Signals that initiate somatic hypermutation of B cells in vitro

Somatic hypermutation is initiated as B lymphocytes proliferate in germinal centers. The signals that switch on the mutation process are unknown. We h ave derived an in vitro system to define signals that will initiate mutatio n in normal, naive splenic B cells. We find that three signals are required to allow detection of somatic mutation in vitro; these are anti-Ig, anti-C D40, and anti-CD38. If any one of these is omitted, mutation remains off. W e show that CD40 is obligatory in vivo, as CD40 knockout mice exhibit no Ag -driven mutation. In contrast, CD38 is not, as CD38 knockout mice mutate no rmally. We believe that, in vitro, CD38, in combination with other stimuli, drives extensive cell division, allowing the detection of mutated sequence s. However, in germinal centers in vivo, proliferative activity is instigat ed by a different molecule. This is the first demonstration of the initiati on of hypermutation in vitro with normal splenic B cells using defined stim uli.