Expression and factor-dependent modulation of the interleukin-3 receptor subunits on human hematopoietic cells.
Interleukin-3 (IL-3) regulates growth and differentiation of multipotential as well as lineage-committed progenitor cells. The human IL-3 receptor (IL-3R) consists of the alpha and common beta (beta c) subunits. The alpha subunit (IL-3R alpha) is specific for IL-3 and binds IL-3 with low affinity. In contrast, the beta c subunit does not bind any cytokine by itself, but forms a high-affinity receptor with IL-3R alpha. As the same beta c subunit also forms high-affinity receptors for IL-5 and granulocyte-macrophage colony-stimulating factor (GM-CSF) with the respective cytokine-specific alpha subunit, the expression of the alpha subunits is responsible for specificity of cytokines. To examine the expression of IL-3R alpha, we have developed a monoclonal antibody (MoAb), N3A. N3A specifically bound to cells expressing IL-3R alpha and immunoprecipitated a 75 Kd glycoprotein, which became 43 Kd on N-glycosidase digestion. N3A and an anti-beta c antibody, CRS1, were used in double color fluorescence-activated cell sorter (FACS) staining with several lineage markers to see the IL-3R expression pattern in peripheral blood (PB), cord blood (CB), and bone marrow (BM) cells. Both IL-3R subunits were expressed on myeloid cell lineages (CD13+, CD14+, CD15Lo, or CD33+). To further study the IL-3R expression on hematopoietic progenitor cells, the CD34+ populations were isolated from both BM and CB cells. Those populations showed positive staining profiles with the N3A MoAb and were weakly stained with the CRS1 MoAb. Furthermore, anti c-kit antibody staining of the CD34+ fraction from CB, but not from BM, showed two intensities and the IL-3R alpha expression seemed to be higher in a fraction of low c-kit expression. Because IL-1, IL-6, G-CSF, stem cell factor (SCF), interferon (IFN)-gamma, and tumor necrosis factor (TNF)-alpha are known to enhance IL-3-dependent colony formation, we have examined whether this enhancement could be correlated with upregulation of the IL-3R expression. Incubation of CD34+ cells with TNF-alpha for 2 days significantly increased the level of beta c and G-CSF increased the number of cells with high level expression of alpha, while other factors did not affect the IL-3R expression. Thus, different cytokines appear to have different mechanisms for enhancement of IL-3-dependent proliferation.
Sato
N
N
Department of Molecular Biology, DNAX Research Institute of Molecular and Cellular Biology, Palo Alto, CA 94304.
Caux
C
C
Kitamura
T
T
Watanabe
Y
Y
Arai
K
K
Banchereau
J
J
Miyajima
A
A
D000911:Antibodies, Monoclonal; D015703:Antigens, CD / Q000032:analysis; D018952:Antigens, CD34; D001854:Bone Marrow Cells; D002460:Cell Line; D016207:Cytokines / Q000494:pharmacology; D005312:Fetal Blood; D005434:Flow Cytometry; D006412:Hematopoietic Stem Cells / Q000378:metabolism*; D006801:Humans; D016130:Immunophenotyping; D066298:In Vitro Techniques; D011233:Precipitin Tests; D016185:Receptors, Interleukin-3 / Q000276:immunology / Q000378:metabolism*
D000911:Antibodies, Monoclonal; D015703:Antigens, CD; D018952:Antigens, CD34; D016207:Cytokines; D016185:Receptors, Interleukin-3