Fusobacterium nucleatum inhibits human T-cell activation by arresting cells in the mid-G1 phase of the cell cycle

Abstract

Fusobacterium nucleatum has been implicated in the pathogenesis of several diseases, including urinary tract infections, bacteremia, pericarditis, otitis media, and disorders of the oral cavity such as pulpal infections, alveolar bone abscesses, and periodontal disease. We have previously demonstrated that sonic extracts of F. nucleatum FDC 364 were capable of inhibiting human T-cell responses to mitogens and antigens. In this study, we have further characterized this immunosuppressive protein (FIP) and initiated experiments to determine its mode of action. The purified FIP has an apparent molecular mass of 90 to 100 kDa; sodium dodecyl sulfate-polyacrylamide gel electrophoresis indicates that the FIP is actually composed of two subunits with molecular masses of 48 and 44 kDa. Purified FIP retained its biological activity and was capable of inhibiting mitogen-induced proliferation of human T cells. Inhibition was dose dependent, and the FIP exhibited a specific activity approximately 250-fold greater than that of the crude extract. Cell cycle analysis indicates that FIP-treated cells were prevented from exiting the G0/G1 phase of the cell cycle. However, FIP did not alter the expression of activation markers (CD69, CD25, and CD71) or interleukin-2 secretion. The latter observations suggest that the T cells did indeed become activated and had entered the G1 phase of the cell cycle. Analysis of the expression of cyclins indicates that the phase of the cell cycle that is FIP sensitive resides somewhere beyond the restriction point of cyclin D2 (early to mid-G1) but prior to that of cyclins D3 and E (mid- to late G1). Finally, analysis of the expression of the proliferating cell nuclear antigen indicates that this is the earliest detectable defect in T cells exposed to FIP. We propose that if a block in the G1 phase of the cell cycle occurs in vivo in lymphocytes, it may result in a state of local and/or systemic immunosuppression. These suppressive effects could alter the nature and consequences of host-parasite interactions, thereby enhancing the pathogenicity of F. nucleatum itself or that of some other opportunistic organisms.