Tn10 insertion specificity is strongly dependent upon sequences immediately adjacent to the target-site consensus sequence.
AUTOR(ES)
Bender, J
RESUMO
Transposon Tn10 inserts preferentially into particular "hotspots" that have been shown by sequence analysis to contain the symmetrical consensus sequence 5'-GCTNAGC-3'. This consensus is necessary but not sufficient to determine insertion specificity. We have mutagenized a known hotspot to identify other determinants for insertion into this site. This genetic dissection of the sequence context of a protein binding site shows that a second major determinant for Tn10 insertion specificity is contributed by the 6-9 base pairs that flank each end of the consensus sequence. Variations in these context base pairs can confer variations of at least 1000-fold in insertion frequency. There is no discernible consensus sequence for the context determinant, suggesting that sequence-specific protein-DNA contacts are not playing a major role. Taken together with previous work, the observations presented suggest a model for the interaction of transposase with the insertion site: symmetrically disposed subunits bind with specific contacts to the major groove of consensus-sequence base pairs, while flanking sequences influence the interaction through effects on DNA helix structure. We also show that the determinants important for insertion into a site are not important for transposition out of that site.
ACESSO AO ARTIGO
http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=49842Documentos Relacionados
- Specificity of Insertion by the Translocatable Tetracycline-Resistance Element Tn10
- Insecticide resistance resulting from an absence of target-site gene product
- Transposon Tn10 provides a promoter for transcription of adjacent sequences.
- Rules for DNA target-site recognition by a lactococcal group II intron enable retargeting of the intron to specific DNA sequences
- Efficient Tn10 transposition into a DNA insertion hot spot in vivo requires the 5-methyl groups of symmetrically disposed thymines within the hot-spot consensus sequence.