calcimycin and Precursor-Cell-Lymphoblastic-Leukemia-Lymphoma

Whether tumor suppressor WWOX (WW domain-containing oxidoreductase) stimulates immune cell maturation is largely unknown. Here, we determined that Tyr-33-phosphorylated WWOX physically binds non-phosphorylated ERK and IκBα in immature acute lymphoblastic leukemia MOLT-4 T cells and in the naïve mouse spleen. The IκBα·ERK·WWOX complex was shown to localize, in part, in the mitochondria. WWOX prevents IκBα from proteasomal degradation. Upon stimulating MOLT-4 with ionophore A23187/phorbol myristate acetate, endogenous IκBα and ERK undergo rapid phosphorylation in <5 min, and subsequently WWOX is Tyr-33 and Tyr-287 de-phosphorylated and Ser-14 phosphorylated. Three hours later, IκBα starts to degrade, and ERK returns to basal or non-phosphorylation, and this lasts for the next 12 h. Finally, expression of CD3 and CD8 occurs in MOLT-4 along with reappearance of the IκBα·ERK·WWOX complex near 24 h. Inhibition of ERK phosphorylation by U0126 or IκBα degradation by MG132 prevents MOLT-4 maturation. By time-lapse FRET microscopy, IκBα·ERK·WWOX complex exhibits an increased binding strength by 1-2-fold after exposure to ionophore A23187/phorbol myristate acetate for 15-24 h. Meanwhile, a portion of ERK and WWOX relocates to the nucleus, suggesting their role in the induction of CD3 and CD8 expression in MOLT-4.

Topics: Active Transport, Cell Nucleus; Animals; Calcimycin; Cell Nucleus; Extracellular Signal-Regulated MAP Kinases; HEK293 Cells; Humans; Jurkat Cells; Mice; Multiprotein Complexes; NF-KappaB Inhibitor alpha; Oxidoreductases; Phosphorylation; Precursor Cell Lymphoblastic Leukemia-Lymphoma; Protein Domains; Proteolysis; Tetradecanoylphorbol Acetate; Tumor Suppressor Proteins; U937 Cells; WW Domain-Containing Oxidoreductase

An immature human T cell line, PER-117, can be induced to secrete interleukin-2 (IL-2). In contrast to mature T cells or the Jurkat cell line, PER-117 cells require interleukin-1 (IL-1) for optimal IL-2 secretion, in addition to calcium ionophore and phorbol 12-myristate 12-acetate (PMA). These requirements mirror the conditions reported to be optimal for normal immature T cell receptor (TCR) negative thymocytes. IL-1 did not substitute for either of the other signals required for IL-2 production, i.e., calcium ionophore or PMA, suggesting that IL-1 activates pathways different from those elicited by the two other stimuli. For optimal effect, IL-1 needed to be provided at the same time as the two other signals. Significantly, a signal provided by a low concentration of PMA (not leading to IL-2 induction by itself in the presence of calcium ionophore) was necessary. The studies reported here provide the first evidence that three signals are required for optimal IL-2 production in a human immature T cell line. PER-117 cells produce substantial levels of IL-2 and thus provide a model to study stage-specific signal transduction and transcriptional activation of the IL-2 gene in IL-1 responsive immature thymocytes.

Topics: Bone Marrow Cells; Calcimycin; Cell Line; Dose-Response Relationship, Drug; Humans; Infant; Interleukin-1; Interleukin-2; Male; Precursor Cell Lymphoblastic Leukemia-Lymphoma; Signal Transduction; T-Lymphocytes; Tetradecanoylphorbol Acetate