Abstract
Ryanodine receptor (RYR) is a Ca(2+) channel that mediates Ca(2+) release from intracellular stores. We have used RT-PCR analysis and examined its expression in primary peripheral mononuclear cells (PBMCs) and in 164 hemopoietic cell lines. In PBMCs, type 1 RYR (RYR1) was expressed in CD19(+) B lymphocytes, but less frequently in CD3(+) T lymphocytes and in CD14(+) monocytes. Type 2 RYR (RYR2) was mainly detected in CD3(+) T cells. Induction of RYR1 and/or RYR2 mRNA was found after treatment with stromal cell-derived factor 1, macrophage-inflammatory protein-1alpha (MIP1alpha) or TGF-beta. Type 3 RYR (RYR3) was not detected in PBMCs. Many hemopoietic cell lines expressed not only RYR1 or RYR2 but also RYR3. The expression of the isoforms was not associated with specific cell lineage. We showed that the RYR-stimulating agent 4-chloro-m-cresol (4CmC) induced Ca(2+) release and thereby confirmed functional expression of the RYR in the cell lines expressing RYR mRNA. Moreover, concordant induction of RYR mRNA with Ca(2+) channel function was found in Jurkat T cells. In untreated Jurkat T cells, 4CmC (>1 mM) had no effect on Ca(2+) release, whereas 4CmC (<400 microM) caused Ca(2+) release after the induction of RYR2 and RYR3 that occurred after treatment with stromal cell-derived factor 1, macrophage-inflammatory protein-1alpha, or TGF-beta. Our results demonstrate expression of all three isoforms of RYR mRNA in hemopoietic cells. Induction of RYRs in response to chemokines and TGF-beta suggests roles in regulating Ca(2+)-mediated cellular responses during the immune response.
Highlights
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RYR3 mRNA was detected at lower levels than RYR1 in skeletal muscle (Fig. 1A)
The RT-PCR products amplified for the RYR1, RYR2, and RYR3 from skeletal muscle, cardiac muscle, and brain, respectively, were digested with restriction enzymes HgaI, BsmI, and HindIII
Summary
We examined a total of 164 human hemopoietic cell lines (36 T cell, 92 B cell, 19 myelomonocytic, 11 megakaryocytic, 3 erythrocytic, and 3 nonlymphocytic, nonmyelocytic) to determine the lineage and differentiation specificity of the expression of the 3 RYRs. The possibility that any isoform of RYR is induced by stimulation with mitogens, chemokines, and other stimuli were investigated to gain insight of the roles of this Ca2ϩ release channel in immune function. Using this selective RT-PCR/RFLP method, we examined the expression of three isoforms of RYR mRNA in T cell, B cell, and monocyte populations purified from nine different donors (Fig. 2 and Table I).
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