A regulatory element in the beta 2-microglobulin promoter identified by in vivo footprinting.

Abstract

Expression of the 12-microglobulin (132-m) and major histocompatibility complex (MHC) class I genes is coordinately regulated. By ligation-mediated polymerase chain reaction, we have analyzed in vivo factor binding to the promoter region of the murine ,12-m gene. In adult spleen, in which ,82-m is expressed, strong protection was found in three elements. Two of these elements, the ,82-m NF-KB binding site and the interferon consensus sequence, are homologous to the regulatory elements of the MHC class I genes and were also found to be protected in spleen. A third protected element, PAM, identified in this work, is unique to the 132-m gene. None of the elements showed protection in brain tissue, in which neither the 132-m nor the MHC class I gene is expressed. In vivo footprinting was also performed with F9 embryonal carcinoma cells, in which expression of the 132-m and MHC class I genes is induced at a low level only upon stimulation with retinoic acid (RA). No in vivo protection was detected before and after RA treatment of F9 cells, indicating that RA induction of 132-m (and MHC class I) expression occurs without detectable in vivo factor occupancy, whereas EL4 T lymphocytes expressing 132-m at a high level exhibited strong protection similar to that in spleen. Despite the lack of in vivo occupancy, the nuclear factors specific for each of the three elements were present in brain tissue and F9 cells as well as in spleen tissue and EL4 cells. We show that PAM, an element identified by its in vivo protection, binds nuclear factors ranging from 40 to 50 kDa in size and is capable of enhancing transcription of a reporter in F9 and other cells. Taken together, these results indicate that in vivo factor occupancy for the ,82-m and MHC class I promoters is coordinated and occurs through a mechanism other than mere expression of relevant factors.

Occupancy of upstream regulatory sites in vivo coincides with major histocompatibility complex class I gene expression in mouse tissues.

“…Consistent with this correlation, we obtained evidence that in vivo protection of the P-2 microglobulin gene also correlates with the expression of the gene (49). However, in vivo protection does not always correlate with expression of the gene.…”

Ligand-dependent occupancy of the retinoic acid receptor beta 2 promoter in vivo.

“…For example, Herrera et al (25) and Dey et al (15) have previously noted that the c-fos promoter is fully occupied prior to growth factor stimulation, with few changes observed following stimulation. Similar constitutive occupancy has been noted for the interferon-stimulated responsive element in several interferon-inducible genes (16,38,43). On the other hand, footprinting of the heat shock element in the human HSP70 gene has been shown to be induced only following heat shock (1).…”

“…Cells were then treated with 0.1% dimethyl sulfate (Kodak) for 2 min at room temperature, and high-molecular-weight DNA was extracted and cleaved with piperidine. Ligation-mediated (LM)-PCR was done with Vent polymerase as described previously (16,20,38). Primers used for analysis of the coding strand were 5'-TGG CAA AGA TAT GAC-3' (+57 to +43), 5'-AGA ATA GAC CCC TCC TGC CTG CCT CGG AGC-3' (+51 to +27), and 5'-ACC CTC CTG CCT CGG AGC TCA CTT CTA-3' (+44 to + 13).…”

NFκB induces overexpression of bovine FcRn

“…The transcriptional regulation of β2 min is similar to that of MHC class I genes and previous reports have indicated that there is a conserved NFκB transcription factor binding site (in addition to interferon regulatory elements) in the promoter of the human and mouse β2 min genes and that is important for the cytokine-induced β2 min expression in lymphoid and myeloid cell types. 48,49 This suggests that the NFκB-mediated induction of the bFcRn α-chain is accompanied with the activation of the mouse β2 min in the APCs and other cytokine-sensitive cells of the bFcRn Tg mice allowing functional overexpression of this interspecies heterodimer receptor. In line with this, our preliminary analysis did not show altered subcellular distribution of the bFcRn in immunized vs. non-immunized spleen sections of Tg mice, although we observed increased bFcRn α-chain staining in different cell types using fluorescence microscopy (data not shown).…”