Escherichia coli RecX inhibits RecA recombinase and coprotease activities in vitro and in vivo.
In Escherichia coli the RecA protein plays a pivotal role in homologous recombination, DNA repair, and SOS repair and mutagenesis. A gene designated recX (or oraA) is present directly downstream of recA in E. coli; however, the function of RecX is unknown. In this work we demonstrated interaction of RecX and RecA in a yeast two-hybrid assay. In vitro, substoichiometric amounts of RecX strongly inhibited both RecA-mediated DNA strand exchange and RecA ATPase activity. In vivo, we showed that recX is under control of the LexA repressor and is up-regulated in response to DNA damage. A loss-of-function mutation in recX resulted in decreased resistance to UV irradiation; however, overexpression of RecX in trans resulted in a greater decrease in UV resistance. Overexpression of RecX inhibited induction of two din (damage-inducible) genes and cleavage of the UmuD and LexA repressor proteins; however, recX inactivation had no effect on any of these processes. Cells overexpressing RecX showed decreased levels of P1 transduction, whereas recX mutation had no effect on P1 transduction frequency. Our combined in vitro and in vivo data indicate that RecX can inhibit both RecA recombinase and coprotease activities.
Stohl
Elizabeth A
EA
Department of Microbiology and Immunology, Feinberg School of Medicine, Northwestern University, Chicago, Illinois 60611, USA.
Brockman
Joel P
JP
Burkle
Kristin L
KL
Morimatsu
Katsumi
K
Kowalczykowski
Stephen C
SC
Seifert
H Steven
HS
D000255:Adenosine Triphosphate / Q000378:metabolism; D001426:Bacterial Proteins / Q000378:metabolism*; D015153:Blotting, Western; D004247:DNA / Q000378:metabolism; D004249:DNA Damage; D004259:DNA-Directed DNA Polymerase; D004307:Dose-Response Relationship, Radiation; D004926:Escherichia coli / Q000201:enzymology*; D029968:Escherichia coli Proteins / Q000378:metabolism; D006868:Hydrolysis; D066298:In Vitro Techniques; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D011938:Rec A Recombinases / Q000378:metabolism*; D011995:Recombination, Genetic; D012697:Serine Endopeptidases / Q000378:metabolism; D013997:Time Factors; D020798:Two-Hybrid System Techniques; D014466:Ultraviolet Rays; D015854:Up-Regulation; D000099160:LexA Repressor Protein
D000255:Adenosine Triphosphate; D001426:Bacterial Proteins; D004247:DNA; D004259:DNA-Directed DNA Polymerase; D029968:Escherichia coli Proteins; D011938:Rec A Recombinases; D012697:Serine Endopeptidases; D000099160:LexA Repressor Protein; C430523:RecX protein, Xanthomonas campestris; C055218:UmuD protein, E coli