Expression of error-prone polymerases in BL2 cells activated for Ig somatic hypermutation

High affinity antibodies are generated in mice and humans by means of somat ic hypermutation (SHM) of variable (V) regions of 1g genes. Mutations with rates of 10(-5)-10(-3) per base pair per generation, about 10(6)-fold above normal, are targeted primarily at V-region hot spots by unknown mechanisms . We have measured mRNA expression of DNA polymerases l, eta, and zeta by u sing cultured Burkitt's lymphoma (BL)2 cells. These cells exhibit 5-10-fold increases in heavy-chain V-region mutations targeted only predominantly to RGYW (R = A or C, Y = C or T, W = T or A) hot spots if costimulated with T cells and IgM crosslinking, the presumed in vivo requirements for SHM. An similar to4-fold increase pol l mRNA occurs within 12 h when cocultured wit h T cells and surface IgM crosslinking. Induction of pols eta and zeta occu r with T cells, IgM crosslinking, or both stimuli. The fidelity of pol 1 wa s measured at RGYW hot-and non-hot-spot sequences situated at nicks, gaps, and double-strand breaks. Pol l formed T.G mispairs at a frequency of 10(-2 ), consistent with SHM-generated C to T transitions, with a 3-fold increase d error rate in hot- vs. non-hot-spot sequences for the single-nucleotide o verhang. The T cell and IgM crosslinking-dependent induction of poi 1 at 12 h may indicate an SHM "triggering" event has occurred. However, pols l, et a, and zeta are present under all conditions, suggesting that their presenc e is not sufficient to generate mutations because both T cell and IgM stimu li are required for SHM induction.