Immune reactions protect our bodies from dangers such as foreign pathogens (bacteria, viruses, fungi), cellular transformation or help during wound healing. Our immune system is able to recognize and fight but also to learn thanks to lymphocytes. B lymhpocytes (B cells) are responsible for antibody production and immunological memory. A B cell carries an antibody structure – a specialized B-cell receptor also called immunoglobulin (Ig) - on its surface.
During a B cell life, there are two time points when the Ig is actively engaged and signals: in early life when Ig is produced and in an immune reaction when Ig engages with an intruder. Between, there is a window of a month or two when B cells are resting.
This evoked curiosity, do B cells need the Ig in a resting state? The laboratory of Prof. Klaus Rajewsky discovered that resting B cells depend on the expression of Ig, without Ig B cells die. A “tonic signal”, a hypothetical signal coming from the Ig in a resting state has been proposed as essential for the maintenance of B cells. However, attempts to identify the trigger which activates such tonic signal have been vain.
In the BCRdangerCOMPETITION project, we addressed alternative mechanisms which might induce B cell death upon loss of the Ig – a cellular stress response, particularly unfolded protein response (UPR). This is initiated once a cell synthetizes more proteins than it can fold (shape into correct confirmation) and in order to re-establish equilibrium induces the expression of proteins involved in stress response, e.g. facilitators of protein folding. If the stress resolves, the cell survives and turns down the stress response, however, if the stress persists, the prolonged overexpression of proteins involved in UPR leads to cell death. Ig is a complex of multiple proteins that assemble inside the cells and are transferred as Ig complex to the cellular surface. When among the proteins forming the Ig complex we delete one essential part, the other Ig proteins remain expressed but cannot form the Ig complex. In theory, they could accumulate, overwhelm the folding machinery of a cell activating the stress response which kills the B cell. To our surprise and contrary to our hypothesis, B cells without Ig were unable to induce the mentioned stress response. We propose a model where the presence of Ig is required to keep the expression of proteins involved in stress response at physiological levels which maintain the function of cellular organelles and anabolic metabolism. Although counterintuitive at first sight, stress response at physiological levels maintained by the presence of Ig is vital for cellular survival. This finding might uncover a novel survival signal that requires the physical presence of Ig to maintain the stress response functional at physiological levels.
In parallel, we addressed another potential mechanism responsible for B cell death upon loss of the Ig: cellular competition. The phenomenon of cellular competition has recently been identified as the cause of disappearance of Ig-negative lymphoma cells (malignant B cells). While lymphoma cells without Ig can grow normally in isolation, they disappear when placed in competition with Ig-positive lymphoma cells. We addressed the extent of cellular competition in primary B cells using the novel CRISPR/Cas9 mediated mutagenesis of primary B cells. We have set up an efficient protocol to induce Ig deletion in primary B cells and observed them under conditions resembling an immune reaction. B cells without Ig were unable to persist and were outcompeted by their Ig-positive counterparts in only two days. So far, our results show that Ig deletion is detrimental for primary B cells growing under conditions resembling an immune reaction and provide experimental evidence for an old observation, namely, that Ig- B cells cannot be detected after an immune reaction in mice or humans. We are currently working on improving our experimental setup to obtain pure Ig-negative B cells aiming to understand how Ig-negative B cells behave in immune cultures in absence of competition.
But why is this basic B cell research important for society? B cells, when healthy, are an indispensable part of our immunity. However, in diseases, such as autoimmunity, allergies or malignant B cell transformation into leukemia or lymphoma, B cells need to be fought against. Therefore, knowledge of basic B cell biology, the signals and pathways which keep B cells alive are necessary in order to target malignant B cells at their weak spots. My research might therefore one day be an important source of information indicating where such weak spots in B cells are.
