can cause a range of diseases that vary in manifestations and outcome depending on the bacterial strain and host susceptibility (1
). In this study, we analyzed host responses after infection with S. pyogenes
strain S165 of type emm6
isolated from a patient with severe invasive disease. We demonstrated that infection induced the apoptosis and shedding of CD46 from host epithelial cells. Recombinant soluble CD46 containing the extracellular protein domains bound to S. pyogenes
in mid-log phase. Furthermore, bacteria survived better in blood of CD46 transgenic mice than in blood of nontransgenic mice lacking CD46, and in vivo infection showed that the presence of human CD46 increased the severity of disease. These data suggest that human CD46 plays an important role during S. pyogenes
CD46 serves as a receptor for several pathogens including strains of measles virus (10
), human herpesvirus type 6 (42
), group A streptococci (38
), adenovirus (13
), and Neisseria
). Among these pathogens, infection by certain strains of measles virus (33
), human herpesvirus type 6 (42
), serogroup B adenovirus (41
), and Neisseria gonorrhoeae
) has been shown to cause CD46 downregulation from the cell surface. The detailed mechanisms of surface CD46 downregulation upon infection by these pathogens remain to be elucidated; however, the decrease in the surface density of CD46 renders the cells more susceptible to lysis by complement, as demonstrated in vitro (44
), and may contribute to the attenuation of these pathogens by the rapid clearing of infected cells.
Elward et al. (11
) and Cole et al. (6
) demonstrated that the chemical induction of apoptosis results in the specific release of CD46. These investigators revealed that apoptotic cells shed CD46 in the form of apoptotic membrane vesicles or blebs and that necrotic cells release soluble CD46 protein (6
). We show that the chemical induction of apoptosis also triggers apoptosis in FaDu epithelial cells and that this results in the shedding of soluble CD46 from the cell surface. After infection of the cell with S. pyogenes
, apoptotic cell death takes place, and CD46 is released into the extracellular environment. CD46 may be released by a specific cleavage at the cell surface or as part of membrane apoptotic blebs. It is possible that a combination of these release processes occurs at different time points during infection. The downregulation of CD46 expression on epithelial cells after Neisseria gonorrhoeae
infection results in the release of membrane vesicles (16
). Future studies will reveal if S. pyogenes
infection triggers the release of CD46 membrane vesicles, a free CD46 cleavage product, or a combination of both. The finding that HLA protein expression remained stable when β1 integrin expression was reduced supports that CD46 is closely linked to β1 integrin, as was previously shown (38
It is well known that bacterial protein or gene expression may vary dramatically between different growth phases or growth conditions (3
). Several virulence factors of S. pyogenes
have been shown to be growth phase dependent (50
). Expression of the M protein, the C5a peptidase, and capsule are maximal at the exponential growth phase, whereas streptococcal pyogenic exotoxins A and B and mitotic factor are maximally expressed in later phases of growth. We found that the best binding of CD46 to S. pyogenes
occurred at mid-log phase. Several lines of evidence support that CD46 can interact with the M protein (14
); however, it cannot be excluded that another surface component of S. pyogenes
binds to CD46 as well.
As shown in the ex vivo blood assay, bacteria were able to survive better in blood from transgenic mice expressing human CD46 than in blood from nontransgenic mice. One possibility is that S. pyogenes uses extracellularly released CD46 to mask and hide from the immune system. Since S. pyogenes is a human-specific pathogen, there must be specific factors of the bacteria and the host that cause this specificity and prevent other species from being affected by severe S. pyogenes disease. The data in this work suggest that human CD46 is one such host factor. However, the in vivo experiments were performed in a murine model system, and it is yet to be determined whether CD46 also has similar properties in humans.
To further evaluate the importance of CD46, we infected transgenic mice expressing CD46. These mice express CD46 in a human-like manner and elicit immune responses similar to that of humans as a consequence of bacterial attack (21
), suggesting that the mouse model is an adequate experimental in vivo system. Experimental infection revealed that CD46 transgenic mice developed a higher level of bacteremia than nontransgenic mice, supporting that CD46 enhances blood survival. In addition, bacterial numbers increased over time and reached the highest level at day 3 postinfection, showing that bacteria were able to grow in blood. It is well known that S. pyogenes
avoids the complement alternative pathway attack by binding soluble complement regulators such as factor H or C4b binding protein on their surface (20
). The data in this work argue that soluble and shed CD46 could also play a role in protecting bacteria against complement alternative pathway attack. The development of arthritis occurred more often in CD46 transgenic mice than in nontransgenic mice but was not always linked to high bacterial blood counts (data not shown), indicating that inflammatory factors are likely involved in this process. Finally, the higher mortality of CD46 transgenic mice than nontransgenic mice argues that CD46 facilitates severe lethal disease. The findings that bacteria survived better in blood from CD46 transgenic mice than in blood from nontransgenic mice and that bacteria preincubated with recombinant CD46 survived better in human blood support that CD46 increases bacterial survival and that soluble CD46 is important for bacterial survival. However, we cannot exclude that the increased virulence in CD46 transgenic mice is caused by enhanced levels of CD46-mediated adherence or invasion of certain cell populations. In future studies, we will further evaluate the role of CD46 in bacterium-host cell interactions.
This study demonstrates that S. pyogenes induces apoptosis and the release of CD46 and that bacteria bind soluble CD46. The interaction with CD46 might help to overcome innate immune defenses since bacteria survived better in blood and were more lethal in the presence of human CD46. However, it cannot be excluded that bacterial interaction with CD46 mediates host cell signal transduction that in turn affects innate immune defenses against the disease. Taken together, the data in this work support that S. pyogenes interacts with soluble free CD46 and that CD46 plays an important role during S. pyogenes infection by contributing to bacterial survival.