MONTI MARIELA ROXANA
Congresos y reuniones científicas
Título:
INFLUENCE OF DNA HETEROLOGIES ON THE STRAND EXCHANGE REACTION CATALYZED BY RecA OF Pseudomonas aeruginosa
Autor/es:
BORGOGNO MARÍA V.; MONTI MARIELA R.; ARGARAÑA CARLOS E.
Reunión:
Congreso; XI Reunión Anual de SAMIGE; 2015
Resumen:
Homologousrecombination (HR) plays key roles in the generation of genetic diversity byreassembling DNA sequences from total or partially homologous DNA molecules. Itallows rapid acquisition of novel functions, driving adaptation and promoting speciation.However, creation of new sequence combinations by HR has to be balanced to givethe highest chance of functionality maintenance of coded proteins. In bacteria,the initial stages of recombination are catalyzed by RecA, which promotes thealignment of two DNA sequences and initiates strand exchange between them. Animportant property of RecA is its tolerance to a limited extent of heterologywhich determines the efficiency of strand exchange between divergent DNA sequencesand the fidelity of HR. Besides RecA, the ability to tolerate heterologiesduring recombination is held in check by the Mismatch Repair System (MRS).Thus, when MRS is either downregulated or inactivated the heterologiesdiscrimination by RecA may be decisive to maintain the integrity of the genome andultimately to avoid mutations. In the present work, we study the influence ofheterologies on HR catalyzed by RecA of Pseudomonas aeruginosa. By a systematicin vitro study of oligonucleotides strand exchange using fluorescence resonanceenergy transfer (FRET) techniques, we examined the efficiency of RecA-catalyzedstrand exchange in presence of 12 different types of single mismatches in threepositions (close to the 5?, middle or 3? end) of the incoming strand, as wellas the presence of small loops. Here we show that most mismatches close to the5? end of the incoming strand had strong inhibitory effect and this wasvariable depending on the type of mismatch. On the other hand, most mismatcheslocated in the middle or 3? end of the incoming strand did not show aninhibitory effect and the strand exchange efficiency was similar amongdifferent types of mismatches. Furthermore, a direct correlation betweenstrength of inhibition and the reversibility of strand exchange reaction wasobserved. Therefore, RecA recognizes different types of mismatches depending onits position suggesting that differences in the intrinsic properties of DNAowing to mismatch type, neighboring nucleotide sequences and the position ofthe mismatch may play a role during the strand exchange. In addition, we foundthat the presence of small loops (+/- 1 or 2 nucleotides) diminishedRecA-mediated strand exchange reaction. These results may suggest that RecAmaintains a guarding role against frameshift mutations, such as it waspreviously described for the MRS proteins. In conclusion, RecA can effectivelycompare DNA segments with different types of heterologies in its search forhomology.
