Investigating the interaction of soluble host proteins (SP-D, C1q and fibronectin) with Mycobacteria

Abstract

Mycobacterium tuberculosis (Mtb), one of the major pathogens of mankind, kills approximately 2 million people each year. Mtb induces inflammation at the site of infection, leading to leakage of serum proteins, which in turn, are likely to come in contact with the pathogen, thus modulate the pathogenesis of tuberculosis. We studied some of these proteins such as surfactant protein D (SP-D), complement protein C1q and fibronectin, which are either produced locally or they leak-out from serum during inflammation, for their interaction with M.smegmatis and BCG. These non-pathogenic mycobacteria were used as model for Mtb. In this study, the recombinant form of truncated human surfactant protein D (rhSP-D) and three globular heads of human C1q (ghA, ghB, and ghC) were expressed in E.coli. The interaction of each of these proteins with mycobacteria and human monocytic cell line THP-1, was examined via ELISA. We demonstrated that rhSP-D, C1q, three globular heads of C1q and fibronectin bind with both mycobacteria and THP-1 cells. Moreover, using rhSP-D and globular heads of C1q, the binding of SP-D and C1q was localised to C-terminal globular regions. The direct effect for each of these proteins on mycobacterial growth, their effect on the uptake and intracellular fate of mycobacteria inside THP-1 cells were also investigated. Direct interaction of rhSP-D and C1q inhibited mycobacterial growth, whereas fibronectin interaction with the mycobacteria increased their growth. RhSP-D inhibited the uptake and growth of mycobacteria inside THP-1 cells, whereas C1q and each individual globular heads of C1q enhanced the uptake of mycobacteria by THP-1 cells. However, C1q protein inhibited BCG growth but enhanced M.smegmatis growth inside these cells and the later activity was localised to ghA. Fibronectin increased the uptake and growth of mycobacteria inside THP-1 cells. Examining the gene expression of inducible nitric oxide synthase, pro-inflammatory and anti-inflammatory cytokines produced by THP-1 cells infected with the proteins treated and untreated mycobacteria, along with cytokine neutralization experiments, suggest that the nitric oxide components and cytokines could be responsible for mycobacterial growth control inside THP-1 cells. These novel and interesting functions of SP-D, C1q, and fibronectin on mycobacteria provide an insight into the modulatory function of these proteins on Mtb infection, and, therefore, in the pathogenesis of tuberculosis.