Abstract

Leptospirosis, a bacterial zoonosis caused by select pathogenic spirochetes belonging to the genus Leptospira , can trigger a systemic inflammatory response syndrome due to a cytokine storm that can occur during infection. However, the molecular mechanisms underlying the extensive inflammatory responses during leptospiral infection remain poorly understood. In this study, we characterized a high temperature requirement protein A (HtrA) homologue encoded by the L. interrogans gene locus LIC12812 , referred herein as LepHtrA. We used AlphaFold 3 to predict the structure of this protein. And we show that LepHtrA is located on the surface of L. interrogans , and its expression is upregulated by increased temperature. An in vitro enzyme assay using purified LepHtrA provides direct evidence of its serine protease activity. We demonstrate that LepHtrA induces a robust pro-inflammatory response in RAW264.7 cells, a mouse macrophage cell line, by promoting their differentiation into pro-inflammatory M1 macrophages. This differentiation leads to an increased production of pro-inflammatory cytokines and the generation of reactive oxygen species, ultimately inducing cellular apoptosis via modulation of Bcl-2/Bax expression. Collectively, our study suggests that LepHtrA is a temperature-induced and surface-exposed antigen that can contribute to cellular inflammation, stress, and apoptosis, potentially influencing the pathogenesis of leptospirosis.