Calreticulin is an interleukin-3-sensitive calcium-binding protein in human basophil leukocytes

Background: IL-3 Enhances basophil histamine release upon stimulation with any known secretagogue. The molecular mechanism behind this regulation is n ot known, although some observations suggest that IL-3 modulates the calciu m part of the signal transduction mechanism. The inhibitory action of gluco corticoids on basophils can be reversed by stimulation with IL-3. Methods: Calcium-binding proteins in the basophil cell line KU812 were iden tified by two-dimensional gel electrophoresis, Calcium-overlay assay, N-ter minal sequence analysis, and mass spectometry. The presence of the same pro teins in purified human basophil leukocytes was established by comigration of KU812 and human basophil proteins on the two-dimensional gels. The expre ssion of the calcium-binding proteins in the absence and presence of IL-3 a nd/or anti-IgE was determined by densitometric measurement of the spots on the two-dimensional gels. Results: Calreticulin was identified on the two-dimensional gel of KU812 pr oteins. A protein with exactly the same migration pattern was found on the gels of proteins from purified human basophils. Immunoblotting with a speci fic antihuman calreticulin antibody confirmed that this protein was calreti culin. Subsequent analysis showed that the expression of calreticulin in th e basophils is upregulated twofold upon stimulation with rhIL-3, even in do ses below those needed for enhancement of histamine release. Conclusions: The expression of calreticulin in human basophil leukocytes is regulated by IL-3. Calreticulin is known to modulate IP3-dependent Ca2+ in flux in different cell systems, and calreticulin overexpression inhibits st eroid-induced transcriptional activation. Therefore, modulation of calretic ulin expression may be one mechanism by which IL-3 exerts its effects on hu man basophils.