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The proteins encoded by the pogo-like Lemi1 element bind the TIRs and subterminal repeated motifs of the Arabidopsis Emigrant MITE: consequences for the transposition mechanism of MITEs.

Loot C, Santiago N, Sanz A, Casacuberta JM - Nucleic Acids Res. (2006)

Bottom Line: We present here evidence for a recent mobility of the Arabidopsis Emigrant MITE and we report on the capacity of the proteins encoded by the related Lemi1 transposon, a pogo-related element, to specifically bind Emigrant elements.Our results show that Lemi1 proteins bind Emigrant TIRs but also bind cooperatively to subterminal repeated motifs.The requirement of internal sequences for the formation of proper DNA/protein structure could affect the capacity of divergent MITEs to be mobilized by distantly related transposases.

View Article: PubMed Central - PubMed

Affiliation: Departament de Genètica Molecular, Laboratori de Genètica Molecular Vegetal, CSIC-IRTA, Jordi Girona 18, 08034 Barcelona, Spain.

ABSTRACT
MITEs (miniature inverted-repeated transposable elements) are a particular class of defective DNA transposons usually present within genomes as high copy number populations of highly homogeneous elements. Although an active MITE, the mPing element, has recently been characterized in rice, the transposition mechanism of MITEs remains unknown. It has been proposed that transposases of related transposons could mobilize MITEs in trans. Moreover, it has also been proposed that the presence of conserved terminal inverted-repeated (TIR) sequences could be the only requirement of MITEs for mobilization, allowing divergent or unrelated elements to be mobilized by a particular transposase. We present here evidence for a recent mobility of the Arabidopsis Emigrant MITE and we report on the capacity of the proteins encoded by the related Lemi1 transposon, a pogo-related element, to specifically bind Emigrant elements. This suggests that Lemi1 could mobilize Emigrant elements and makes the Lemi1/Emigrant couple an ideal system to study the transposition mechanism of MITEs. Our results show that Lemi1 proteins bind Emigrant TIRs but also bind cooperatively to subterminal repeated motifs. The requirement of internal sequences for the formation of proper DNA/protein structure could affect the capacity of divergent MITEs to be mobilized by distantly related transposases.

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Structure and sequences of the Lemi1 elements from different Arabidopsis ecotypes. (A) The structure of the Columbia Lemi1 element is shown compared with that of a consensus Emigrant element. The percentage of identity between both sequences in the common regions is shown on the top of the Emigrant scheme. The two orfs of Lemi1, as well as the position of the stop codon interrupting the orf, and of the putative intron are shown. The position of the two AccI (A) used for cloning (see Materials and Methods) is shown. (B) Comparison of the DNA sequences of three polymorphic regions (the region containing the stop codon in Columbia, STOP; the region containing an insertion in Co-1 and Co-4 ecotypes, INSERTION; and the region of the putative acceptor splicing site, AS) of Lemi1 from seven different Arabidopsis ecotypes: Moscow (Ms-0), RLD, Dijon-G, Columbia (Col-0), Coimbra-4 (Co-4), Coimbra-1 (Co-1) and two alleles of Tsu-0. The accession numbers of the sequences are DQ888704, DQ888705, DQ888706, At2g06660, DQ888708, DQ888709, DQ888710 and DQ888711, respectively. The polypeptide encoded by the insertion sequence is shown on the bottom of the corresponding sequence. The plant consensus sequence of the splicing acceptor site sequence (numbers below sequences represent the percentage occurrence of indicated bases) is shown underneath the corresponding sequences.
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fig2: Structure and sequences of the Lemi1 elements from different Arabidopsis ecotypes. (A) The structure of the Columbia Lemi1 element is shown compared with that of a consensus Emigrant element. The percentage of identity between both sequences in the common regions is shown on the top of the Emigrant scheme. The two orfs of Lemi1, as well as the position of the stop codon interrupting the orf, and of the putative intron are shown. The position of the two AccI (A) used for cloning (see Materials and Methods) is shown. (B) Comparison of the DNA sequences of three polymorphic regions (the region containing the stop codon in Columbia, STOP; the region containing an insertion in Co-1 and Co-4 ecotypes, INSERTION; and the region of the putative acceptor splicing site, AS) of Lemi1 from seven different Arabidopsis ecotypes: Moscow (Ms-0), RLD, Dijon-G, Columbia (Col-0), Coimbra-4 (Co-4), Coimbra-1 (Co-1) and two alleles of Tsu-0. The accession numbers of the sequences are DQ888704, DQ888705, DQ888706, At2g06660, DQ888708, DQ888709, DQ888710 and DQ888711, respectively. The polypeptide encoded by the insertion sequence is shown on the bottom of the corresponding sequence. The plant consensus sequence of the splicing acceptor site sequence (numbers below sequences represent the percentage occurrence of indicated bases) is shown underneath the corresponding sequences.

Mentions: It has been proposed that Emigrant elements originated by a severe deletion of a putative pogo-like transposon called Lemi1 that shows extensive sequence homology with Emigrant elements (12). Lemi1 is present only in one copy of the Arabidopsis Columbia ecotype, and displays an orf potentially coding for a pogo-like transposase interrupted by a STOP codon in position 39 of the protein and a frameshift in position 385 of the protein [(12) and Figure 2A]. It has been proposed that the splicing of a putative intron could allow overcoming the frameshift (12). The sequence of the putative donor and acceptor splicing sites (data not shown and Figure 2B, respectively) perfectly fit the consensus for plant introns (14) and a consensus branch point sequence is found at the correct distance from the acceptor AG (data not shown). In order to get insight on the original structure of the Lemi1 element we have analysed the Lemi1 sequences of different Arabidopsis ecotypes. Lemi1 is present as a single-copy element in all the Arabidopsis ecotypes that we have analysed, and we have not been able to obtain evidences of mobility of Lemi1 in those genomes (data not shown). Using primers complementary to internal Lemi1 sequences we have amplified, sequenced and compared Lemi1 sequences obtained from seven different Arabidopsis ecotypes. In spite of the high similarity found (from 94 to 100% identical over 1245 bp), the Lemi1 sequences are polymorphic at particularly important positions (Figure 2B). The Lemi1 sequence does not contain the STOP codon found in Columbia in four different ecotypes, Ms-0, RLD and Dijon-G, and is probably present in two different alleles (only one of them containing the STOP) in the Tsu-0 ecotype, as we have obtained two types of sequences when amplifying this region from this ecotype. On the other hand, the Lemi1 sequence has an insertion of 56 bp in the region of the frameshift that restores the coding capacity in a single orf in Coimbra-1 and Coimbra-4 (Figure 2B). The presence of this insertion in Lemi1-related sequences of Gossypium hirsutum, Solanum demisum and Medicago truncatula (data not shown) suggests that the original Lemi1 element had a single orf of 1559 bp. Nevertheless, Coimbra-1 and Coimbra-4 also have a difference with respect to the Lemi1 sequences found in all other ecotypes at one of the two invariable nucleotides of the putative splicing acceptor site (Figure 2B). The presence of an acceptor splicing consensus site exclusively in the Lemi1 sequences where the orf is interrupted by a frameshift (Figure 2B) could also indicate the functionality of a spliced protein.


The proteins encoded by the pogo-like Lemi1 element bind the TIRs and subterminal repeated motifs of the Arabidopsis Emigrant MITE: consequences for the transposition mechanism of MITEs.

Loot C, Santiago N, Sanz A, Casacuberta JM - Nucleic Acids Res. (2006)

Structure and sequences of the Lemi1 elements from different Arabidopsis ecotypes. (A) The structure of the Columbia Lemi1 element is shown compared with that of a consensus Emigrant element. The percentage of identity between both sequences in the common regions is shown on the top of the Emigrant scheme. The two orfs of Lemi1, as well as the position of the stop codon interrupting the orf, and of the putative intron are shown. The position of the two AccI (A) used for cloning (see Materials and Methods) is shown. (B) Comparison of the DNA sequences of three polymorphic regions (the region containing the stop codon in Columbia, STOP; the region containing an insertion in Co-1 and Co-4 ecotypes, INSERTION; and the region of the putative acceptor splicing site, AS) of Lemi1 from seven different Arabidopsis ecotypes: Moscow (Ms-0), RLD, Dijon-G, Columbia (Col-0), Coimbra-4 (Co-4), Coimbra-1 (Co-1) and two alleles of Tsu-0. The accession numbers of the sequences are DQ888704, DQ888705, DQ888706, At2g06660, DQ888708, DQ888709, DQ888710 and DQ888711, respectively. The polypeptide encoded by the insertion sequence is shown on the bottom of the corresponding sequence. The plant consensus sequence of the splicing acceptor site sequence (numbers below sequences represent the percentage occurrence of indicated bases) is shown underneath the corresponding sequences.
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Related In: Results  -  Collection

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fig2: Structure and sequences of the Lemi1 elements from different Arabidopsis ecotypes. (A) The structure of the Columbia Lemi1 element is shown compared with that of a consensus Emigrant element. The percentage of identity between both sequences in the common regions is shown on the top of the Emigrant scheme. The two orfs of Lemi1, as well as the position of the stop codon interrupting the orf, and of the putative intron are shown. The position of the two AccI (A) used for cloning (see Materials and Methods) is shown. (B) Comparison of the DNA sequences of three polymorphic regions (the region containing the stop codon in Columbia, STOP; the region containing an insertion in Co-1 and Co-4 ecotypes, INSERTION; and the region of the putative acceptor splicing site, AS) of Lemi1 from seven different Arabidopsis ecotypes: Moscow (Ms-0), RLD, Dijon-G, Columbia (Col-0), Coimbra-4 (Co-4), Coimbra-1 (Co-1) and two alleles of Tsu-0. The accession numbers of the sequences are DQ888704, DQ888705, DQ888706, At2g06660, DQ888708, DQ888709, DQ888710 and DQ888711, respectively. The polypeptide encoded by the insertion sequence is shown on the bottom of the corresponding sequence. The plant consensus sequence of the splicing acceptor site sequence (numbers below sequences represent the percentage occurrence of indicated bases) is shown underneath the corresponding sequences.
Mentions: It has been proposed that Emigrant elements originated by a severe deletion of a putative pogo-like transposon called Lemi1 that shows extensive sequence homology with Emigrant elements (12). Lemi1 is present only in one copy of the Arabidopsis Columbia ecotype, and displays an orf potentially coding for a pogo-like transposase interrupted by a STOP codon in position 39 of the protein and a frameshift in position 385 of the protein [(12) and Figure 2A]. It has been proposed that the splicing of a putative intron could allow overcoming the frameshift (12). The sequence of the putative donor and acceptor splicing sites (data not shown and Figure 2B, respectively) perfectly fit the consensus for plant introns (14) and a consensus branch point sequence is found at the correct distance from the acceptor AG (data not shown). In order to get insight on the original structure of the Lemi1 element we have analysed the Lemi1 sequences of different Arabidopsis ecotypes. Lemi1 is present as a single-copy element in all the Arabidopsis ecotypes that we have analysed, and we have not been able to obtain evidences of mobility of Lemi1 in those genomes (data not shown). Using primers complementary to internal Lemi1 sequences we have amplified, sequenced and compared Lemi1 sequences obtained from seven different Arabidopsis ecotypes. In spite of the high similarity found (from 94 to 100% identical over 1245 bp), the Lemi1 sequences are polymorphic at particularly important positions (Figure 2B). The Lemi1 sequence does not contain the STOP codon found in Columbia in four different ecotypes, Ms-0, RLD and Dijon-G, and is probably present in two different alleles (only one of them containing the STOP) in the Tsu-0 ecotype, as we have obtained two types of sequences when amplifying this region from this ecotype. On the other hand, the Lemi1 sequence has an insertion of 56 bp in the region of the frameshift that restores the coding capacity in a single orf in Coimbra-1 and Coimbra-4 (Figure 2B). The presence of this insertion in Lemi1-related sequences of Gossypium hirsutum, Solanum demisum and Medicago truncatula (data not shown) suggests that the original Lemi1 element had a single orf of 1559 bp. Nevertheless, Coimbra-1 and Coimbra-4 also have a difference with respect to the Lemi1 sequences found in all other ecotypes at one of the two invariable nucleotides of the putative splicing acceptor site (Figure 2B). The presence of an acceptor splicing consensus site exclusively in the Lemi1 sequences where the orf is interrupted by a frameshift (Figure 2B) could also indicate the functionality of a spliced protein.

Bottom Line: We present here evidence for a recent mobility of the Arabidopsis Emigrant MITE and we report on the capacity of the proteins encoded by the related Lemi1 transposon, a pogo-related element, to specifically bind Emigrant elements.Our results show that Lemi1 proteins bind Emigrant TIRs but also bind cooperatively to subterminal repeated motifs.The requirement of internal sequences for the formation of proper DNA/protein structure could affect the capacity of divergent MITEs to be mobilized by distantly related transposases.

View Article: PubMed Central - PubMed

Affiliation: Departament de Genètica Molecular, Laboratori de Genètica Molecular Vegetal, CSIC-IRTA, Jordi Girona 18, 08034 Barcelona, Spain.

ABSTRACT
MITEs (miniature inverted-repeated transposable elements) are a particular class of defective DNA transposons usually present within genomes as high copy number populations of highly homogeneous elements. Although an active MITE, the mPing element, has recently been characterized in rice, the transposition mechanism of MITEs remains unknown. It has been proposed that transposases of related transposons could mobilize MITEs in trans. Moreover, it has also been proposed that the presence of conserved terminal inverted-repeated (TIR) sequences could be the only requirement of MITEs for mobilization, allowing divergent or unrelated elements to be mobilized by a particular transposase. We present here evidence for a recent mobility of the Arabidopsis Emigrant MITE and we report on the capacity of the proteins encoded by the related Lemi1 transposon, a pogo-related element, to specifically bind Emigrant elements. This suggests that Lemi1 could mobilize Emigrant elements and makes the Lemi1/Emigrant couple an ideal system to study the transposition mechanism of MITEs. Our results show that Lemi1 proteins bind Emigrant TIRs but also bind cooperatively to subterminal repeated motifs. The requirement of internal sequences for the formation of proper DNA/protein structure could affect the capacity of divergent MITEs to be mobilized by distantly related transposases.

Show MeSH
Related in: MedlinePlus