Different regulatory pathways employed in cytokine-enhanced expression of secretory component and epithelial HLA class I genes

Abstract

The transmembrane secretory component (SC, or pIg receptor) plays a crucial role in mucosal immunity by translocating dimeric IgA and pentameric IgM through exocrine epithelia. This receptor is up-regulated by cytokines in parallel with increased epithelial HLA expression. By use of the human epithelial cell line HT-29m3, we show that IFN-+ , TNF- § and IL-4 activate transcription of the SC gene. This activation was slow, suggesting mediation via newly synthesized protein factors. IFN-+ and TNF- § , but not IL-4, also up-regulated expression of HLA class I genes. However, this gene induction was rapid and did not depend on new protein synthesis. Nuclear run-on experiments showed that the transcription rate of HLA class I genes nearly peaked after only 30 min of IFN-+ or TNF- § stimulation, whereas the SC transcription rate did not peak until after 20-36 h of IFN-+ , TNF- § or IL-4 stimulation. Gel electrophoresis mobility shift assays demonstrated binding of nuclear proteins from cytokine-stimulated HT-29 cells to consensus elements in the promoter of the SC gene, involving the binding site for the nuclear factor-O B p50 subunit after TNF- § stimulation, and IFN-stimulated response element after IFN-+ stimulation (and weakly after TNF- § ). Our observations in vitro likely parallel events in vivo by which activated mucosal T cells and macrophages enhance pIg receptor-mediated external transport of secretory IgA and IgM and up-regulate epithelial HLA expression.

De Novo Synthesized RelB Mediates TNF-Induced Up-Regulation of the Human Polymeric Ig Receptor

“…The magnitude of mRNA up-regulation by TNF varied somewhat between experiments, but the delayed response (after 4 h) and the timing of the steady state plateau were reproducible. Importantly, the results were in accordance with our previous report (10). The kinetics of TNF-mediated up-regulation of the pIgR-derived reporter gene pSC1 was investigated by transient transfection in HT-29.m3 cells and stimulation with TNF for various time periods as described in Materials and Methods (Fig.…”

“…The proinflammatory cytokine TNF is a key mediator of host protection against infection (8,9) and is known to up-regulate the transcription of pIgR by a delayed and protein synthesis-dependent mechanism (10). The best-characterized mediator of TNF action is the NF-B family of latent, rapidly activated transcription factors (TFs) (11,12).…”

A Novel NF-κB/Rel Site in Intron 1 Cooperates with Proximal Promoter Elements to Mediate TNF-α-Induced Transcription of the Human Polymeric Ig Receptor

“…TNF-␣-mediated transcriptional up-regulation of pIgR/SC was shown to depend on NF-B/Rel activation (25) and an IFN-stimulated response element (ISRE) located in exon 1 (26,27). However, the effect mediated by described DNA elements could not account for the degree of up-regulation indicated by the increased level of mRNA and by nuclear run-on experiments (18). In this report, we describe a novel 748-bp TNF-␣-responsive region in the 5.7-kb intron 1, located ϳ4.1 kb downstream of the transcriptional start site.…”

“…For mutation of the exon 1 ISRE and the NF-B/Rel site in intron 1 (pSC45-pSC47, pSC51-pSC54, pSC56 -pSC58), point mutations were designed that changed the nucleotide from a purine to the noncomplementary pyrimidine and vice versa (i.e., A7C, T7G). For mutation of the NF-B/Rel site in the pIgR/SC promoter (pSC55-pSC58), the same mutation was introduced (GGG3 CCC) as used for EMSA by Nilsen et al (18). The integrity of the vector-insert boundary of all subcloned DNA fragments, as well as all mutations, was confirmed by sequencing with the cycle sequencing kit (Amersham International, Slough, U.K.), or by the sequencing service offered by MediGenomix (Martinsried, Germany).…”

“…Transfected cells were either left untreated or stimulated with 10 ng/ml recombinant human TNF-␣ for 12 or 24 h (as indicated in the figures). A time point of 24 h was initially chosen based on earlier experiments with the endogenous pIgR/SC gene (18). However, to minimize the effect of TNF-␣ on the internal control plasmid (pRL-PGK), we later chose 12 h for cytokine incubation, which yielded similar levels of pIgR/SC gene induction as 24 h. The luciferase activity of both the reporter gene (Firefly luciferase) and the internal control plasmid pRL-PGK (Renilla luciferase) was measured in a luminometer (Victor; Wallace, Turku, Finland) with the Dual Luciferase Reporter Assay System (Promega).…”