The Ezrin-Radixin-Moesin (ERM) proteins regulate B lymphocyte activation via their effect on BCR diffusion and microclustering. This relies on their ability to dynamically tether the plasma membrane with actin filaments that is in turn facilitated by phosphorylation of the conserved threonine residue in the actin-binding domain. Here, we describe a novel function of ezrin in regulating JNK activation that is mediated by phosphorylation of a tyrosine (Y353) residue that is unconserved with moesin and radixin. BCR, but not CD40, TLR4 or CXCR5 stimulation, induced phosphorylation of ezrin at Y353 in mouse splenic B cells. Ezrin existed in a preformed complex with Syk in unstimulated B cells and underwent Syk-dependent phosphorylation upon anti-IgM stimulation. Y353-phosphorylated ezrin co-localized with the BCR within minutes of stimulation and co-trafficked with the endocytosed BCRs through the early and late endosomes. The T567 residue of ezrin was rephosphorylated in late endosomes and at the plasma membrane at later times of BCR stimulation. Expression of a non-phosphorylatable Y353F mutant of ezrin specifically impaired JNK activation. BCR crosslinking induced the association of Y353-phosphorylated ezrin with JNK and its kinase MKK7, and spatial co-localization with phosphorylated JNK in the endosomes. The YFP-tagged Y353F mutant displayed reduced co-localization with the endocytosed BCR as compared to wild type Ezrin-YFP. Taken together, our data identify a novel role for ezrin as a spatial adaptor that couples JNK signaling components to the BCR signalosome, thus facilitating JNK activation.

SUMMARYIgA nephropathy (IgAN) is defined by the predominant deposition of IgA immune complexes (IC) in the glomerular mesangium. Interaction between IgA immune complexes and mesangial cells (MC) could be a linchpin for the genesis of IgAN. We studied the modulation of MC expression of IgA receptors (FcaR) by selected cytokines. Binding of 125 I-IgA to quiescent human MC showed 2 : 55 × 10 5 sites/cell with an affinity (Ka) of 3 : 2 × 10 7 M -1 . Addition of selected recombinant cytokines had no significant influence on Ka, but increased the number of sites/cell relative to unstimulated cells. Northern hybridization using the pHuFcaR cDNA probe showed time-dependent increases in mRNA expression in stimulated versus control cells. IL-6 and tumour necrosis factor-alpha (TNF-a) had a biphasic effect on the FcaR mRNA level; at 48 h, IL-6 increased steady state mRNA levels about six-fold relative to control, TNF-a increased mRNA four-fold, and interferon-gamma (IFN-g) induced FcaR mRNA twofold. By reverse transcriptase-polymerase chain reaction (RT-PCR), the FcaR expressed on human MC appears highly homologous to that expressed by U937 cells. Altered FcaR expression in response to cytokines may influence the pathogenesis of IgAN by affecting deposition and/or clearance of IgA-IC in the mesangium.

CsA induces apoptosis in various renal cell lines, and this effect is mediated by the induction of iNOS via p53. These effects may contribute to the acellular fibrosis characteristic of late CsA nephrotoxicity.

We found that Amadori-configurated GA at furosine concentrations similar to those in diabetic patients favoured EC apoptosis through enhancement of iNOS activity. We propose that this process may be involved in the progressive cellular loss occurring in vascular and glomerular diabetic sclerosis.

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