SummaryComplement is viewed as a critical serum-operative component of innate immunity, with processing of its key component, C3, into activation fragments C3a and C3b confined to the extracellular space. We report here that C3 activation also occurred intracellularly. We found that the T cell-expressed protease cathepsin L (CTSL) processed C3 into biologically active C3a and C3b. Resting T cells contained stores of endosomal and lysosomal C3 and CTSL and substantial amounts of CTSL-generated C3a. While “tonic” intracellular C3a generation was required for homeostatic T cell survival, shuttling of this intracellular C3-activation-system to the cell surface upon T cell stimulation induced autocrine proinflammatory cytokine production. Furthermore, T cells from patients with autoimmune arthritis demonstrated hyperactive intracellular complement activation and interferon-γ production and CTSL inhibition corrected this deregulated phenotype. Importantly, intracellular C3a was observed in all examined cell populations, suggesting that intracellular complement activation might be of broad physiological significance.

Factor H-related proteins (FHRs) are a group of partly characterized complement proteins that are thought to promote complement activation by competing binding of factor H (FH) to surface-bound C3b. Among them, FHR-1 is remarkable because is associated with atypical hemolytic uremic syndrome (aHUS) and other important diseases. Using a combination of biochemical, immunological, nuclear magnetic resonance and computational approaches, we have characterized a series of FHR-1 mutants (including two associated with aHUS) and have unraveled the molecular bases of the so-called de-regulation activity of FHR-1. In contrast with FH, FHR-1 lacks the capacity to bind sialic acids, which prevents C3b-binding competition between FH and FHR-1 in host cell surfaces. aHUS-associated FHR-1 mutants are pathogenic because they have acquired the capacity to bind sialic acids, which increases FHR-1 avidity for surface-bound C3-activated fragments and results in C3b-binding competition with FH. FHR-1 binds to native C3, in addition to C3b, iC3b and C3dg. This unexpected finding suggests that the mechanism by which surface-bound FHR-1 promotes complement activation is the attraction of native C3 to the cell surface. Whilst C3b-binding competition with FH is limited to aHUS-associated mutants, all surface-bound FHR-1 promote complement activation, which is delimited by the FHR-1/FH activity ratio. Our data indicate that the FHR-1 de-regulation activity is important to sustain complement activation and C3 deposition at complement activating surfaces. They also support that abnormally elevated FHR-1/FH activity ratios would perpetuate a pathological complement dysregulation at complement activating surfaces, which may explain the association of FHR-1 quantitative variations with diseases.