Asunto(s)
Endorribonucleasas/genética , Escherichia coli/genética , Complejos Multienzimáticos/genética , Polirribonucleótido Nucleotidiltransferasa/genética , ARN Helicasas/genética , ARN Bacteriano/genética , Cromatografía en Gel , Electroforesis en Gel de Poliacrilamida , Endorribonucleasas/aislamiento & purificación , Hidrólisis , Complejos Multienzimáticos/aislamiento & purificación , Polirribonucleótido Nucleotidiltransferasa/aislamiento & purificación , ARN Helicasas/aislamiento & purificaciónRESUMEN
IS911 transposition involves a free circular transposon intermediate where the terminal inverted repeat sequences are connected. Transposase synthesis is usually driven by a weak promoter, p(IRL), in the left end (IRL). Circle junction formation creates a strong promoter, p(junc), with a -35 sequence located in the right end and the -10 sequence in the left. p(junc) assembly would permit an increase in synthesis of transposase from the transposon circle, which would be expected to stimulate integration. Insertion results in p(junc) disassembly and a return to the low p(IRL)- driven transposase levels. We demonstrate that p(junc) plays an important role in regulating IS911 transposition. Inactivation of p(junc) strongly decreased IS911 transposition when transposase was produced in its natural configuration. This novel feedback mechanism permits transient and controlled activation of integration only in the presence of the correct (circular) intermediate. We have also investigated other members of the IS3 and other IS families. Several, but not all, IS3 family members possess p(junc) equivalents, underlining that the regulatory mechanisms adopted to fine-tune transposition may be different.