Intracellular competition regulates B lymphocyte differentiation

The production of antibodies, with their potential to recognise unique targets and prevent repeat infections, is an important aspect of immune health. In order to generate free antibodies, the cells responsible, B cells, must undergo a differentiation step to transform from lymphoblast to antibody secreting cell (ASC). This differentiation step prevents further antibody modifications and hence the timing for optimal immunity requires a delicate balance between expanding useful clones and providing early protection. How differentiation is controlled to achieve this balance for an effective immune response is of great interest. In this study, the progression from naive B cells to ASC was investigated in the context of an emerging model for competing cell fates. By this model, alternate cell fates, such as division, death and differentiation, are pursued independently in individual cells but are in competition such that events which occur earlier prevent those that require more time from being observed. Evaluation and testing of this model requires careful measurement of distributions of times to fates which is only possible with single cell fate tracking. Here I have developed and applied methods for live cell imaging and analysis for assessing and evaluating cell fate changes over time. Using these methods, several modes of regulating differentiation times were revealed. Low levels of stimulation through CD40 produced a greater proportion of antibody secreting cells per generation as division is slowed and more time is allowed for differentiation, consistent with competing cell fates. A second mechanism was found where increasing division numbers directly reduced the amount of time required for cells to differentiate, without modulating division times, ensuring the natural development of ASC during the ongoing immune response. A direct method of uncensoring was explored where cell cycle inhibitors were used to prevent division, with the hypothesis that more cells wou