Analysis of somatic mutation activity in multiple V kappa genes involved in the response to 2-phenyl-5-oxazolone

We have studied somatic mutation activity early in a response to 2-phenyl-5-oxazolone coupled to ovalbumin (phOx-OVA). Although the V kappa Ox1 gene rearranged to J kappa 5 is known to predominate in this response, other closely related V kappa genes are involved. We compared the introduction of point mutations into V kappa Ox1 genes and into a set of related V kappa genes rearranged to the same J kappa segment at two time points after primary immunization. The result showed that quantitation of mutations in a single rearrangement substrate leads to an underestimation of the total mutational activity. There is pronounced somatic mutation activity early within genes that may be absent later in the response. We also show that multiple somatic mutations can be detected in B cells from draining lymph nodes after foot-pad injection with phOx-OVA already at day 7 after immunization. The data suggest a system in which mutation acts early in the response on a wide range of substrates and that selection and expansion of high affinity paratopes occurs later.