Hyacinth Sterling

MFR, a Putative Receptor Mediating the Fusion of Macrophages

ABSTRACT We had previously identified a macrophage surface protein whose expression is highly induced, transient, and specific, as it is restricted to actively fusing macrophages in vitro and in vivo. This protein is recognized by monoclonal antibodies that block macrophage fusion. We have now purified this protein and cloned its corresponding cDNA. This protein belongs to the superfamily of immunoglobulins and is similar to immune antigen receptors such as the T-cell receptor, B-cell receptor, and viral receptors such as CD4. We have therefore named this protein macrophage fusion receptor (MFR). We show that the extracellular domain of MFR prevents fusion of macrophages in vitro and therefore propose that MFR belongs to the fusion machinery of macrophages. MFR is identical to SHPS-1 and BIT and is a homologue of P84, SIRPα, and MyD-1, all of which have been recently cloned and implicated in cell signaling and cell-cell interaction events.

Regulation of ROMK1 Channels by Protein-tyrosine Kinase and -tyrosine Phosphatase

We have used the two-electrode voltage clamp technique and the patch clamp technique to investigate the regulation of ROMK1 channels by protein-tyrosine phosphatase (PTP) and protein-tyrosine kinase (PTK) in oocytes coexpressing ROMK1 and cSrc. Western blot analysis detected the presence of the endogenous PTP-1D isoform in the oocytes. Addition of phenylarsine oxide (PAO), an inhibitor of PTP, reversibly reduced K+ current by 55% in oocytes coinjected with ROMK1 and cSrc. In contrast, PAO had no significant effect on K+ current in oocytes injected with ROMK1 alone. Moreover, application of herbimycin A, an inhibitor of PTK, increased K+ current by 120% and completely abolished the effect of PAO in oocytes coexpressing ROMK1 and cSrc. The effects of herbimycin A and PAO were absent in oocytes expressing the ROMK1 mutant R1Y337A in which the tyrosine residue at position 337 was mutated to alanine. However, addition of exogenous cSrc had no significant effect on the activity of ROMK1 channels in inside-out patches. Moreover, the effect of PAO was completely abolished by treatment of oocytes with 20% sucrose and 250 μg/ml concanavalin A, agents that inhibit the endocytosis of ROMK1 channels. Furthermore, the effect of herbimycin A is absent in the oocytes pretreated with either colchicine, an inhibitor of microtubules, or taxol, an agent that freezes microtubules. We conclude that PTP and PTK play an important role in regulating ROMK1 channels. Inhibiting PTP increases the internalization of ROMK1 channels, whereas blocking PTK stimulates the insertion of ROMK1 channels.

Superoxide Anions Are Involved in Mediating the Effect of Low K Intake on c-Src Expression and Renal K Secretion in the Cortical Collecting Duct

We previously demonstrated that low K intake stimulated the expression of c-Src and that stimulation of protein tyrosine kinase inhibited ROMK channel activity (Wei, Y., Bloom, P., Lin, D. H., Gu, R. M., and Wang, W. H. (2001) Am. J. Physiol. 281, F206-F212). Decreases in dietary K content significantly increased \batchmode \documentclass[fleqn,10pt,legalpaper]{article} \usepackage{amssymb} \usepackage{amsfonts} \usepackage{amsmath} \pagestyle{empty} \begin{document} \(\mathrm{O}_{{\dot{2}}}^{-}\) \end{document} levels and the phosphorylation of c-Jun, a transcription factor, in renal cortex and outer medulla. The role of \batchmode \documentclass[fleqn,10pt,legalpaper]{article} \usepackage{amssymb} \usepackage{amsfonts} \usepackage{amsmath} \pagestyle{empty} \begin{document} \(\mathrm{O}_{{\dot{2}}}^{-}\) \end{document} and related products such as H2O2 in stimulating the expression of protein tyrosine kinase is suggested by the observation that addition of 50-200 μm H2O2 increased the phosphorylation of c-Jun and the expression of c-Src in M1 cells, a mouse collecting duct principal cell line. The effect of H2O2 on c-Src expression was completely abolished with cyclohexamide or actinomycin D. The treatment of animals on a K-deficient (KD) diet with tempol for 7 days significantly decreased the production of \batchmode \documentclass[fleqn,10pt,legalpaper]{article} \usepackage{amssymb} \usepackage{amsfonts} \usepackage{amsmath} \pagestyle{empty} \begin{document} \(\mathrm{O}_{{\dot{2}}}^{-}\) \end{document}, c-Jun phosphorylation, and c-Src expression. Moreover, low K intake decreased the activity of ROMK-like small conductance channels from 1.37 (control K diet) to 0.5 in the cortical collecting duct and increased the tyrosine phosphorylation of ROMK in the renal cortex and outer medulla. In contrast, the tempol treatment not only increased channel activity to 1.1 in the cortical collecting duct but also decreased the tyrosine phosphorylation of ROMK from rats on a KD diet. Finally, suppressing \batchmode \documentclass[fleqn,10pt,legalpaper]{article} \usepackage{amssymb} \usepackage{amsfonts} \usepackage{amsmath} \pagestyle{empty} \begin{document} \(\mathrm{O}_{{\dot{2}}}^{-}\) \end{document} production with tempol significantly increased renal K excretion measured with metabolic cage and lowered the plasma K concentration in comparison with those on a KD diet alone without tempol. We conclude that \batchmode \documentclass[fleqn,10pt,legalpaper]{article} \usepackage{amssymb} \usepackage{amsfonts} \usepackage{amsmath} \pagestyle{empty} \begin{document} \(\mathrm{O}_{{\dot{2}}}^{-}\) \end{document} and related products play a role in mediating the effect of low K intake on c-Src expression and in suppressing ROMK channel activity and renal K secretion.