Upstream activating sequence

About: Upstream activating sequence is a research topic. Over the lifetime, 1633 publications have been published within this topic receiving 100112 citations.

Sequences Just Upstream of the Simian Immunodeficiency Virus Core Enhancer Allow Efficient Replication in the Absence of NF-κB and Sp1 Binding Elements

Abstract: The proviral genomes of all retroviruses are flanked by repetitive sequences called long terminal repeats (LTRs) (38). Each LTR consists of three regions, U3, R, and U5 (38). The U3 region of the primate lentiviruses contains (i) sequences required for integration into the host cell genome and (ii) major transcriptional control elements: the TATA box motif and binding sites for the transcription factors NF-κB and Sp1 (9, 10, 12, 17, 18, 28, 34, 36). The U3 regions of human immunodeficiency virus type 1 (HIV-1) and simian immunodeficiency virus (SIV) are considerably longer than those of other lentiviruses, ranging from about 450 to 560 bp in length (27). About 330 to 400 bp of the sequences upstream of the NF-κB and Sp1 binding sites overlap the nef open reading frame. After infection of rhesus macaques with a nef-defective SIV variant, large deletions accumulated within a 334-bp region of U3 that overlaps the nef open reading frame (22) (Fig. ​(Fig.1).1). Further analysis confirmed that this region serves predominantly as the Nef coding region (13). Similar deletions were observed in infection with nef-defective HIV-1 (4, 23) (Fig. ​(Fig.1).1). In one of these long-term nonprogressors of HIV-1 infection, the predominant deletion observed early in infection was located in the nef-unique region, and long deletions in U3 accumulated over time (23). These results suggest that in the absence of an intact nef gene, these upstream U3 sequences (US sequences) are lost and may not be advantageous for SIVmac and HIV-1 replication in vivo. FIG. 1 Locations of deletions in the nef-unique and U3 regions observed in HIV-1 and SIV infection. (A) Deletions observed in a long-term nonprogressor of HIV-1 infection at early and late time points (23). (B) Additional deletions observed in the U3 region ... These naturally occurring deletions (NOD) removed up to 65% of the HIV-1 and SIVmac U3 regions and almost the entire nef-unique region. However, some elements of well-documented importance, like the polypurine tract, the NF-κB and Sp1 binding sites, the TATA box motif, and sequences required for proviral integration, were not affected (Fig. ​(Fig.1)1) (22, 23). In both HIV-1 and SIVmac U3, also 60 to 90 bp just upstream of the NF-κB binding site(s) were always preserved (22, 23). For HIV-1, it has been documented that the cell-type-specific cellular transcription factors USF, LEF-1, and Ets-1 bind to this region (Fig. ​(Fig.1)1) and activate transcription synergistically with Sp1 and NF-κB on chromatin-assembled DNA (1, 6, 20, 30, 35). Furthermore, point mutations in these binding sites decreased viral replication (21). Relatively little is known about the relevance of the sequences just upstream of the single NF-κB binding site in the SIVmac LTR for viral transcription and replication. Several lines of evidence suggest the presence of an important enhancer element in this region. For instance, some transcription factors that bind just upstream of the NF-κB binding site in the LTRs of SIVmac (33, 40) and the closely related HIV-2 have been described (24–26). Most strikingly, SIVmac239 containing deletions of all NF-κB and Sp1 binding sites replicated with kinetics similar to those of the wild-type SIVmac239 clone in lymphoid cells and caused AIDS in rhesus macaques (14, 15). In contrast, HIV-1 containing deletions or substitutions in all NF-κB and Sp1 elements is not competent for replication (34). In this study, we have analyzed the influence of deletions and point mutations in the region upstream of the core enhancer region. Since 334 bp of US sequences are clearly dispensable for SIV LTR function in vivo (22), we focused on the 65 bp just upstream of the NF-κB binding site, designated the US65 region (Fig. ​(Fig.2).2). Deletion of this region had only a moderately negative effect on transcriptional activity in transient assays and did not reduce viral replication in several cell lines, including primary rhesus macaque peripheral blood mononuclear cells (rPBMC), when either the single NF-κB binding site or the four Sp1 binding sites were present. Therefore, we analyzed SIVmac239 LTR variants containing deletions in the US sequences in conjunction with deletion of the entire core enhancer. We report that a purine-rich site (purine-rich box 2 [PuB2]) in the 26 bp of sequences located just upstream of the NF-κB binding site binds to the transcription factor Elf-1 and that this regulatory element allows efficient viral replication in the absence of the entire core enhancer region. FIG. 2 Mutations in the SIVmac239 U3 region. The SIVmac239 sequence has been published by Regier and Desrosiers (32). The NF-κB and Sp1 binding sites, the stop codon of nef, two PuB sites, a region with homology to the USF binding site, the peri-κB ...

Two upstream elements activate transcription of a major histocompatibility complex class I gene in vitro

Abstract: Expression of major histocompatibility complex (MHC) class I genes exhibits unique tissue and developmental specificity In an effort to study molecular mechanisms of MHC class I gene regulation, an in vitro transcription system has been established In B cell nuclear extracts a template DNA containing the mouse H-2Ld promoter sequence accurately directed RNA polymerase II-dependent transcription of a G-free cassette A conserved class I regulatory complex previously shown to moderately enhance promoter activity in vivo enhanced transcription in vitro by 2-3 fold Much of this enhancement was accounted for by a 40 bp fragment within the complex, which was capable of activating a basal H-2Ld promoter in either orientation Farther downstream, another element called site B was identified, which independently activated MHC class I transcription in vitro by 2-4 fold Site B bound a specific nuclear factor(s) through an NF-1 binding site but not through a neighboring CCAAT site The functional significance of site B in vivo was demonstrated in transfection experiments in which site B enhanced MHC class I promoter activity to a degree comparable to that seen in vitro With the identification of the two upstream activators, MHC class I genes may serve as a model to study roles of sequence-specific DNA-binding proteins in transcription in vitro

A unique, short sequence determines p53 gene basal and UV-inducible expression in normal human cells.

Abstract: A unique, short sequence determines p53 gene basal and UV-inducible expression in normal human cells