PubMed Health⌕ Search

Biomedical subjects

W Herr

Publications and source records attributed to W Herr.

At least 91 records · Page 5Linked to original sources
PubMed1991

Functional similarities between human immunodeficiency virus type 1 and simian virus 40 kappa B proto-enhancers.

To search for broadly active enhancer elements within the human immunodeficiency virus type 1 (HIV-1) long terminal repeat, we have used a proto-enhancer amplification assay. In this assay, the enhancer region of simian virus 40 (SV40) is replaced by heterologous regulatory sequences. Upon passage in African green monkey kidney cells. SV40 growth revertants can arise by amplification (usually duplication) of active protoenhancers within the heterologous sequences. Most of the HIV-1 U3 regulatory sequences were assayed; only amplification of one or both of the HIV-1 enhancer core kappa B motifs consistently resulted in viable SV40 virus. Examination of the cell-specific enhancer activity of the individual HIV-1 kappa B proto-enhancers showed that, like the broadly active SV40 kappa B proto-enhancer (C proto-enhancer), they are all active in noninduced cell lines of either lymphoid (H9 and Jurkat) or nonlymphoid (HeLa and CV-1) origin. Unexpectedly, one of three kappa B point mutants that exhibit little or no activity in unstimulated cells is as highly induced in stimulated Jurkat cells as are the wild-type kappa B proto-enhancers. This point mutation shows that kappa B-related proto-enhancers can display markedly different activation properties in unstimulated cells yet still activate transcription to similar levels in stimulated cells.

Animals↗
PubMed1990

Differential transcriptional activation by Oct-1 and Oct-2: interdependent activation domains induce Oct-2 phosphorylation.

The ubiquitous Oct-1 and lymphoid Oct-2 POU homeodomain transcription factors bind to the same DNA sequence but differ in their activation potential. Oct-2 is a positive, negative, or neutral regulator of beta-globin transcription depending on the position and sequence of multimerized binding sites. To activate transcription, Oct-2 relies on two interdependent nonacidic domains, an N-terminal glutamine-rich region and a C-terminal serine-, threonine-, and proline-rich region. Oct-1 also contains a functional glutamine-rich region but fails to activate beta-globin transcription in our assay because the Oct-1 C-terminus is inactive, indicating that differential activation by Oct-1 and Oct-2 is determined by the combination of multiple activation domains. Oct-2 displays a unique phosphorylation pattern that is absent from molecules lacking one or the other activation domain, suggesting the activation domains have a role in inducing protein phosphorylation.

Amino Acid Sequence↗
PubMed1990

The gene for the ubiquitous octamer-binding protein Oct-1 is on human chromosome 1, region cen-q32, and near Ly-22 and Ltw-4 on mouse chromosome 1.

Oct-1 is a sequence-specific transcription and DNA replication factor that recognizes the octameric sequence ATGCAAAT. This protein shares an extended region of sequence similarity, called the POU domain, with the lymphoid-specific transcription factor Oct-2, the pituitary-specific transcription factor Pit-1, and the Caenorhabditis elegans cell lineage gene product unc-86. Two subdomains, POU-related homeobox and POU-specific box, lie within the POU domain. Unlike other POU or homeodomain proteins, Oct-1 is ubiquitously expressed. A ubiquitous 95- to 100-kDa protein that has the same DNA-binding properties and is variously referred to as OTF-1, NFIII, or OBP100 is probably identical to Oct-1. The human gene was mapped by Southern blot analyses of human x rodent hybrid cell lines to chromosome 1, region cen-q32. In Chinese hamster x mouse hybrid cell lines the mouse locus was also mapped to chromosome 1. With the BXD (from progenitor strains C57BL/6J and DBA/2J) set of recombinant inbred strains of mice, the murine gene was localized on the linkage map of mouse chromosome 1. There were no recombinants among 26 strains with Ly-22 and 1 recombinant among 24 strains with Ltw-4. The gene symbol is OTF1 for humans and Oct-1 for mouse.

Animals↗
PubMed1989

The Oct-1 homoeodomain directs formation of a multiprotein-DNA complex with the HSV transactivator VP16.

The herpes simplex virus transactivator VP16 participates in the formation of a multiprotein-DNA complex with the ubiquitous octamer-motif-binding factor Oct-1. Complex formation is dependent on specific amino acids in the Oct-1 homoeodomain which are in positions analogous to positive control mutations in helix 2 of the lambda phage repressor helix-turn-helix motif, indicating that this structure is an ancient target for protein-protein interactions mediating transcriptional control.

Amino Acid Sequence↗
PubMed1988

The POU domain is a bipartite DNA-binding structure.

The POU domain (pronounced 'pow') is a highly charged 155-162-amino-acid (aa) region of sequence similarity contained within three mammalian transcription factors. Pt-1 (ref. 2), Oct-1 (ref. 3) and Oct-2 (ref. 4), and the product of the nematode gene unc-86 (ref. 5) which is involved in determining neural cell lineage. This domain consists of two subdomains, a C-terminal homoeo domain and an N-terminal POU-specific region separated by a short nonconserved linker; the sequence relationship shows that the POU homoeo domains form a distinct POU-related family. In the ubiquitous and lymphoid-specific octamer-motif binding proteins Oct-1 and Oct-2, the POU domain is sufficient for sequence-specific DNA binding. Homoeobox domains contain a helix-turn-helix DNA-binding motif, first identified in bacterial repressors. The helix-turn-helix region of the POU domain is important for DNA binding and, in other classes of homoeo-containing proteins, the entire homoeo domain is sufficient for DNA binding; thus the new POU-specific region could be involved in other functions such as protein-protein interactions. Nevertheless, we show here that in fact the POU domain is a novel bipartite DNA-binding structure in which the POU homoeo and POU-specific regions form two subdomains that are both required for DNA binding but are held together by a flexible linker.

Base Sequence↗
PubMed1988

The SV40 enhancer contains two distinct levels of organization.

The SV40 transcriptional enhancer is composed of separate 15 to 20 base-pair-(bp)-long enhancer elements that cooperate with one another or duplicates of themselves to enhance transcription. These elements are bipartite, being composed of subunits, called enhansons, that can be duplicated or interchanged to create new enhancer elements. Enhansons differ from the enhancer elements because they are very sensitive to changes in spacing. This prototypic enhancer, therefore, contains two distinct levels of organization, each of which requires redundancy to be effective.

Base Sequence↗
PubMed1988

OBP100 binds remarkably degenerate octamer motifs through specific interactions with flanking sequences.

We have used the 100-kD HeLa cell octamer-binding protein OBP100 as a model to study flexible DNA sequence recognition by promoter-binding proteins. OBP100 binds to the conserved octamer motif ATGCAAAT found in numerous promoters and additionally to two degenerate octamer motifs (sites I and II) within the SV40 enhancer region. We show here that OBP100 binds the herpes simplex virus immediate early promoter TAATGARAT (R = purine) motif itself, extending the flexibility of OBP100 sequence recognition to sequences that bear very little resemblance (four matches over a 14-bp region). Nevertheless, a progression of OBP100-binding sites can be established that links the sequences of these two apparently unrelated binding sites by incremental steps. Mutational and chemical modification interference analyses of a degenerate octamer binding site (SV40 site II) show that specific sequences, which are not normally conserved but flank the degenerate octamer motif, can compensate for the degeneracy in the octamer core sequence. Thus, different regions of the binding site sequence (core or flanking) can diverge separately but not independently of one another. These results suggest that flexible DNA sequence recognition arises because there are few obligatory contact sites for OBP100 binding, but, rather, specific binding reflects the sum of many independent interactions.

Animals↗
PubMed1988

The ubiquitous octamer-binding protein Oct-1 contains a POU domain with a homeo box subdomain.

The octamer motif ATGCAAAT is recognized indistinguishably by two mammalian transcription factors: one that is expressed ubiquitously and referred to here as Oct-1, and another, Oct-2, that is expressed in lymphoid cells. We report the cDNA cloning of the human oct-1 gene, which encodes Oct-1, by screening lambda gt11 recombinant phage in situ for octamer motif-specific DNA binding. One lambda gt11 recombinant expressed a beta-galactosidase-octamer-binding fusion protein with a DNA-binding specificity indistinguishable from human HeLa cell Oct-1 protein. As expected for a ubiquitously expressed protein, Oct-1 mRNA is expressed in all five human and two mouse cell lines tested. Polyclonal rabbit antiserum raised against the beta-galactosidase fusion protein shows that the DNA-binding domains of Oct-1 and Oct-2 proteins are related antigenically. Deletion analysis of the 743-amino-acid-long oct-1 open reading frame shows that the DNA-binding activity lies within a central highly charged domain of 160 amino acids. Comparison of the Oct-1 and Oct-2 sequences reveals that this domain is nearly identical between the two proteins. Highly similar domains are also present in the pituitary-specific transcription factor Pit-1 and the Caenorhabditis elegans unc-86 cell lineage gene product (see Herr et al. 1988). Within this shared POU (Pit-1, Oct-1 and Oct-2, unc-86) domain (pronounced 'pow') lie two subdomains: a POU-related homeo box and a POU-specific box. The Oct-1 protein is unique among the POU-related proteins and other homeo box proteins because it is expressed ubiquitously.

Amino Acid Sequence↗
PubMed1988

Activation of the U2 snRNA promoter by the octamer motif defines a new class of RNA polymerase II enhancer elements.

The recent discovery that the activation domains of transcriptional activators (e.g., GAL4) from a number of species are interchangeable has led to the concept of a general mechanism for activation of RNA polymerase II genes. We have examined the different activities of the SV40 octamer motif ATGCAAAG in B cells and in HeLa cells in the context of either the beta-globin promoter, a TATA-box-containing mRNA promoter, or the U2 snRNA promoter, which contains a snRNA-specific proximal element. In the context of the beta-globin promoter, the octamer motif is a B-cell-specific enhancer element, whereas it is a ubiquitous enhancer element for the U2 snRNA promoter. The U2 promoter is unique in that it is not activated by enhancer elements that activate the beta-globin promoter, and a hybrid U2 promoter containing the upstream activating sequence UASG is not stimulated by a yeast GAL4 trans-activator. Together, these observations suggest that in the context of the U2 promoter, the octamer motif defines a new class of RNA polymerase II enhancer elements, which bind transcription factors that trans-activate gene expression by a different mechanism than the general mechanism mentioned above. These results are discussed in light of the possibility that the ubiquitous octamer binding protein Oct-1 and the B-cell-specific octamer binding protein Oct-2 are involved in the activation of the U2 and beta-globin promoters, respectively.

B-Lymphocytes↗
PubMed1988

Simian virus 40 revertant enhancers exhibit restricted host ranges for enhancer function.

We have assayed the cell-specific activity of a matched set of four enhancers found in viral revertants derived from simian virus 40 (SV40) enhancer mutants. These enhancers all contain 71-base-pair duplications that span identical regions or, in one case, the same region shifted by 2 nucleotides. The four enhancers differ, however, in that each one either carries a different wild-type pair of the genetically defined SV40 enhancer A, B, or C elements, with the other two elements mutated, or carries all three elements mutated. The three enhancers carrying two copies of a wild-type element effectively enhance transcription in CV-1 and HeLa cells, but only the enhancer containing a duplicated wild-type C element exhibits activity in NIH 3T3 cells. These results show that the ability of the A, B, and C elements to compensate for one another is cell specific and that selection for enhancer function in one cell type can generate enhancers with different cell-specific activities. These results are consistent with the hypothesis that tandem duplication of multiple distinct enhancer elements, as in wild-type strains of SV40 (e.g., the 72-base-pair repeat), has the property of expanding the host range of an enhancer.

Animals↗
PubMed1987

Discrete elements within the SV40 enhancer region display different cell-specific enhancer activities.

The SV40 enhancer contains three genetically defined elements, called A, B and C, that can functionally compensate for one another. By using short, synthetic DNA oligonucleotides, we show that each of these elements can act autonomously as an enhancer when present as multiple tandem copies. Analysis of a progressive series of B element oligomers shows a single element is ineffective as an enhancer and that the activity of two or more elements increases with copy number. Assay in five different cell lines of two separate enhancers containing six tandem copies of either the B or C element shows that these elements possess different cell-specific activities. Parallel oligomer enhancer constructs containing closely spaced double point mutations display no enhancer activity in any of the cell lines tested, indicating that these elements represent single units of enhancer function. These elements contain either a 'core' or 'octamer' consensus sequence but these consensus sequences alone are not sufficient for enhancer activity. The different cell-specific activities of the B and C elements are consistent with functional interactions with different trans-acting factors. We discuss how tandem duplication of such dissimilar elements, as in the wild-type SV40 72-bp repeats, can serve to expand the conditions under which an enhancer can function.

Animals↗
PubMed1987

A 100-kD HeLa cell octamer binding protein (OBP100) interacts differently with two separate octamer-related sequences within the SV40 enhancer.

Numerous eukaryotic upstream promoter and enhancer regions contain a functional octamer sequence ATGCAAAT. We have examined the interactions between an octamer binding protein isolated from HeLa cells and the SV40 and immunoglobulin heavy-chain (IgH) gene enhancers. A partially purified octamer binding activity forms a single complex with the IgH enhancer octamer in a gel retardation assay, but two complexes with a SV40 enhancer fragment containing a single 72-bp element. By using point mutants and both dimethyl sulfate and diethyl pyrocarbonate modification interference assays, we show that the SV40 complexes result from binding of a factor to the octamer-related sequence ATGCAAAG (Octa1) and to an adjacent previously unidentified octamer-related sequence ATGCATCT (Octa2). The base-specific interactions with Octa1 and Octa2 differ; chemical modifications over a 10-bp sequence TATGCAAAGC affect Octa1 binding whereas Octa2 binding is affected by modifications spanning a 13-bp sequence ATGCATCTCAATT in which the octamer-like sequence is not centered. The octamer binding activity has been purified extensively by a DNA affinity precipitation procedure and SDS-polyacrylamide gel electrophoresis. The purified protein, OBP100, has an apparent molecular weight of 100 kD and binds both SV40 Octa1 and Octa2, as well as the IgH enhancer. The distinct interactions of OBP100 with the differently sized Octa1 and Octa2 binding sites suggest remarkably flexible sequence recognition between OBP100 and its binding sites.

Base Sequence↗
PubMed1987

Activation of mutated simian virus 40 enhancers by amplification of wild-type enhancer elements.

We show that duplication of any one of three separate simian virus 40 enhancer elements, A, B, or C, can compensate for loss of function in the remaining two. Simian virus 40 revertants containing point mutations within the A and C (dpm16) or B and C (dpm26) enhancer elements contain tandem duplications that include the remaining wild-type element. These simple tandem duplications can create enhancers 25-fold more active than that of the parental mutant. These revertants can arise by illegitimate recombination between heterologous viral genomes. This was demonstrated by the recombinants resulting from a mixed infection with the viruses dpm16 and dpm2, which contain mutations in the A and C elements and the B element, respectively.

Base Sequence↗
PubMed1986

The SV40 enhancer is composed of multiple functional elements that can compensate for one another.

We present evidence that the SV40 enhancer consists of three functional units, A, B, and C, each of which can cooperate with the others or with duplicates of itself to enhance transcription. We show that, when element C, containing the core consensus sequence, is inactivated by point mutations, revertants with restored enhancer function contain duplications of either one or both of the elements A and B. To search for additional elements, we isolated revertants of a mutant with the three elements mutated. These revertants do not identify any other elements; instead, enhancer function is effectively restored by "double duplications," in which the first duplication event either partially or entirely recreates one of the three elements A, B, and C and the second duplication then creates two copies of the newly created sequence.

Base Sequence↗
PubMed1985

Diethyl pyrocarbonate: a chemical probe for secondary structure in negatively supercoiled DNA.

Purine residues located within regions of DNA that have the potential to form left-handed Z-helical structures are modified preferentially by diethyl pyrocarbonate; this hyperreactivity is dependent on the degree of negative superhelicity of the circular DNA molecules. As negative superhelical density increases, guanosines in a 32-base-pair alternating G-C sequence and adenosines (but not guanosines) in a 64-base-pair alternating A-C/G-T sequence become 5- to 10-fold more reactive to diethyl pyrocarbonate. The negative superhelical densities at which enhanced reactivity occurs are similar to those reported for the point at which left-handed helices form within plasmids carrying these DNA sequences. Probing of negatively supercoiled pBR322 with diethyl pyrocarbonate reveals a hyperreactive region 31 base pairs in length of which only 9 base pairs are a perfect alternating purine and pyrimidine sequence; the reactivity of purines within this sequence indicates that purines in the anti conformation, or guanosines in the syn conformation with neighboring 3' thymidines, are not hyperreactive in the Z-DNA form.

Adenosine↗
PubMed1984

Nucleotide sequence of AKV murine leukemia virus.

AKV is an endogenous, ecotropic murine leukemia virus that serves as one of the parents of the recombinant; oncogenic mink cell focus-forming viruses that arise in preleukemic AKR mice. I report the 8,374-nucleotide-long sequence of AKV, as determined from the infectious molecular clone AKR-623. The 5'-leader sequence of AKV extends to nucleotide 639, after which lies a long open reading frame encoding the gag and pol gene products. The reading frame is interrupted by a single amber codon separating the gag and pol genes. The pol gene overlaps the env gene within the 3' region of the AKV genome. The nucleotide sequence of the 5' region of AKV reveals the following features. (i) The 5'-leader sequence lacks any AUG codon to initiate translation of gPr80gag, suggesting that gPr80gag is not required for the replication of AKV. (ii) A short portion of the leader region diverges in sequence from the closely related Moloney murine leukemia virus and appears to be related to a sequence highly repeated in eucaryotic genomes. (iii) As in Moloney murine leukemia virus, there is a potential RNA secondary structure flanking the amber codon that separates the gag and pol genes. This structure might function as a regulatory protein binding site that controls the relative levels of synthesis of the gag and pol precursors. The nucleotide sequence of the 3' region of AKV is compared with sequences reported previously from both infectious and noninfectious molecular clones of AKV.

Amino Acid Sequence↗