Biomedical subjects

Ulrich Siebenlist

Publications and source records attributed to Ulrich Siebenlist.

14 recordsLinked to original sources

PubMed2006

Improved antibacterial host defense and altered peripheral granulocyte homeostasis in mice lacking the adhesion class G protein receptor CD97.

CD97 is a member of the adhesion family of G protein-coupled receptors. Alternatively spliced forms of CD97 bind integrins alpha5beta1 and alphavbeta3, decay accelerating factor, or dermatan sulfate. CD97 is expressed on myeloid cells at high levels and a variety of other cell types at lower levels. Little is known about the physiological function of CD97. To begin dissecting the function of CD97, we evaluated the immune response of CD97 null mice to systemic infection by Listeria monocytogenes. CD97 null mice were significantly more resistant to listeriosis than matched wild-type mice. A major determinant of the difference in survival appeared to be the comparatively more robust accumulation of granulocytes in the blood and in infected livers of CD97 null mice within 18 h of inoculation, correlating with a decrease in the number of bacteria. CD97 null mice also displayed a mild granulocytosis in the nonchallenged state. Because there is a strong suggestion that CD97 functions in an adhesive capacity, we examined the migratory properties of granulocytes in CD97 null mice. In chimeric animals, CD97 null and wild-type granulocytes migrated similarly, as determined by inflammation-induced emigration from the bone marrow and accumulation in the peritoneum. Granulocyte development in the bone marrow of CD97 null mice was comparable to that of wild-type mice, and CD97 deficiency did not appear to stimulate granulocytosis secondary to peripheral inflammation and resultant granulocyte colony-stimulating factor induction, unlike various other models of adhesion deficiencies. Our results suggest that CD97 plays a role in peripheral granulocyte homeostasis.

Animals↗

PubMed2006

NF-kappaB2 is required for the establishment of central tolerance through an Aire-dependent pathway.

NF-kappaB2-deficient mice have impaired T and B cell responses. We found, however, that in these mice there was severe infiltration of lymphocytes into multiple organs and increased activity of autoantibodies to peripheral tissue antigens in a manner similar to that of autoimmune regulator-deficient (Aire-deficient) mice. We further demonstrated that NF-kappaB2 was required for thymic Aire gene transcriptional regulation. The Nfkb2(-/-) thymus had distinct cortical and medullar structures, but reduced Aire and target gene expression of peripheral tissue antigens. Engraftment of Nfkb2(-/-) thymic stroma to nude mice recapitulated the autoimmune phenotype of the native Nfkb2(-/-) mice, confirming a key defect in central tolerance. Lymphotoxin beta receptor (LTbetaR) ligation-induced Aire gene expression was also largely abolished in the absence of NF-kappaB2. Thus NF-kappaB2 downstream of LTbetaR plays an important role in the regulation of central tolerance in an Aire-dependent manner.

Animals↗

PubMed2006

Transcription impairment and cell migration defects in elongator-depleted cells: implication for familial dysautonomia.

Mutations in IKBKAP, encoding a subunit of Elongator, cause familial dysautonomia (FD), a severe neurodevelopmental disease with complex clinical characteristics. Elongator was previously linked not only with transcriptional elongation and histone acetylation but also with other cellular processes. Here, we used RNA interference (RNAi) and fibroblasts from FD patients to identify Elongator target genes and study the role of Elongator in transcription. Strikingly, whereas Elongator is recruited to both target and nontarget genes, only target genes display histone H3 hypoacetylation and progressively lower RNAPII density through the coding region in FD cells. Interestingly, several target genes encode proteins implicated in cell motility. Indeed, characterization of IKAP/hELP1 RNAi cells, FD fibroblasts, and neuronal cell-derived cells uncovered defects in this cellular function upon Elongator depletion. These results indicate that defects in Elongator function affect transcriptional elongation of several genes and that the ensuing cell motility deficiencies may underlie the neuropathology of FD patients.

Carrier Proteins↗

PubMed2005

Opposing roles for RelB and Bcl-3 in regulation of T-box expressed in T cells, GATA-3, and Th effector differentiation.

CD4+ T cells with a block in the NF-kappaB signaling pathway exhibit decreases in Th1 responses and diminished nuclear levels of multiple transactivating NF-kappaB/Rel/IkappaB proteins. To determine the lineage-intrinsic contributions of these transactivators to Th differentiation, T cells from mice deficient in specific subunits were cultured in exogenous cytokines promoting either Th1 or Th2 differentiation. RelB-deficient cells exhibited dramatic defects in Th1 differentiation and IFN-gamma production, whereas no consistent defect in either Th1 or Th2 responses was observed with c-Rel-deficient cells. In sharp contrast, Bcl-3-null T cells displayed no defect in IFN-gamma production, but their Th2 differentiation and IL-4, IL-5, and IL-13 production were significantly impaired. The absence of RelB led to a dramatic decrease in the expression of T-box expressed in T cells and Stat4. In contrast, Bcl-3-deficient cells exhibited decreased GATA-3, consistent with evidence that Bcl-3 can transactivate a gata3 promoter. These data indicate that Bcl-3 and RelB exert distinct and opposing effects on the expression of subset-determining transcription factors, suggesting that the characteristics of Th cell responses may be regulated by titrating the stoichiometry of transactivating NF-kappaB/Rel/IkappaB complexes in the nuclei of developing helper effector cells.

Animals↗

PubMed2005

Early growth response proteins EGR-4 and EGR-3 interact with immune inflammatory mediators NF-kappaB p50 and p65.

Here, we characterize the basis for the T-cell-specific activity of the human zinc-finger protein early growth response factor 4 (EGR-4). A yeast two-hybrid screen showed interaction of EGR-4 with NF-kappaB p50. Using recombinant proteins, stable physical complex formation was confirmed for EGR-4 and EGR-3 with p50 and with p65 using glutathione-S-transferase pull-down assays and surface-plasmon-resonance and peptide-spot analyses. In vivo interaction of EGR-4 and EGR-3 with NF-kappaB p65 was demonstrated by immunoprecipitation experiments and fluorescence-resonance-energy transfer (FRET) analysis showing interaction in the nucleus of transfected Jurkat T cells. In transfection assays, EGR-p50 complexes were transcriptionally inactive and EGR-p65 complexes strongly activated transcription of the promoters of the human genes encoding the cytokines interleukin 2, tissue necrosis factor alpha and ICAM-1. The EGR-p65 complexes increased reporter-gene activity about 100-fold and thus exceeded the transcriptional activities of the p65 homodimer and the p65/p50 heterodimers. The major interaction domain for p65 was localized within the third zinc finger of EGR-4 using deletion mutants for pull-down assays and peptide-spot assays. By computer modeling, this interaction domain was localized to an alpha-helical region and shown to have the central amino acids surface exposed and thus accessible for interaction. In summary, in T cells, the two zinc-finger proteins EGR-4 and EGR-3 interact with the specific nuclear mediator NF-kappaB and control transcription of genes encoding inflammatory cytokines.

Amino Acid Sequence↗

PubMed2005

Control of lymphocyte development by nuclear factor-kappaB.

The evolutionarily conserved nuclear factor-kappaB family of transcription factors is known to have a crucial role in rapid responses to stress and pathogens, inducing transcription of many genes that are essential for host defence. Now, studies of mice that are deficient in nuclear factor-kappaB-family members (or deficient in the activation of these factors) reveal that nuclear factor-kappaB is extensively involved in the development of T cells and B cells. And, as we review here, although these factors have several roles, their primary cell-autonomous function is to ensure lymphocyte survival at various developmental stages. This function is subverted in numerous diseases and can lead, for example, to survival of self-reactive lymphocytes or tumour cells.

Animals↗

PubMed2004

GSK3-mediated BCL-3 phosphorylation modulates its degradation and its oncogenicity.

The oncoprotein BCL-3 is a nuclear transcription factor that activates NF-kappaB target genes through formation of heterocomplexes with p50 or p52. BCL-3 is phosphorylated in vivo, but specific BCL-3 kinases have not been identified so far. In this report, we show that BCL-3 is a substrate for the protein kinase GSK3 and that GSK3-mediated BCL-3 phosphorylation, which is inhibited by Akt activation, targets its degradation through the proteasome pathway. This phosphorylation modulates its association with HDAC1, -3, and -6 and attenuates its oncogenicity by selectively controlling the expression of a subset of newly identified target genes such as SLPI and Cxcl1. Our results therefore suggest that constitutive BCL-3 phosphorylation by GSK3 regulates BCL-3 turnover and transcriptional activity.

Animals↗

PubMed2004

BCL-3 and NF-kappaB p50 attenuate lipopolysaccharide-induced inflammatory responses in macrophages.

Lipopolysaccharide (LPS) induces expression of tumor necrosis factor alpha (TNFalpha) and other pro-inflammatory cytokines in macrophages. Following its induction, TNFalpha gene transcription is rapidly attenuated, in part due to the accumulation of NF-kappaB p50 homodimers that bind to three kappaB sites in the TNFalpha promoter. Here we have investigated the inhibitory role of BCL-3, an IkappaB-like protein that interacts exclusively with p50 and p52 homodimers. BCL-3 was induced by LPS with delayed kinetics and was associated with p50 in the nucleus. Forced expression of BCL-3 suppressed LPS-induced transcription from the TNFalpha promoter and inhibited two artificial promoters composed of TNFalphakappaB sites that preferentially bind p50 dimers. BCL-3-mediated repression was reversed by trichostatin A and was enhanced by overexpression of HDAC-1, indicating that transcriptional attenuation involves recruitment of histone deacetylase. Analysis of macrophages from p50 and BCL-3 knock-out mice revealed that both transcription factors negatively regulate TNFalpha expression and that BCL-3 inhibits IL-1alpha and IL-1beta. In contrast, induction of the anti-inflammatory cytokine IL-10 was reduced in BCL-3 null macrophages. BCL-3 was not required for the production of p50 homodimers but BCL-3 expression was severely diminished in p50-deficient cells. Together, these findings indicate that p50 and BCL-3 function as anti-inflammatory regulators in macrophages by attenuating transcription of pro-inflammatory cytokines and activating IL-10 expression.

Animals↗

PubMed2003

Lymphotoxin beta receptor induces sequential activation of distinct NF-kappa B factors via separate signaling pathways.

Lymphotoxin beta receptor (LTbetaR)-induced activation of NF-kappaB in mouse embryo fibroblasts was mediated by the classical pathway and by an alternative or second pathway. The classical pathway involved the IkappaB kinase (IKK)beta- and IKKgamma-dependent degradation of IkappaBalpha and resulted in the rapid but transient activation of primarily RelA-containing NF-kappaB dimers. The alternative or second pathway proceeded via NF-kappaB-inducing kinase (NIK)-, IKKalpha-, and protein synthesis-dependent processing of the inhibitory NF-kappaB2 p100 precursor protein to the p52 form and resulted in a delayed but sustained activation of primarily RelB-containing NF-kappaB dimers. This second pathway was independent of the classical IKK complex, which is governed by its central IKKgamma regulatory subunit. The sequential engagement of two distinct pathways, coupled with the negative feedback inhibition of RelA complexes by NF-kappaB-induced resynthesis of IkappaBalpha, resulted in a pronounced temporal change in the nature of the NF-kappaB activity during the course of stimulation. Initially dominant RelA complexes were replaced with time by RelB complexes. Therefore, the alternative activation path mediated by processing of p100 was necessary for sustained NF-kappaB activity in mouse embryo fibroblasts in response to LTbetaR stimulation. Based on the phenotype of mice deficient in various components of the LTbetaR-induced activation of p100 processing, we conclude that this pathway is critically involved in the function of stromal cells during the generation of secondary lymphoid organ microarchitectures.

Animals↗

PubMed2003

Expression of either NF-kappaB p50 or p52 in osteoclast precursors is required for IL-1-induced bone resorption.

Interleukin (IL)-1 is implicated in postmenopausal- and inflammation-mediated bone loss. Its expression is regulated by NF-kappaB and vice versa. To examine the role of NF-kappaB p50 and p52 (they are required for osteoclast formation during embryonic development) in IL-1-induced resorption, we used various NF-kappaB knockout (KO) mice, including p50-/- and p52-/- single KO, p50-/- and p52+/- (3/4KO), and p50-/- and p52-/- double KO (dKO) mice. IL-1 increased blood calcium and bone resorption in wild-type (wt), p50, and p52 single KO mice, but not in 3/4KO or dKO mice. Osteoclast formation was impaired in bone marrow cultures from 3/4KO compared with single KO and wt mice treated with IL-1. IL-1 receptor expression was similar in colony forming unit-granulocyte macrophage (CFU-GM) colony cells from wt and dKO mice. However, IL-1 promoted CFU-GM colony formation and survival as well as the formation, activity, and survival of osteoclasts generated from these colonies from wt mouse splenocytes, but not from dKO splenocytes. No difference in expression of the osteoclast regulatory cytokines, RANKL, and OPG, was observed in osteoblasts from wt and dKO mice. Thus, expression of either NF-kappaB p50 or p52 is required in osteoclasts and their precursors, rather than osteoblasts, for IL-1-mediated bone resorption.

Active Transport, Cell Nucleus↗

PubMed2003

NF-kappaB activating scaffold proteins as signaling molecules and putative therapeutic targets.

Activation of transcription factors such as NF-kappa B occurs through signaling pathways involving sequential phosphorylation of a variety of substrates by distinct kinases. Proper assemby and activation of these kinases require interaction with non-enzymatic and essential partners named scaffold proteins. Here, we describe how the NF-kappa B activating scaffold proteins involved in the signaling pathways triggered by the pro-inflammatory cytokines TNF alpha, IL-1 beta and by the CD40 ligand play such roles. We also illustrate the human genetic diseases that are linked to mutations affecting genes coding for these proteins. We suggest that these scaffold proteins may be specifically targeted by novel therapeutical agents for the treatment of inflammation or cancers.

CD40 Antigens↗

PubMed2002

BAFF-induced NEMO-independent processing of NF-kappa B2 in maturing B cells.

NF-kappa B is usually activated by signal-induced, ubiquitin-mediated degradation of its inhibitor, I kappa B. This process is initiated by phosphorylation of I kappa B by the I kappa B kinase (IKK) complex, predominantly by the IKK beta catalytic subunit, and requires the regulatory subunit IKK gamma (NEMO). Another activation pathway, with no known physiological inducers, involves ubiquitin-mediated processing of the NF-kappa B2 inhibitory protein p100 and is dependent on phosphorylation of p100 by IKK alpha. We show here that B cell-activating factor (BAFF) activates this second pathway and that this requires the BAFF receptor (BAFF-R), the NF-kappa B-inducing kinase (NIK) and protein synthesis, but not NEMO. This NEMO-independent cascade is physiologically relevant for the survival and, hence, progression of maturing splenic B cells.

Animals↗

PubMed2002

Association of the adaptor TANK with the I kappa B kinase (IKK) regulator NEMO connects IKK complexes with IKK epsilon and TBK1 kinases.

Canonical activation of NF-kappa B is mediated via phosphorylation of the inhibitory I kappa B proteins by the I kappa B kinase complex (IKK). IKK is composed of a heterodimer of the catalytic IKK alpha and IKK beta subunits and a presumed regulatory protein termed NEMO (NF-kappa B essential modulator) or IKK gamma. NEMO/IKK gamma is indispensable for activation of the IKKs in response to many signals, but its mechanism of action remains unclear. Here we identify TANK (TRAF family member-associated NF-kappa B activator) as a NEMO/IKK gamma-interacting protein via yeast two-hybrid analyses. This interaction is confirmed in mammalian cells, and the domains required are mapped. TANK was previously shown to assist NF-kappa B activation in a complex with TANK-binding kinase 1 (TBK1) or IKK epsilon, two kinases distantly related to IKK alpha/beta, but the underlying mechanisms remained unknown. Here we show that TBK1 and IKK epsilon synergize with TANK to promote interaction with the IKKs. The TANK binding domain within NEMO/IKK gamma is required for proper functioning of this IKK subunit. These results indicate that TANK can synergize with IKK epsilon or TBK1 to link them to IKK complexes, where the two kinases may modulate aspects of NF-kappa B activation.

Adaptor Proteins, Signal Transducing↗

PubMed2002

NF-kappaB p50 and p52 expression is not required for RANK-expressing osteoclast progenitor formation but is essential for RANK- and cytokine-mediated osteoclastogenesis.

Expression of RANKL by stromal cells and of RANK and both NF-kappaB p50 and p52 by osteoclast precursors is essential for osteoclast formation. To examine further the role of RANKL, RANK, and NF-KB signaling in this process, we used NF-kappaB p50-/- ;p52-/- double knockout (dKO) and wild-type (WT) mice. Osteoclasts formed in cocultures of WT osteoblasts with splenocytes from WT mice but not from dKO mice, a finding unchanged by addition of RANKL and macrophage colony-stimulating factor (M-CSF). NF-kappaB dKO splenocytes formed more colony-forming unit granulocyte macrophage (CFU-GM) colonies than WT cells, but no osteoclasts were formed from dKO CFU-GM colonies. RANKL increased the number of CFU-GM colonies twofold in WT cultures but not in dKO cultures. Fluorescence-activated cell sorting (FACS) analysis of splenocytes from NF-kappaB dKO mice revealed a two-to threefold increase in the percentage of CD11b (Mac-1) and RANK double-positive cells compared with WT controls. Treatment of NF-kappaB dKO splenocytes with interleukin (IL)-1, TNF-alpha, M-CSF, GM-CSF, and IL-6 plus soluble IL-6 receptor did not rescue the osteoclast defect. No increase in apoptosis was observed in cells of the osteoclast lineage in NF-kappaB dKO or p50-/-;p52+/- (3/4KO) mice. Thus, NF-kappaB p50 and p52 expression is not required for formation of RANK-expressing osteoclast progenitors but is essential for RANK-expressing osteoclast precursors to differentiate into TRAP+ osteoclasts in response to RANKL and other osteoclastogenic cytokines.

Animals↗