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Transpositionally active episomal hAT elements.

O'Brochta DA, Stosic CD, Pilitt K, Subramanian RA, Hice RH, Atkinson PW - BMC Mol. Biol. (2009)

Bottom Line: V(D)J signal joints, which resemble episomal transposable elements, have been considered non-recombinogenic products of V(D)J recombination and a safe way to dispose of excised chromosomal sequences.Up to 50% of hobo/Hermes episomes contained two intact, inverted-terminal repeats and 86% of these contained from 1-1000 bp of intercalary DNA.Episomal hobo/Hermes elements were recovered from Musca domestica (a natural host of Hermes), Drosophila melanogaster (a natural host of hobo) and transgenic Drosophila melanogaster and Aedes aegypti (with autonomous Hermes elements).

View Article: PubMed Central - HTML - PubMed

Affiliation: Center for Biosystems Research, University of Maryland Biotechnology Institute, Rockville, MD 20850, USA. obrochta@umbi.umd.edu

ABSTRACT

Background: hAT elements and V(D)J recombination may have evolved from a common ancestral transposable element system. Extrachromosomal, circular forms of transposable elements (referred to here as episomal forms) have been reported yet their biological significance remains unknown. V(D)J signal joints, which resemble episomal transposable elements, have been considered non-recombinogenic products of V(D)J recombination and a safe way to dispose of excised chromosomal sequences. V(D)J signal joints can, however, participate in recombination reactions and the purpose of this study was to determine if hobo and Hermes episomal elements are also recombinogenic.

Results: Up to 50% of hobo/Hermes episomes contained two intact, inverted-terminal repeats and 86% of these contained from 1-1000 bp of intercalary DNA. Episomal hobo/Hermes elements were recovered from Musca domestica (a natural host of Hermes), Drosophila melanogaster (a natural host of hobo) and transgenic Drosophila melanogaster and Aedes aegypti (with autonomous Hermes elements). Episomal Hermes elements were recovered from unfertilized eggs of M. domestica and D. melanogaster demonstrating their potential for extrachromosomal, maternal transmission. Reintegration of episomal Hermes elements was observed in vitro and in vivo and the presence of Hermes episomes resulted in lower rates of canonical Hermes transposition in vivo.

Conclusion: Episomal hobo/Hermes elements are common products of element excision and can be maternally transmitted. Episomal forms of Hermes are capable of integration and also of influencing the transposition of canonical elements suggesting biological roles for these extrachromosomal elements in element transmission and regulation.

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Map of episomal Hermes integration sites in the target plasmid pGDV1. The x axis refers to the linear form of the target plasmid and the position of the chloramphenicol resistance gene (CamR) is shown. The numbers on the x-axis refer to the length of the plasmid in basepairs. The y-axis is the number of integrations. Positive and negative values refer to elements that integrated into the target in opposite orientations. Five different classes of Hermes episomes were tested for their ability to integrate with each differing only in the amount of intercalary DNA (n) located between the inverted terminal repeats. The unweighted consensus sequence of the Hermes target site is graphically represented with a sequence logo [58] constructed with WebLogo [59].
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Figure 3: Map of episomal Hermes integration sites in the target plasmid pGDV1. The x axis refers to the linear form of the target plasmid and the position of the chloramphenicol resistance gene (CamR) is shown. The numbers on the x-axis refer to the length of the plasmid in basepairs. The y-axis is the number of integrations. Positive and negative values refer to elements that integrated into the target in opposite orientations. Five different classes of Hermes episomes were tested for their ability to integrate with each differing only in the amount of intercalary DNA (n) located between the inverted terminal repeats. The unweighted consensus sequence of the Hermes target site is graphically represented with a sequence logo [58] constructed with WebLogo [59].

Mentions: The recombination potential of a variety of Hermes episomes was tested directly using a cell free Hermes transposition assay with purified Hermes transposase, episomal Hermes elements and a target plasmid. Hermes episomes with n nucleotides of intercalary DNA between the inverted repeats were tested where n was 0, 1, 4, 5, 17, 37,69 and 120 nucleotides. All episomal forms of Hermes tested were capable of transposition resulting in canonical 8 bp target site duplications with the sequence nTnnnnAn (Table 6 and Figure 3). The distribution of integration events within the target indicated that certain sites were preferred as we have previously described (Figure 3) although the apparent hot spots of integration in vitro are not the same as reported in vivo (Table 6) [37,38]. Integrations at nucleotide 94 of the target plasmid were recovered frequently however it is not known whether the primary nucleotide sequence of this target site (ATTGAGAT) is the major determinant of this site's preferred status.


Transpositionally active episomal hAT elements.

O'Brochta DA, Stosic CD, Pilitt K, Subramanian RA, Hice RH, Atkinson PW - BMC Mol. Biol. (2009)

Map of episomal Hermes integration sites in the target plasmid pGDV1. The x axis refers to the linear form of the target plasmid and the position of the chloramphenicol resistance gene (CamR) is shown. The numbers on the x-axis refer to the length of the plasmid in basepairs. The y-axis is the number of integrations. Positive and negative values refer to elements that integrated into the target in opposite orientations. Five different classes of Hermes episomes were tested for their ability to integrate with each differing only in the amount of intercalary DNA (n) located between the inverted terminal repeats. The unweighted consensus sequence of the Hermes target site is graphically represented with a sequence logo [58] constructed with WebLogo [59].
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2803484&req=5

Figure 3: Map of episomal Hermes integration sites in the target plasmid pGDV1. The x axis refers to the linear form of the target plasmid and the position of the chloramphenicol resistance gene (CamR) is shown. The numbers on the x-axis refer to the length of the plasmid in basepairs. The y-axis is the number of integrations. Positive and negative values refer to elements that integrated into the target in opposite orientations. Five different classes of Hermes episomes were tested for their ability to integrate with each differing only in the amount of intercalary DNA (n) located between the inverted terminal repeats. The unweighted consensus sequence of the Hermes target site is graphically represented with a sequence logo [58] constructed with WebLogo [59].
Mentions: The recombination potential of a variety of Hermes episomes was tested directly using a cell free Hermes transposition assay with purified Hermes transposase, episomal Hermes elements and a target plasmid. Hermes episomes with n nucleotides of intercalary DNA between the inverted repeats were tested where n was 0, 1, 4, 5, 17, 37,69 and 120 nucleotides. All episomal forms of Hermes tested were capable of transposition resulting in canonical 8 bp target site duplications with the sequence nTnnnnAn (Table 6 and Figure 3). The distribution of integration events within the target indicated that certain sites were preferred as we have previously described (Figure 3) although the apparent hot spots of integration in vitro are not the same as reported in vivo (Table 6) [37,38]. Integrations at nucleotide 94 of the target plasmid were recovered frequently however it is not known whether the primary nucleotide sequence of this target site (ATTGAGAT) is the major determinant of this site's preferred status.

Bottom Line: V(D)J signal joints, which resemble episomal transposable elements, have been considered non-recombinogenic products of V(D)J recombination and a safe way to dispose of excised chromosomal sequences.Up to 50% of hobo/Hermes episomes contained two intact, inverted-terminal repeats and 86% of these contained from 1-1000 bp of intercalary DNA.Episomal hobo/Hermes elements were recovered from Musca domestica (a natural host of Hermes), Drosophila melanogaster (a natural host of hobo) and transgenic Drosophila melanogaster and Aedes aegypti (with autonomous Hermes elements).

View Article: PubMed Central - HTML - PubMed

Affiliation: Center for Biosystems Research, University of Maryland Biotechnology Institute, Rockville, MD 20850, USA. obrochta@umbi.umd.edu

ABSTRACT

Background: hAT elements and V(D)J recombination may have evolved from a common ancestral transposable element system. Extrachromosomal, circular forms of transposable elements (referred to here as episomal forms) have been reported yet their biological significance remains unknown. V(D)J signal joints, which resemble episomal transposable elements, have been considered non-recombinogenic products of V(D)J recombination and a safe way to dispose of excised chromosomal sequences. V(D)J signal joints can, however, participate in recombination reactions and the purpose of this study was to determine if hobo and Hermes episomal elements are also recombinogenic.

Results: Up to 50% of hobo/Hermes episomes contained two intact, inverted-terminal repeats and 86% of these contained from 1-1000 bp of intercalary DNA. Episomal hobo/Hermes elements were recovered from Musca domestica (a natural host of Hermes), Drosophila melanogaster (a natural host of hobo) and transgenic Drosophila melanogaster and Aedes aegypti (with autonomous Hermes elements). Episomal Hermes elements were recovered from unfertilized eggs of M. domestica and D. melanogaster demonstrating their potential for extrachromosomal, maternal transmission. Reintegration of episomal Hermes elements was observed in vitro and in vivo and the presence of Hermes episomes resulted in lower rates of canonical Hermes transposition in vivo.

Conclusion: Episomal hobo/Hermes elements are common products of element excision and can be maternally transmitted. Episomal forms of Hermes are capable of integration and also of influencing the transposition of canonical elements suggesting biological roles for these extrachromosomal elements in element transmission and regulation.

Show MeSH
Related in: MedlinePlus