The enzymatic component of Drosophila melanogaster chorion is the Pxd peroxidase

Konstandi, Ourania; Papassideri, Issidora S.; Stravopodis, Dimitrios J.; Kenoutis, Christos A.; Hasan, Zulfiqar; Katsorchis, Theodoros; Wever, Ron; Margaritis, Lukas H.

doi:10.1016/j.ibmb.2005.04.005

Cited by 31 publications

(36 citation statements)

References 34 publications

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“…Similarly to what has been previously reported for D. melanogaster (Konstandi et al, 2005), a single copy homologous gene (bPxd) is presented to occur in the B. oleae genome, according to the Southern blotting analysis of genomic DNA (Sambrook et al, 1989) purified from adult flies, subsequently digested with different restriction endonucleases (EcoRI, BamHI, PstI and HindIII) and finally, after band-separation and blotting, hybridized against the 0.5 kb RT-PCR fragment directly generated from total RNA extracts of dissected ovaries, containing choriogenic-specific follicles (Fig. 1A).…”

Section: The Gene Amplification Rule Does Not Apply For the Single Cosupporting

confidence: 89%

“…In the present study, we demonstrate that the B. oleae chorion peroxidase, bPxd, a highly homologous member with the Drosophila Pxd peroxidase (Konstandi et al, 2005), plays an essential role and participates with a dual fashion in the chorion assembly and formation process. The totality of our conclusions derived from (a) Southern blotting, (b) Northern blotting, (c) semi-quantitative RT-PCR reactions, (d) in situ hybridization analysis, (e) Western blotting and (f) immunolocalization approaches (Light Microscopy immunodetection and Transmission Electron Microscopy immunogold labelling), compared with previously reported data concerning the Drosophila Pxd peroxidase (Konstandi et al, 2005), strongly suggest for the presence of a highly conserved mechanism of chorion hardening process among all Diptera.…”

Section: Introductionmentioning

confidence: 60%

“…The totality of our conclusions derived from (a) Southern blotting, (b) Northern blotting, (c) semi-quantitative RT-PCR reactions, (d) in situ hybridization analysis, (e) Western blotting and (f) immunolocalization approaches (Light Microscopy immunodetection and Transmission Electron Microscopy immunogold labelling), compared with previously reported data concerning the Drosophila Pxd peroxidase (Konstandi et al, 2005), strongly suggest for the presence of a highly conserved mechanism of chorion hardening process among all Diptera. Thus, by exploiting the diverse and especially the genetically developed, technologies for pest control, the B. oleae chorion peroxidase, bPxd, could comprise a crucial protein to be directly targeted in vivo, in order to develop highly successful protocols for a large variety of agricultural applications.…”

Section: Introductionmentioning

confidence: 70%

“…Two specific primers were designed, the forward pQUE and the reverse pQUErev, according to sequences corresponding to conserved regions among insect and mammalian peroxidase genes (Konstandi et al, 2005). A 0.5 kb cDNA fragment was amplified through a reverse transcription-PCR (RT-PCR) approach, using as a template the total RNA preparation purified from dissected B. oleae ovaries and subsequently cloned into the pGEM-T-easy vector, according to manufacturer's instructions.…”

Section: Strong Phylogenetic Conservation Of Bpxd Cdnamentioning

confidence: 99%

“…The upper part of (A,B) shows a developed autoradiograph, while the bottom part illustrates the ethidium bromide-stained agarose gel before transfer. The 0.5 kb bPxd cDNA fragment was purified and used for high stringency Southern blotting of B. oleae genomic DNA (gDNA) isolated from adult flies (females without ovaries and males) and subsequently digested with the EcoRI, BamHI, PstI and HindIII restriction endonucleases (single or double incuba-Comparative analysis of the DNA sequencing data revealed an identity of 98% with the Drosophila Pxd cDNA clone, previously isolated by Konstandi et al (2005), exhibiting only two "silent" nucleotide differences (without any impact on the amino acid sequence of the protein) and also showing significant homologies with the mammalian myeloperoxidase (MPO), thyroid peroxidase (TPO) and eosinophil peroxidase (EOS) (data not shown). The cloned 0.5 kb bPxd cDNA fragment likely corresponds to the catalytic domain (CD) of the B. oleae putative peroxidase and thus was selected as the most suitable probe for all hybridization experiments.…”

Section: A B Tionsmentioning

confidence: 99%

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The dual role of chorion peroxidase in Bactrocera oleae chorion assembly

Konstandi¹,

Papassideri²,

Stravopodis³

et al. 2006

Int. J. Dev. Biol.

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In the present study, we reveal for the first time that the Bactrocera oleae chorion peroxidase (bPxd) participates essentially in B. oleae chorion formation and clearly represents the homologous member of Drosophila melanogaster chorion peroxidase (Pxd). Comparative sequence analysis disclosed that the bPxd cDNA semi-central region, which encodes for the putative catalytic domain of the enzyme, exhibits great homology (98%) with its Pxd counterpart. Thus, it is very likely that bPxd is highly responsible for the chorion hardening process, through protein cross-linking mediated by the formation of di-and tri-tyrosine bonds. Distinct molecular weight bPxd RNA transcripts were detected in Northern blotting analysis of total RNA extracts of adult flies (2.9 and 1.7 kb) and ovaries (2.2 kb). The ovarian-specific bPxd RNA transcript is selectively expressed in the follicle cell layer during the late stages of oogenesis 12-14, as revealed by in situ hybridization. Moreover, reverse transcription reactions confirmed the stage-specific developmental regulation of the bPxd gene, which is maximally expressed during stage 13. Western blotting with the rabbit anti-rAePO polyclonal antibody revealed three immunoreactive bands of 76, 66 and 54 kDa in crude protein extracts from adult flies, while in larva and purified chorion preparations, a unique 54 kDa band was clearly detected. Immunolocalization experiments revealed that bPxd peroxidase constitutes an essential structural chorionic component, being abundantly localized in all the successive chorionic layers and vitelline membrane as well.

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Section: The Gene Amplification Rule Does Not Apply For the Single Cosupporting

confidence: 89%

Section: Introductionmentioning

confidence: 60%

Section: Introductionmentioning

confidence: 70%

Section: Strong Phylogenetic Conservation Of Bpxd Cdnamentioning

confidence: 99%

Section: A B Tionsmentioning

confidence: 99%

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