Signal-specific and phosphorylation-dependent RelB degradation: a potential mechanism of NF-kappa B control
ABSTRACT RelB is an unusual member of the Rel/NF-kappaB family of transcription factors which are involved in oncogenic processes. Due to a relaxed control by the IkappaBs, the cytosolic NF-kappaB inhibitors, RelB is constitutively expressed in the nuclei of lymphoid cells. We show here that RelB is inducibly degraded upon activation of T cells in a fashion similar to the IkappaBs. However, RelB degradation differs from that of IkappaBs since it is not induced by TNFalpha but only by T cell receptor or TPA/ionomycin stimulation. Moreover, RelB degradation occurs in three steps: (i) after stimulation RelB is rapidly phosphorylated at amino acids Thr84 and Ser552 followed by (ii) an N-terminal cut and, finally, (iii) the complete degradation in the proteasomes. Since mutation of the two phosphoacceptor sites to non-acceptor sites abolished RelB phosphorylation in vivo and led to the stabilization of the mutated RelB(DM), site-specific phosphorylation appears to be a necessary prerequisite for RelB degradation. RelB is a crucial regulator of NF-kappaB-dependent gene expression. Thus, the signal-induced degradation of RelB should be an important control mechanism of NF-kappaB activity.
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ABSTRACT: Platelets are small anucleate blood cells derived from megakaryocytes. In addition to their pivotal roles in hemostasis, platelets are the smallest, yet most abundant, immune cells and regulate inflammation, immunity, and disease progression. Although platelets lack DNA, and thus no functional transcriptional activities, they are nonetheless rich sources of RNAs, possess an intact spliceosome, and are thus capable of synthesizing proteins. Previously, it was thought that platelet RNAs and translational machinery were remnants from the megakaryocyte. We now know that the initial description of platelets as "cellular fragments" is an antiquated notion, as mounting evidence suggests otherwise. Therefore, it is reasonable to hypothesize that platelet transcription factors are not vestigial remnants from megakaryocytes, but have important, if only partly understood functions. Proteins play multiple cellular roles to minimize energy expenditure for maximum cellular function; thus, the same can be expected for transcription factors. In fact, numerous transcription factors have non-genomic roles, both in platelets and in nucleated cells. Our lab and others have discovered the presence and non-genomic roles of transcription factors in platelets, such as the nuclear factor kappa β (NFκB) family of proteins and peroxisome proliferator-activated receptor gamma (PPARγ). In addition to numerous roles in regulating platelet activation, functional transcription factors can be transferred to vascular and immune cells through platelet microparticles. This method of transcellular delivery of key immune molecules may be a vital mechanism by which platelet transcription factors regulate inflammation and immunity. At the very least, platelets are an ideal model cell to dissect out the non-genomic roles of transcription factors in nucleated cells. There is abundant evidence to suggest that transcription factors in platelets play key roles in regulating inflammatory and hemostatic functions.Frontiers in Immunology 02/2015; 6:48. DOI:10.3389/fimmu.2015.00048
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ABSTRACT: Heightened inflammation, including expression of COX-2, is associated with COPD pathogenesis. RelB is an NF-κB family member that attenuates COX-2 in response to cigarette smoke by a mechanism that may involve the miRNA miR-146a. There is no information on the expression of RelB in COPD or if RelB prevents COX-2 expression through miR-146a. RelB, Cox-2 and miR-146a levels were evaluated in lung fibroblasts and blood samples derived from non-smokers (Normal) and smokers (At Risk) with and without COPD by qRT-PCR. RelB and COX-2 protein levels were evaluated by western blot. Human lung fibroblasts from Normal subjects and smokers with and without COPD, along with RelB knock-down (siRNA) in Normal cells, were exposed to cigarette smoke extract (CSE) in vitro and COX-2 mRNA/protein and miR-146a levels assessed. Basal expression of RelB mRNA and protein were significantly lower in lung cells derived from smokers with and without COPD, the latter of which expressed more Cox-2 mRNA and protein in response to CSE. Knock-down of RelB in Normal fibroblasts increased Cox-2 mRNA and protein induction by CSE. Basal miR-146a levels were not different between the three groups, and only Normal fibroblasts increased miR-146a expression in response to smoke. There was a positive correlation between systemic RelB and Cox-2 mRNA levels and circulating miR-146a levels were higher only in GOLD stage I subjects. Our data indicate that RelB attenuates COX-2 expression in lung structural cells, such that loss of pulmonary RelB may be an important determinant in the aberrant, heightened inflammation associated with COPD pathogenesis.Respiratory research 05/2015; 16(1):54. DOI:10.1186/s12931-015-0214-6 · 3.38 Impact Factor
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ABSTRACT: Toll-like receptors (TLRs) recognize a wide range of pathogen-associated molecular patterns (PAMP) and are preferentially expressed in innate immune cells. TLR-mediated activation of these cells activates the adaptive immune system. However, it has become clear that TLRs are not only expressed but functionally active in CD4T cells. The intestines are continuously exposed to TLR ligands, including lipopolysaccharide (LPS), a TLR4 ligand, and TLR4 is expressed higher in Th17 cells than Th1 and Th2 cells. In addition, development of Th17 cells in the gut mucosa is more dependent on gut microbiota than Th1, Th2, and Treg. Thus, we examined whether LPS directly regulates Th17 differentiation. LPS directly stimulated Th17 differentiation in vitro. In Th17 cells, LPS increased phosphorylation of NF-κB1, resulting in an increase of p50, the processed form of NF-κB1, whereas it decreased phosphorylation of RelB, leading to the up-regulation of RelB. Subcutaneous injection of LPS increased the frequency of IL-17 producing cells in inguinal lymph nodes, worsening experimental autoimmune encephalomyelitis (EAE). Additionally, expression of TLR1, TLR2, TLR4, and TLR5 was reduced upon T cell activation and LPS showed modest effect on TLR4 expression. These findings provide the first evidence that TLR4 activation directly regulate Th17 differentiation. Copyright © 2015. Published by Elsevier B.V.Immunology letters 03/2015; 54(1). DOI:10.1016/j.imlet.2015.03.003 · 2.37 Impact Factor