B cell activation involves nanoscale receptor reorganizations and inside-out signaling by Syk

Abstract

Binding of antigen to the B cell antigen receptor (BCR) initiates a multitude of events resulting in B cell activation. How the BCR becomes signaling-competent upon antigen binding is still a matter of controversy. Using a high-resolution proximity ligation assay (PLA) to monitor the conformation of the BCR and its interactions with co-receptors at a 10–20 nm resolution, we provide direct evidence for the opening of BCR dimers during B cell activation. We also show that upon binding Syk opens the receptor by an inside-out signaling mechanism that amplifies BCR signaling. Furthermore, we found that on resting B cells, the coreceptor CD19 is in close proximity with the IgD-BCR and on activated B cells with the IgM-BCR, indicating nanoscale reorganization of receptor clusters during B cell activation.Our immune system protects us against diseases by recognizing invading pathogens, such as bacteria and viruses, and launching a response to eliminate them. In vertebrates, like mice and humans, this immune response often involves white blood cells called B cells, which make antibodies.B cells can recognize a huge number of different molecules called antigens, including those from pathogens, with the help of their antigen receptors. These receptors are proteins that span the surface membrane of the B cells, such that most the receptor is outside of the cell, with the rest being inside the cell. When an antigen binds to the outside portion of a B cell receptor, that B cell becomes activated. The B cell then starts to multiply, and to produce antibodies that bind to that antigen and hence mark a pathogen for attack by the immune system.For many years it was thought that two copies of the receptors had to be brought together for the B cell antigen receptor to activate the B cell. However, other research revealed that the receptors tend to cluster together in the membrane, even before an antigen is recognized. Now, Kläsener, Maity et al. have used techniques that can essentially measure the distance between two B cell antigen receptors, even when they are just a few billionths of a meter (or nanometers) apart. This revealed that the receptors start very close together, and actually move further away from each other when the B cells are activated.Kläsener, Maity et al. also found that an enzyme, called spleen tyrosine kinase (Syk), is needed to separate the receptors. Further experiments revealed that Syk does this by binding to the so-called ‘signaling motif’ of the receptors, which is inside the cell: this causes the receptors to change shape, forcing the parts outside the cell to move apart. Furthermore, Kläsener, Maity et al. found that other proteins in the surface membrane called co-receptors—which cooperate with the B cell antigen receptors to activate a B cell—were also re-organized when B cells became activated.It is likely that most other membrane proteins are also organized in clusters that are only nanometers across. As such, the techniques described by Kläsener, Maity et al. will now allow the study of membrane organization at the nanoscale; which, as yet, has remained largely unexplored.