Central Role of the Holliday Junction Helicase RuvAB in vlsE Recombination and Infectivity of Borrelia burgdorferi

Abstract

Antigenic variation plays a vital role in the pathogenesis of many infectious bacteria and protozoa including Borrelia burgdorferi, the causative agent of Lyme disease. VlsE, a 35 kDa surface-exposed lipoprotein, undergoes antigenic variation during B. burgdorferi infection of mammalian hosts, and is believed to be a critical mechanism by which the spirochetes evade immune clearance. Random, segmental recombination between the expressed vlsE gene and adjacent vls silent cassettes generates a large number of different VlsE variants within the infected host. Although the occurrence and importance of vlsE sequence variation is well established, little is known about the biological mechanism of vlsE recombination. To identify factors important in antigenic variation and vlsE recombination, we screened transposon mutants of genes known to be involved in DNA recombination and repair for their effects on infectivity and vlsE recombination. Several mutants, including those in BB0023 (ruvA), BB0022 (ruvB), BB0797 (mutS), and BB0098 (mutS-II), showed reduced infectivity in immunocompetent C3H/HeN mice. Mutants in ruvA and ruvB exhibited greatly reduced rates of vlsE recombination in C3H/HeN mice, as determined by restriction fragment polymorphism (RFLP) screening and DNA sequence analysis. In severe combined immunodeficiency (C3H/scid) mice, the ruvA mutant retained full infectivity; however, all recovered clones retained the ‘parental’ vlsE sequence, consistent with low rates of vlsE recombination. These results suggest that the reduced infectivity of ruvA and ruvB mutants is the result of ineffective vlsE recombination and underscores the important role that vlsE recombination plays in immune evasion. Based on functional studies in other organisms, the RuvAB complex of B. burgdorferi may promote branch migration of Holliday junctions during vlsE recombination. Our findings are consistent with those in the accompanying article by Dresser et al., and together these studies provide the first examples of trans-acting factors involved in vlsE recombination.

Characterization of the Operon Encoding the Holliday Junction Helicase RuvAB from Mycoplasma genitalium and Its Role in mgpB and mgpC Gene Variation

“…The activity of RuvAB has also been implicated in the antigenic variation systems of vlsE in Borrelia burgdorferi and pilE in Neisseria gonorrhoeae, both of which depend upon recombination (44)(45)(46)(47). In the case of N. gonorrhoeae, another branch migration helicase, RecG, has been implicated (47).…”

Effect of Borrelia burgdorferi OspC at the Site of Inoculation in Mouse Skin

“…The molecule to which OspC binds in the mammalian host might be a signal to which the bacteria respond by changing the composition of the outer membrane, either at the transcriptional or at the posttranscriptional level. For example, the induction of antigenic variation in VlsE is critical for B. burgdorferi survival in the mammalian host (16,37) and relies on an as-yet-unknown signal in the mammalian environment. In fact, the interaction of B. burgdorferi with endothelial cells in vitro was reported previously to induce vlsE expression (31), although VlsE expression is not equivalent to variation.…”

DNA Recombination Strategies During Antigenic Variation in the African Trypanosome

“…To date, however, the machinery needed for an HR reaction of such a short length is unknown. Gene conversion during antigenic variation that is independent of the HR recombinase is not unique to T. brucei: Borrelia burgdorferi is a tick-borne spirochete where recombination of silent vls gene segments into the single vlsE expression site is unaffected by mutation of RecA (169), though it does require RuvAB, a helicase that processes Holliday junction HR intermediates (170,171). Like T. brucei, how homology-directed strand transfer initiates in B. burgdorferi without RecA/RAD51 is unknown, though the potential involvement of a novel, telomere-directed resolvase would be very interesting (172).…”

“…Elevated switching by crossover in T. brucei RTR mutants (117,145) provides evidence not only that reciprocal recombination can occur but also that it is normally suppressed between BES. However, the formation and resolution of Holliday junction intermediates has not been demonstrated, such as by testing for a role of Holliday junction resolvases (219), which have been shown to act in both N. gonorrhoeae and B. burgdorferi antigenic variation (131,170,171). It should be noted that it remains unclear whether any of the above reactions can account for VSG SGC, and this will be an important next phase of investigation, given the importance of this reaction to the parasite.…”