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Crry, but not CD59 and DAF, is indispensable for murine erythrocyte protection in vivo from spontaneous complement
Takashi Miwa1, Lin Zhou, Brendan Hilliard
1Center for Experimental Therapeutics and Department of Pharmacology, University of Pennsylvania School of Medicine, Philadelphia 19104, USA.
Insights
Mice lacking decay-accelerating factor (DAF) or CD59 on erythrocytes did not develop anemia. Complement receptor 1 (CR1)-related gene y (Crry) was essential for preventing complement-mediated erythrocyte destruction in mice.
Area of Science:
- Immunology
- Complement System
- Hematology
Background:
- Decay-accelerating factor (DAF) and CD59 are membrane proteins that regulate complement activation.
- CD59 deficiency in humans causes erythrocyte sensitivity to complement lysis, suggesting its critical role.
- The in vivo roles of DAF and CD59 in erythrocytes remain to be fully elucidated.
Purpose of the Study:
- To investigate the in vivo significance of CD59 and DAF in protecting erythrocytes from complement attack.
- To compare the roles of CD59, DAF, and complement receptor 1 (CR1)-related gene y (Crry) in murine erythrocyte protection.
Main Methods:
- Generation and analysis of CD59 knockout and CD59/DAF double-knockout mice.
- Assessment of erythrocyte sensitivity to complement lysis in vitro.
- Evaluation of erythrocyte lifespan and complement deposition in vivo.
- Study of Crry-deficient mouse erythrocytes.
Main Results:
- CD59-deficient and CD59/DAF-double-deficient mouse erythrocytes showed high sensitivity to antibody-induced complement lysis in vitro.
- Neither CD59 knockout nor CD59/DAF double-knockout mice developed spontaneous hemolytic anemia, and their erythrocytes had normal in vivo lifespan.
- Crry-deficient erythrocytes were rapidly cleared from circulation, exhibiting increased spontaneous C3 deposition compared to DAF-deficient erythrocytes.
Conclusions:
- CD59 and DAF are not essential for protecting murine erythrocytes from spontaneous complement attack.
- Crry, a C3 inhibitor, is crucial for regulating complement activation on mouse erythrocytes.
- CR1, not DAF, may be the primary regulator of spontaneous C3 activation on human erythrocytes.
Abstract:
Decay-accelerating factor (DAF) and CD59 are 2 glycosylphosphatidylinositol-anchored membrane proteins that inhibit complement activation at the C3 and C5b-9 step, respectively. CD59 is considered critical for protecting erythrocytes from spontaneous complement attack, as deficiency of CD59 or CD59/DAF, but not of DAF alone, on human erythrocytes renders them sensitive to complement lysis in paroxysmal nocturnal hemoglobinuria syndrome. To evaluate the relative roles of CD59 and DAF in vivo, we have generated and studied a CD59 knockout and a CD59/DAF double-knockout mouse. CD59-deficient and CD59/DAF-double-deficient mouse erythrocytes were highly sensitive to antibody-induced complement lysis in vitro, yet neither CD59 knockout nor CD59/DAF double-knockout mouse developed spontaneous hemolytic anemia. Consistent with the latter observation, erythrocytes from the 2 strains of mutant mice were shown to have a normal lifespan in vivo. In contrast, mouse erythrocytes deficient in complement receptor 1 (CR1)-related gene y (Crry), a membrane C3 inhibitor with DAF and membrane cofactor protein activities, were rapidly eliminated from the circulation by a complement-dependent mechanism. Compared with DAF-deficient erythrocytes, Crry-deficient erythrocytes incurred higher levels of spontaneous C3 deposition in vivo. These findings demonstrate that CD59 and DAF are not indispensable on murine erythrocytes. Rather, effective C3 regulation on the cell surface, provided by Crry rather than DAF, is necessary for mouse erythrocytes to resist spontaneous complement attack. Our results raise the possibility that proper control of C3 activation may also be critical on human erythrocytes, where CR1 but not DAF could be the principal regulator of spontaneous C3 activation.