongly suggest that these three genes play roles in ecdysteroid-dependent exoskeleton construction and differentiation and also point to a possible role for the two tweedle genes in the formation of the cuticle (peritrophic membrane) that internally lines the gut.

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Related in: MedlinePlus

Abundance of AmelTwdl1, AmelTwdl2 and Ampxd transcripts in the integument of ligated (L) abdomens compared to non-ligated controls (NL).Transcript levels were investigated at days (d) 1, 2, 3, 4, 5 and 6 after the abdominal ligature of newly ecdysed pupae. Transcript levels were assessed by semiquantitative RT-PCR assays followed by electrophoresis of the amplified cDNAs in ethidium bromide stained agarose gels. The Amactin gene was used as endogenous control.
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pone-0020513-g006: Abundance of AmelTwdl1, AmelTwdl2 and Ampxd transcripts in the integument of ligated (L) abdomens compared to non-ligated controls (NL).Transcript levels were investigated at days (d) 1, 2, 3, 4, 5 and 6 after the abdominal ligature of newly ecdysed pupae. Transcript levels were assessed by semiquantitative RT-PCR assays followed by electrophoresis of the amplified cDNAs in ethidium bromide stained agarose gels. The Amactin gene was used as endogenous control.

Mentions: The amounts of the AmelTwdl1, AmelTwdl2 and Ampxd transcripts did not increase in the integument of ligated abdomens as they did in intact abdomens, at least up to the fifth day after ligation (Figure 6). By impairing the flow of hemolymph, and consequently of ecdysteroids, to the abdomen, the ligature also temporarily impaired the increase in the abdominal levels of AmelTwdl1, AmelTwdl2 and Ampxd transcripts. This result suggests that abdominal exposure to the ecdysteroid peak that induces apolysis (see Figure 3) is critical for the increase in the levels of all of these transcripts in pharate adults. This result also reinforces the fact that Ampxd encodes a genuine cuticular peroxidase involved in cuticle sclerotization, which, like the CP genes studied here, depends on the ecdysteroid pulse for maximal activity.


Ecdysteroid-dependent expression of the tweedle and peroxidase genes during adult cuticle formation in the honey bee, Apis mellifera.

Soares MP, Silva-Torres FA, Elias-Neto M, Nunes FM, Simões ZL, Bitondi MM - PLoS ONE (2011)

Abundance of AmelTwdl1, AmelTwdl2 and Ampxd transcripts in the integument of ligated (L) abdomens compared to non-ligated controls (NL).Transcript levels were investigated at days (d) 1, 2, 3, 4, 5 and 6 after the abdominal ligature of newly ecdysed pupae. Transcript levels were assessed by semiquantitative RT-PCR assays followed by electrophoresis of the amplified cDNAs in ethidium bromide stained agarose gels. The Amactin gene was used as endogenous control.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3105072&req=5

pone-0020513-g006: Abundance of AmelTwdl1, AmelTwdl2 and Ampxd transcripts in the integument of ligated (L) abdomens compared to non-ligated controls (NL).Transcript levels were investigated at days (d) 1, 2, 3, 4, 5 and 6 after the abdominal ligature of newly ecdysed pupae. Transcript levels were assessed by semiquantitative RT-PCR assays followed by electrophoresis of the amplified cDNAs in ethidium bromide stained agarose gels. The Amactin gene was used as endogenous control.
Mentions: The amounts of the AmelTwdl1, AmelTwdl2 and Ampxd transcripts did not increase in the integument of ligated abdomens as they did in intact abdomens, at least up to the fifth day after ligation (Figure 6). By impairing the flow of hemolymph, and consequently of ecdysteroids, to the abdomen, the ligature also temporarily impaired the increase in the abdominal levels of AmelTwdl1, AmelTwdl2 and Ampxd transcripts. This result suggests that abdominal exposure to the ecdysteroid peak that induces apolysis (see Figure 3) is critical for the increase in the levels of all of these transcripts in pharate adults. This result also reinforces the fact that Ampxd encodes a genuine cuticular peroxidase involved in cuticle sclerotization, which, like the CP genes studied here, depends on the ecdysteroid pulse for maximal activity.

Bottom Line: Gene sequencing and annotation validated the formerly predicted tweedle genes, and revealed a novel gene, Ampxd, in the honey bee genome.The effect of this hormone was confirmed in vivo by tying a ligature between the thorax and abdomen of early pupae to prevent the abdominal integument from coming in contact with ecdysteroids released from the prothoracic gland.Together, the data strongly suggest that these three genes play roles in ecdysteroid-dependent exoskeleton construction and differentiation and also point to a possible role for the two tweedle genes in the formation of the cuticle (peritrophic membrane) that internally lines the gut.

View Article: PubMed Central - PubMed

Affiliation: Departamento de Biologia, Faculdade de Filosofia, Ciências e Letras de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, São Paulo, Brazil.

ABSTRACT
Cuticle renewal is a complex biological process that depends on the cross talk between hormone levels and gene expression. This study characterized the expression of two genes encoding cuticle proteins sharing the four conserved amino acid blocks of the Tweedle family, AmelTwdl1 and AmelTwdl2, and a gene encoding a cuticle peroxidase containing the Animal haem peroxidase domain, Ampxd, in the honey bee. Gene sequencing and annotation validated the formerly predicted tweedle genes, and revealed a novel gene, Ampxd, in the honey bee genome. Expression of these genes was studied in the context of the ecdysteroid-coordinated pupal-to-adult molt, and in different tissues. Higher transcript levels were detected in the integument after the ecdysteroid peak that induces apolysis, coinciding with the synthesis and deposition of the adult exoskeleton and its early differentiation. The effect of this hormone was confirmed in vivo by tying a ligature between the thorax and abdomen of early pupae to prevent the abdominal integument from coming in contact with ecdysteroids released from the prothoracic gland. This procedure impaired the natural increase in transcript levels in the abdominal integument. Both tweedle genes were expressed at higher levels in the empty gut than in the thoracic integument and trachea of pharate adults. In contrast, Ampxd transcripts were found in higher levels in the thoracic integument and trachea than in the gut. Together, the data strongly suggest that these three genes play roles in ecdysteroid-dependent exoskeleton construction and differentiation and also point to a possible role for the two tweedle genes in the formation of the cuticle (peritrophic membrane) that internally lines the gut.

Show MeSH
Related in: MedlinePlus