The Trypanosoma brucei VSG-macrophage interface: Recognition, signaling, and innate immune regulation

The work outlined in this thesis describes the immunoregulatory properties of the variant surface glycoprotein of Trypanosoma brucei and the mechanisms of immune activation and modulation associated with recognition of this molecule by macrophages. Increasing evidence has documented that membrane-form (mfVSG) and soluble VSG (sVSG) can induce proinflammatory macrophage activating effects or downregulatory effects. However, little is known about how either of these molecules is recognized by the macrophage to induce these responses. We hypothesized that transfer of the GPI-anchored mfVSG molecule occurs between trypanosomes and host red and white cell populations, which results in immune-dependent responses by or against cells containing transferred VSG. Complement-mediated lysis studies indicated that mfVSG transfer occurs between trypanosome and erythrocyte membranes and suggests that transferred mfVSG may be responsible for anemia in infected animals. However, transfer to monocyte and macrophage membranes was not detected.; Analysis of the sVSG molecule revealed that receptor-mediated recognition of sVSG by macrophages induces the NF-kappaB cascade. Macrophage activation by sVSG following treatment with the TRAF6 inhibitory peptide DIVK resulted in enhanced degradation of IkappaBalpha, suggesting both that TRAF6-dependent Toll-like receptor (TLR) activation of the pathway alone is not required for signaling and that TLR pathway components may negatively regulate expression of sVSG-induced signaling. Binding of sVSG to the macrophage membrane is saturable and inhibited by the macrophage scavenger receptor-specific ligand mBSA, and activation of the NF-kappaB cascade requires recognition and internalization of this molecule through an actin-dependent, clathrin-independent, acidification-independent phagocytic pathway. Further studies show that sVSG stimulates inhibitory and activating NF-kappaB subunit transcription factor complexes that regulate activation and modulation of gene promoter activity. Nuclear translocated NF-kappaB complexes consist primarily of the transcriptionally inhibitory p50/p50 homodimer. Overall, these seemingly diverse responses demonstrate that sVSG can directly induce both activating and inhibitory responses in macrophages and are the first example of a parasite immunomodulatory molecule capable of controlling immune responses through both positive and negative regulatory means.