Organization and Functional Analysis of the Mouse Transporter Associated with Antigen Processing 2 Promoter

Abstract

In accordance with the key role of MHC class I molecules in the adaptive immune response against viruses, they are expressed by most cells, and their expression can be enhanced by cytokines. The assembly and cell surface expression of class I complexes depend on a continuous peptide supply. The peptides are generated mainly by the proteasome and are transported to the endoplasmic reticulum by a peptide transport pump consisting of two subunits, TAP1 and TAP2. The proteasome low molecular weight polypeptide (2 and 7), as well as TAP (1 and 2) genes, are coordinately regulated and are induced by IFNs. Despite this coordinate regulation, examination of tumors shows that these genes can be discordantly down-regulated. In pursuing a molecular explanation for these observations, we have characterized the mouse TAP2 promoter region and 5′-flanking sequence. We show that the 5′ untranslated regions of TAP2 genes have a characteristic genomic organization that is conserved in both the mouse and the human. The mouse TAP2 promoter belongs to a class of promoters that lack TATA boxes but contain a MED1 (multiple start site element downstream) sequence. Accordingly, transcription is initiated from multiple sites within a 100-nucleotide window. An IFN regulatory factor 1 (IRF1)/IRF2 binding site is located in this region and is involved in both basal and IRF1-induced TAP2 promoter activity. The implication of the extensive differences found among the promoters of class I heavy chain, low molecular weight polypeptide, and TAP genes, all encoding proteins involved in Ag presentation, is discussed.

B Lymphocyte-Induced Maturation Protein (Blimp)-1, IFN Regulatory Factor (IRF)-1, and IRF-2 Can Bind to the Same Regulatory Sites

“…3). The relative affinity of Blimp-1 for each oligonucleotide was calculated from the ratio of the oligonucleotide required to compete half of the bound complex relative to the c-myc oligonucleotide required to compete half of the complex, X 50 /c-myc 50 . Thus, ratios under 1 are sequences for which Blimp-1 has a higher affinity than for the c-myc sequence; ratios greater than 1 are sequences for which Blimp-1 has a lower affinity than for the c-myc sequence.…”

“…Self competition with the c-myc sequence and the nonspecific sequence, C2, is also shown. The affinity of each site was determined by calculating the ratio of the concentration of competitor oligonucleotide required to block 50% of Blimp-1 binding over the concentration of c-myc sequence required to block 50% of Blimp-1 binding (X 50 /c-myc 50 ). Values greater than 1 represent lower affinity binding sites compared with the c-myc gene, and values Ͻ1 represent a higher affinity binding site compared with the c-myc gene.…”

Identification of E2F1 as an Important Transcription Factor for the Regulation of Tapasin Expression

“…These include (i) CREB for constitutive expression and the IFN-sensitive response element for an IFN-␥-induced expression of the HC (40,(43)(44)(45)(46) and the bidirectional TAP1/LMP2 promoter (8,41) and (ii) the cooperating transcription factors IRF2 and STAT1 for constitutive expression as well as IRF1 and STAT1 for the IFN-␥-mediated enhanced TAP1/LMP2 expression (41,42,(47)(48)(49)(50)(51). A similar regulation pattern was observed for murine TAP2, which is controlled by CREB and IRF for constitutive and/or IFN-␥-controlled activity (52,53). Both the human and the murine tapasin promoters contain one IFN-sensitive response element and two IFN␥-activated sequence motif consensus sequences in the 5Ј-UTR of the gene (54,55).…”

Characterization and allelic variation of the transporters associated with antigen processing (TAP) genes in the domestic dog (Canis lupus familiaris)

“…From a minimum of six colonies from each PCR, identical sequences of 631 bp (TAP1) and 516 bp (TAP2) were obtained, indicating a single transcription start site (TSS) for both, which differed from predicted sites. Typical of genes whose promoters lack a TATA box, transcription of TAP1 and TAP2 is usually initiated from multiple sites (Arons et al, 2001; Kishi et al, 1993; Wright et al, 1995). The finding of a single TSS may represent an idiosyncrasy of the DH82 cells, the effect of IFN-γ, or both; for example, Arons et al (2001) found multiple TSSs in murine TAP2 using a T-cell leukemia line, but similarly, observed only one TSS in transcripts from IFN-γ-treated transformed fibroblasts.…”

“…Typical of genes whose promoters lack a TATA box, transcription of TAP1 and TAP2 is usually initiated from multiple sites (Arons et al, 2001; Kishi et al, 1993; Wright et al, 1995). The finding of a single TSS may represent an idiosyncrasy of the DH82 cells, the effect of IFN-γ, or both; for example, Arons et al (2001) found multiple TSSs in murine TAP2 using a T-cell leukemia line, but similarly, observed only one TSS in transcripts from IFN-γ-treated transformed fibroblasts. Multiple start site element downstream (MED)1 sequences (GCTCCC/G) were found in both TAP1 and TAP 2, which are common elements in TATA-less promoters that have multiple initiation sites, so presumably other TSSs are used in canine TAP genes, but 5′-RACE analysis of other canine cell lines will be needed to confirm this supposition.…”