Functional nuclei of LPS-nonresponder C3H/HeJ mice after transfer into LPS-responder C3H/HeN cells by cell fusion

Abstract

Splenic lymphocytes of C3H/HeJ mice are defective in responding to the mitogenic activity of bacterial lipopolysaccharide (LPS), which has been attributed to a mutation at a single gene locus on chromosome 4 (ref. 5). It was suggested that C3H/HeJ mice lack a cell-surface receptor necessary for LPS recognition. However, specific binding of 125I-labelled lipid-A of LPS to B cells was found to be equivalent in LPS-responder and LPS-nonresponder C3H/HeJ, C57BL/10 ScCR and C57BL/10 ScN mice. This suggests that the genetic defect in C3H/HeJ mice is related to triggering mechanisms rather than to LPS binding. We recently established a method for separating mouse splenic lymphocytes into karyoplasts (minicells) and cytoplasts (enucleated cells) with high purity. This method makes possible the exchange of nuclei from one type of lymphocyte to another. In the present study, karyoplasts were purified from LPS-nonresponder C3H/HeJ mice and transferred to mitomycin C-treated or X ray-irradiated splenic lymphocytes taken from LPS-responder C3H/HeN mice by fusion with polyethyleneglycol. These reconstituted hybrid cells could respond to LPS, suggesting that nuclei from C3H/HeJ mice could be activated by LPS if proper signals were provided from cell-surface receptors. Moreover, responsiveness to concanavalin A (Con A) of mitomycin C-treated or X ray-irradiated lymphocytes was also recovered by the microinjection of karyoplasts from untreated lymphocytes.