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I Boros

Publications and source records attributed to I Boros.

At least 37 records · Page 2Linked to original sources
PubMed1994

Expansion of CREB's DNA recognition specificity by Tax results from interaction with Ala-Ala-Arg at positions 282-284 near the conserved DNA-binding domain of CREB.

The transactivator protein of human T-lymphotropic virus type I (HTLV-I), Tax, forms multiprotein complexes with the ubiquitous transcription factor CREB and the CREB/ATF-1 heterodimer. The interaction between Tax and CREB is highly specific and results in increased binding of the Tax/CREB complexes to the HTLV-I 21-bp repeats. Despite the extensive sequence similarities between CREB and ATF-1, Tax interacts with ATF-1 only marginally. Compared with CREB, Tax/CREB exhibits greatly increased DNA recognition specificity and preferentially assembles on a consensus binding site, GGGGG(T/A)TGACG(T/C)(A/C)TA(T/C)C-CCCC, homologous to the HTLV-I 21-bp repeats. Here we report that Tax affects CREB binding to the Tax-inducible DNA elements by interacting with the basic-leucine zipper (bZip) domain of CREB. We show by domain switching that the basic region in CREB bZip can confer on c-Jun and ATF-1 leucine zippers the ability to interact with Tax in vitro. Mutational analyses further demonstrate that the amino acid residues of CREB critical for Tax/CREB interaction are Ala-Ala-Arg at positions 282-284 (AAR284), immediately upstream of the highly conserved DNA-binding domain (R/K)XX(R/K) N(R/K)XAAXX(S/C)RX(R/K)(K/R) characteristic of all bZip proteins. Specific amino acid substitutions in AAR284 of CREB weakened or abolished Tax/CREB interaction, whereas reciprocal changes in ATF-1 allowed it to interact with Tax. These results support a model in which the specific interaction between Tax and the AAR284 residues near the DNA-binding domain of CREB results in a multiprotein complex with altered DNA recognition property. This protein complex assembles selectively on the viral Tax-responsive 21-bp repeats to augment transcription.

Activating Transcription Factor 1↗
PubMed1994

Renal hypertrophy in streptozotocin-diabetic rats.

In male adolescent rats kidney mass attained maximum terminal value already at week 1 after streptozotocin treatment, and this value did not change over the next two weeks. At week 1 the kidney size of diabetic rats was greater than that of untreated controls, whereas at week 3 renal mass was the same in both groups. In adult rats with final body and kidney mass streptozotocin treatment failed to induce further increment in renal growth, and the kidney size of diabetic animals was the same as the final kidney size of age-matched control animals. The identity of terminal maximum values for kidney mass in male rats suggests rather an accelerated kidney growth than a true hypertrophic response in diabetes. Similar conclusion can be drawn from our observations made on the compensatory growth of the solitary kidney in diabetic rats with prior uninephrectomy.

Animals↗
PubMed1993

Member of the CREB/ATF protein family, but not CREB alpha plays an active role in BLV tax trans activation in vivo.

The trans activator protein of Bovine Leukaemia Virus (tax) increases the rate of transcription from the virus promoter through 21 bp sequences located in three tandem copies in the virus LTR. Based on data obtained by three different experimental approaches we concluded that the central CRE-like motif found in each of the BLV 21 bp repeats plays an important and indispensable role in tax mediated trans activation. These include (i) in vivo analysis of the function of mutant 21 bp sequences in transient transfection, (ii) gel mobility shift assay to show that CREB binds to BLV 21 bp repeats in vitro and (iii) the demonstration that the production of antisense CREB mRNA inhibits tax trans activation. Further studies with different deletion mutant CREB proteins suggest that although CREB alpha can interact with factors involved in BLV trans activation, it does not promote transcription initiation; consequently some other member/s of the CREB/ATF family must be involved.

Activating Transcription Factors↗
PubMed1993

Expression in Escherichia coli and in vitro processing of HIV-1 p24 fusion protein.

Recombinant HIV-1 p24/p25 gag proteins were obtained from Escherichia coli using a cleavable fusion strategy. The fusion protein contains 280 amino acid residues of staphylococcal Protein A and 317 amino acid residues of p24/p25 flanking with the recognition/cleavage sequences for HIV protease. Fusion protein expressed under the control of lambda phage promoter PR was purified by IgG-Sepharose affinity chromatography. The p24/p25 part of the fusion protein was released by recombinant HIV protease in vitro. After a second IgG-Sepharose affinity chromatography, the purified p24/p25 proteins were obtained in milligram quantities. The HIV-1 p24/p25 protein displayed antigenicity similar to those of native counterparts confirmed by Western blot assays and the Abbott antigen test.

Blotting, Western↗
PubMed1993

[The effect of H2-receptor blockers (cimetidine and ranitidin) on parotid gland secretion and salivary carbonic anhydrase activity in the rat].

It has been demonstrated, that the H2-receptor antagonists: cimetidine and ranitidine-administered in doses of the same order of magnitude as reported for the inhibition of rat gastric secretion and development of the experimentally induced gastric and duodenal ulcer-are capable of increasing significantly the parotid secretion evoked by the cholinergic stimulant bethanechol. The salivary carbonic anhydrase activity was also elevated after combined treatment of bethanechol(+)-cimetidine. The similar action of the two H2-antagonists appears to suggest that their effect on salivation is connected probably with the cholinergic activity and it is concluded that the combination of bethanechol with these blockers may improve the esophageal clearance, an action that may be beneficial in the treatment of the gastroesophageal reflux disease.

Animals↗
PubMed1991

Mutational analysis of the HTLV-I trans-activator, Tax.

The gene coding for the trans-activating factor (Tax) of the human T-cell leukemia virus, type I (HTLV-I) was mutagenized in vitro using oligonucleotide-directed mutagenesis and recombinant DNA techniques. All except one of the mutagenized tax constructs failed to trans-activate the HTLV-I LTR in a eukaryotic test system. Moreover, negative Tax mutant Arg-39----Gly was found to be trans-dominant. This observation suggests that Tax contains distinct functional domains mediating different interactions of the protein in the process of trans-activation.

Base Sequence↗
PubMed1991

Analysis of the complex transcription termination region of the Escherichia coli rrnB gene.

The complex terminator region of the Escherichia coli rrnB gene was analyzed by subcloning the terminators T1 and T2 and the inverted repeats IR1 and IR2 individually, or in various combinations, in a normal or inverted orientation into a terminator probe vector. The in vivo terminating efficiency was assayed by measuring the galactokinase activity encoded by the downstream galK gene. Termination efficiencies of all fragments were compared in two constructs, differing in the presence or absence of readthrough translation over the investigated terminator signal. The following main conclusions were drawn. (a) T1 and T2 are both efficient terminators in isolated forms. (b) IR1 and IR2 have some terminating effect (much lower than the proper terminators), especially in the inverted orientation. Their presence modifies the effect of the proper terminators in a quite unpredictable way, especially if these regions are translated. (c) The terminators are not symmetrical; in the inverted orientation T1 is practically inactive and T2 termination is reduced. (d) Translation radically decreases the efficiency of the terminators. (e) Several sequences in the rrnB gene, upstream of the terminator region (one in the 16S RNA and one in the 5S RNA coding region), are very efficient in vivo terminators in the inverted orientation.

Base Sequence↗
PubMed1990

[Chromatographic study of salivary carbonic anhydrase isoenzymes in rats].

Authors could purify two isoenzymes (designated CA1 and CA2) from the submandibular gland but only one (CA1) from the submandibular saliva by gel filtration and affinity chromatography. They concluded, that CA1 is a high molecular weight (approximately 90 kD) membrane-bound glycoprotein and CA2 is a cytosolic isoenzyme with lower molecular weight (30 kD) comparing to CA1.

Animals↗
PubMed1989

[Recurrent dissection of aortic aneurysm with pulmonary complications].

The authors demonstrate a case of aorta aneurysm. The patient had survived despite the relatively rare and unfavourable complications presented in the observed consecutive aortic dissections. Both dissections had led to severe complications of the respiratory tract, to hemothorax and to hemoptysis.

Aged↗
PubMed1989

Indirect binding of human T-cell leukemia virus type I tax1 to a responsive element in the viral long terminal repeat.

Several laboratories have demonstrated that tandem copies of the human T-cell leukemia virus type I 21-base-pair (bp) repeat cloned upstream of either a homologous or heterologous promoter increase transcription in the presence of tax1 protein. In this report, we provide evidence for a second tax1-responsive sequence in the viral long terminal repeat. Analysis of human T-cell leukemia virus type I promoter deletion mutants and plasmids containing cloned oligonucleotide motifs demonstrated that this 47-bp sequence, located between -117 and -163, confers responsiveness to tax1. We further demonstrated that proteins present in HeLa nuclear extracts bind specifically to this tax1-responsive sequence. Mutants that affected in vivo activity also decreased in vitro binding. Using an in vitro binding assay, we demonstrated that tax1 interacts indirectly with the 47-bp sequence, most likely through protein-protein interaction. Thus, while tax1 does not bind directly to DNA to enhance transcription, it may influence sequence-specific responses by interacting with the primary DNA-protein complex.

Animals↗
PubMed1988

In vivo and in vitro autoprocessing of human immunodeficiency virus protease expressed in Escherichia coli.

The viral-specific protease of human immunodeficiency virus (HIV) has been expressed as a lacZ-protease fusion protein. This fusion protein contains protease cleavage sites at the gag/protease and protease/reverse transcriptase junctions and undergoes autoprocessing in vivo when expressed in Escherichia coli. The purified lacZ-protease fusion protein precursors also exhibit autoproteolytic activity in vitro. One cleavage product of the autoprocessing reactions is a 10-kDa protein that cross-reacts with peptide antisera prepared against the putative protease sequence. Consistent with the notion that HIV protease is an acid protease, its autoproteolytic activity is inhibited in alkaline buffers and by pepstatin A. The in vivo and in vitro autocleavage assays for HIV protease together with the overproduction of the protease should facilitate design and testing of therapeutic agents that inhibit gag-pol polyprotein processing and HIV virion maturation.

DNA Restriction Enzymes↗
PubMed1988

An Escherichia coli gene in search of a function.

The rrnB gene of Escherichia coli is preceded by an open reading frame, which is cotranscribed with rrnB both in vivo and in vitro. It has earlier been shown that a 289 amino acid protein corresponding to this gene is actually synthesized in E. coli. In this paper we show that: (1.) The transcription of this gene diminishes the stringent response of the P1 promoter of the linked rrnB gene, but this is a cis effect and is not mediated by the protein product of the gene. (2.) The functional integrity of this gene seems to be essential, because efforts to replace it by a plasmid-coded copy mutagenized by Tn5 completely failed. (3.) The protein product of this gene was strongly overproduced by a recombinant plasmid, exploiting the principle of "translational coupling". This overproduction did not change the phenotype of the host cell significantly. The protein was purified to apparent electrophoretic homogeneity.

Amino Acid Sequence↗
PubMed1988

Characterization of cellular factors that interact with the human T-cell leukemia virus type I p40x-responsive 21-base-pair sequence.

Transcriptional activation of the human T-cell leukemia virus type I (HTLV-I) long terminal repeat (LTR) by viral protein p40x requires a 21-base-pair (bp) sequence which is repeated three times within the LTR. This sequence contains a core octanucleotide (TGACGTCT) which has been attributed to be a cyclic-AMP (cAMP)-responsive element. We demonstrate here that the HTLV-I LTR can be specifically stimulated by cAMP regulators and have identified four proteins in HeLa cells that bind to the HTLV-I 21-bp sequence. We correlated the in vitro binding and transcriptional activity of one of these cellular factors (Mr, 180,000) to the trans-activation of the HTLV-I LTR by p40x. Point mutations were generated within the cAMP octanucleotide of the HTLV-I 21-bp sequence that simultaneously abolished biological responsiveness to trans-activation by p40x and to stimulation by cAMP. We found that these mutations also eliminated the binding of the 180-kilodalton HeLa factor to the HTLV-I 21-bp element. In the absence of a demonstrable DNA-binding property for p40x, we hypothesize that cellular proteins are involved, possibly through signal transduction pathways, in its trans-activation of responsive promoters.

Base Sequence↗
PubMed1987

New regulatory features of the promoters of an Escherichia coli rRNA gene.

Recombinant plasmids were constructed by fusing either promoter p1 or p2 or both promoters of the rrnB gene of Escherichia coli to a DNA fragment coding for the N-terminal alpha-peptide of beta-galactosidase. These plasmids contained various lengths of the 5'-leader region of rRNA as the 5'-terminal end of the alpha-peptide messenger. In some cases the entire 5'-terminal rRNA-coding sequence was removed, and alpha-peptide synthesis was governed by rac promoters formed by fusion of rrnBp2 and lac promoters. By measuring the level of alpha peptide, conclusions could be drawn about the activities of the promoters under various physiological conditions. It was found that the rate of transcription starting from promoter p1 or p2 might vary more than 10-fold during the growth cycle, showing a sharp maximum during outgrowth from the stationary phase into exponential growth or during nutritional shift-up. The target sequence of this regulation was localized to the leader region of the rrnB gene.

Base Sequence↗
PubMed1986

The role of upstream sequences in determining the strength of an rRNA promoter of E. coli.

In vitro transcription experiments were carried out with recombinant plasmids containing the promoters of the rrnB gene of Escherichia coli, and with deletion mutants lacking various lengths of the AT-rich sequence upstream from the P1 promoter of that gene. The main conclusions are as follows: The in vitro transcriptional activity of the P1 and P2 promoters of the rrnB gene are an order of magnitude higher on closed-circular (supercoiled) templates than on linear DNA; the strong P1 and P2 promoters are heparin-sensitive on linear templates, and on circular DNA only P2 is heparin-resistant; removal of the upstream AT-rich region did not decrease the apparent in vitro strength of the P1 promoter under standard conditions (50 mM KCl, high RNA polymerase/DNA ratio); at higher salt concentrations, or with a lower RNA polymerase/DNA ratio, the deletion mutants displayed much lower in vitro transcriptional activity than the wild-type, and the apparent weakening of the P1 promoter was roughly proportional to the length of the deleted AT-rich sequence. The implications of these findings for the possible in vivo role of the AT-rich region are discussed.

Base Sequence↗