The lysozyme primary transcript has been shown to be slowly spliced, particularly in LPS-activated myelomonocytic HD11 cells. In this study, Northern blot analysis shows that the splicing of lysozyme pre-mRNA in LPS-activated cells is significantly delayed by treatment with KN-62, a selective inhibitor of the Ca2+/calmodulin-dependent protein kinase II (CaMKII), but not with Go 6976 and herbimycin A, inhibitors of Ca2+-dependent PKC and PTK, respectively. In vitro kinase assay using autocamtide 2 as specific substrate for CaMKII demonstrates that KN-62, when added to the extract from HD11 cells, inhibits selectively CaMKII activity. Treatment of HD11 cells with cycloheximide, a potent inhibitor of protein synthesis, results in a transient increase in lysozyme pre-mRNA levels, whilst the mature mRNA levels are not increased. Moreover, neither cycloheximide nor KN-62 has any effect on the glyceraldehyde-3-phosphate dehydrogenase pre-mRNA splicing. Together, our results indicate that phosphorylation by CaMKII, and probably new protein synthesis may be required for the lysozyme pre-mRNA processing.