Deciphering the Role of Leptospira Surface Protein LigA in Modulating the Host Innate Immune Response

Abstract

Leptospira, a zoonotic pathogen, is known to infect various hosts and can establish persistent infection. This remarkable ability of bacteria is attributed to its potential to modulate (activate or evade) the host immune response by exploiting its surface proteins. We have identified and characterized the domain of the variable region of Leptospira immunoglobulin-like protein A (LAV) involved in immune modulation. The 11th domain (A11) of the variable region of LigA (LAV) induces a strong TLR4 dependent innate response leading to subsequent induction of humoral and cellular immune responses in mice. A11 is also involved in acquiring complement regulator FH and binds to host protease Plasminogen (PLG), there by mediating functional activity to escape from complement-mediated killing. The deletion of A11 domain significantly impaired TLR4 signaling and subsequent reduction in the innate and adaptive immune response. It also inhibited the binding of FH and PLG thereby mediating killing of bacteria. Our study discovered an unprecedented role of LAV as a nuclease capable of degrading Neutrophil Extracellular Traps (NETs). This nuclease activity was primarily mediated by A11. These results highlighted the moonlighting function of LigA and demonstrated that a single domain of a surface protein is involved in modulating the host innate immune defenses, which might allow the persistence of Leptospira in different hosts for a long term without clearance.

LigA formulated in AS04 or Montanide ISA720VG induced superior immune response compared to alum, which correlated to protective efficacy in a hamster model of leptospirosis

“…The LAV was purified as described previously ( 74 ). Briefly, BL21(DE3) carrying the expression plasmid (pET28a-LAV) were grown at 37°C overnight on LB broth containing 50 μg/ml kanamycin, and the expression of the protein was induced with 1 mM isopropyl β-D-1-thiogalactoside (IPTG).…”

Screening of Surface-Exposed Lipoproteins of Leptospira Involved in Modulation of Host Innate Immune Response

“…Despite identification and characterization of several surface proteins of Leptospira involved in innate activation and/or complement evasion, not much effort is made toward the identification of immunomodulatory surface proteins, which are simultaneously involved in both immune activation and evasion. Recently, our group has demonstrated the immunomodulatory role of Leptospira immunoglobulin-like protein A (LigA) and characterized the domain involved in activation and evasion from host innate immune response ( Kumar et al, 2021 ). Considering the functional redundancy in Leptospira surface proteins, in the present study, we aimed to identify and characterize the surface-exposed lipoproteins having this immunomodulatory or dual role that may help in better understanding their critical role in host–pathogen interaction.…”

“…Upon encounter with neutrophils, Leptospira can induce NET; however, to evade NETosis, whether it expresses nucleases has not been reported yet ( Scharrig et al, 2015 ). For the first time, our group reported the role of LigA in degrading NET by its nuclease activity ( Kumar et al, 2021 ). Considering the functional redundancy in surface proteins of Leptospira , it is likely that other surface-exposed lipoproteins might also possess nuclease activity.…”

“…The concentration of purified protein was estimated using the Bradford reagent (Sigma B6916). The recombinant variable region of Leptospira immunoglobulin like protein A (LAV) was purified as described previously ( Kumar et al, 2021 ).…”

Pathogen-Derived Nucleases: An Effective Weapon for Escaping Extracellular Traps

“…Leptospira immunoglobulin-like proteins (Lig) are surface proteins expressed in pathogenic strains of Leptospira , including LigA, LigB, and LigC. The Lig proteins were involved in leptospiral adhesion, complement resistance, and modulation of hemostasis and play a vital role in invasion and immune evasion ( 117 ).The recombinant variable region of LigA was able to degrade DNA with both endonuclease and exonuclease activities, and could degrade PMA-induced NETs ( 118 ).…”