PubMed Health⌕ Search

Biomedical subjects

S Habib

Publications and source records attributed to S Habib.

At least 19 recordsLinked to original sources
PubMed2000

Differential activity of two non-hr origins during replication of the baculovirus Autographa californica nuclear polyhedrosis virus genome.

The identification of potential baculovirus origins of replication (ori) has involved the generation and characterization of defective interfering particles that contain major genomic deletions yet retain their capability to replicate by testing the replication ability of transiently transfected plasmids carrying viral sequences in infected cells. So far, there has not been any evidence to demonstrate the actual utilization of these putative origins in Autographa californica multinucleocapsid nuclear polyhedrosis virus (AcMNPV) replication. By using the method of origin mapping by competitive PCR, we have obtained quantitative data for the ori activity of the HindIII-K region and the ie-1 promoter sequence in AcMNPV. We also provide evidence for differential activity of the two ori in the context of the viral genome through the replication phase of viral infection. Comparison of the number of molecules representing the HindIII-K and ie-1 origins vis-à-vis the non-ori polH region in a size-selected nascent DNA preparation revealed that the HindIII-K ori is utilized approximately 14 times more efficiently than the ie-1 region during the late phase of infection. HindIII-K also remains the more active ori through the early and middle replication phases. Our results provide in vivo evidence in support of the view that AcMNPV replication involves multiple ori that are activated with vastly different efficiencies during the viral infection cycle.

Animals↗
PubMed1999

CDR1, a multidrug resistance gene from Candida albicans, contains multiple regulatory domains in its promoter and the distal AP-1 element mediates its induction by miconazole.

We previously demonstrated that the CDR1 gene, encoding a multidrug transporter in Candida albicans, is differentially upregulated by various drugs and steroids. In order to get an insight into the molecular basis of the induction of this gene we analyzed its promoter region. The transcription start site was mapped to 63 nucleotides upstream of the initiating ATG. Reporter assays revealed the presence of four upstream activating and four upstream repressing sequence domains along the entire promoter. Like the native gene, promoter-luciferase recombinants showed enhanced activity in response to various stresses like drugs, human steroid hormones and heavy metals. Mutational analysis demonstrated that while the proximal promoter (-345/+1) contains all the regulatory domains required for its induction by various other stresses, the miconazole response is mediated via the distal promoter (-857/-1147), harboring an AP-1 site. The involvement of the AP-1 element in mediating the latter effect was evident by an increase in AP-1 binding activity following miconazole treatment.

Antifungal Agents↗
PubMed1998

Lack of inducible nitric oxide synthase in bronchial epithelium: a possible mechanism of susceptibility to infection in cystic fibrosis.

Cystic fibrosis (CF) is an inherited disorder associated with severe inflammation and repeated bacterial infection and colonization in the lung. Airway epithelium is involved in defence against bacteria, but this system may be defective in CF. Pro-inflammatory cytokines can stimulate the expression of inducible nitric oxide synthase (iNOS), an enzyme generating nitric oxide, which functions as an important mediator in host defence mechanisms. To understand better the poor resistance to infections in the CF lung, the expression of the iNOS gene was investigated in explanted lungs from patients with cystic fibrosis (n = 13), bronchiectasis (n = 3), emphysema (n = 14), and in normal lungs (n = 8). In addition, bronchial epithelial cell lines were examined to study iNOS gene expression in vitro. Strong immunoreactivity for iNOS was seen in inflammatory cells and bronchial epithelium in all the diseased lungs, except for bronchial epithelium in CF. Quantitative analysis showed a significant reduction in the area of epithelium immunostained in CF [CF 6.8 +/- 1.6 (% +/- SEM); emphysema 18.2 +/- 2.8; normal 9.6 +/- 0.8, P < 0.01], regardless of steroid treatment. These results were supported by in situ hybridization of iNOS mRNA, which showed a pattern of gene expression in CF, emphysema, and normal lung which paralleled that of protein immunoreactivity. Stimulation with cytokines (IL-1 beta, TNF-alpha, and IFN-gamma) increased the expression of iNOS mRNA detected by reverse transcriptase-polymerase chain reaction (RT-PCR) in cultures of normal (16HBE14o-), but not CF (CFBE41o-, with delta F508 CFTR mutation) epithelial cells. Expression of iNOS in inflammatory cells suggests that the gene is normal in CF. Absence of iNOS from bronchial epithelium may be due to low expression of the gene resulting from abnormalities in the signalling system that normally causes induction, such as cytokine receptors, second messengers or transcription factors. The resulting deficiency of the nitric oxide defence system may be relevant to the susceptibility of CF patients to pulmonary bacterial colonization.

Adult↗
PubMed1998

Determinants of regional myocardial function in patients with chronic significant coronary stenosis or occlusion.

BACKGROUND: Our aim was to study the effect of collateral flow, stenosis severity, antegrade flow, and location of the lesion on regional myocardial function in patients with chronic left anterior descending (LAD) coronary artery disease. METHODS AND RESULTS: Seventy-four patients who underwent coronary angiography and ventriculography were divided into three groups: group A (n = 9), patients with normal coronary angiogram and ventriculogram; group B (n = 32), patients with LAD stenosis >75%; and group C, 33 patients with LAD occlusion. The effect of collateral flow, stenosis severity, location, and antegrade flow on regional myocardial function in the LAD territory was studied. Regional function was impaired in both groups B and C. In group B, univariate analysis confirmed that antegrade flow had a significant effect on regional function (p < 0.02). With the use of multiple regression, none of the other variables had an additional independent effect. In group C, no significant correlation was found between regional function and the study variables by univariate analysis; however, multiple regression revealed a significant correlation between anterobasal function, the lesion site (p = 0.01), and collateral flow (p = 0.02). CONCLUSIONS: In patients with LAD stenosis, antegrade flow has the strongest effect on regional myocardial function, with no additional effect of visible collaterals. In patients with LAD occlusion, both angiographic collateral flow and the site of the lesion have a significant effect on the function of the anterobasal area.

Blood Flow Velocity↗
PubMed1998

The modulatory effects of bradykinin B1 and B2 receptor antagonists upon viscero-visceral hyper-reflexia in a rat model of visceral hyperalgesia.

This study assessed the relative involvement of the two bradykinin (Bk) receptors (B1 and B2) in the viscero-visceral hyper-reflexia (VVH) and plasma extravasation observed in an animal model of cystitis. The effects of the competitive receptor antagonists des-Arg9[Leu8]-Bk (B1) and HOE 140 (B2) were tested both in prophylactic (pre-inflammation administration) and therapeutic (post-inflammation administration) scenarios. Compared with control animals, des-Arg9[Leu8]-Bk had no effect on the hyper-reflexic response of the bladder to inflammation unless it was administered 5 h after inflammation. However, HOE 140 was able to attenuate the inflammation-induced viscero-visceral hyper-reflexia (VVH) at doses of 1 mg/kg, 2 mg/kg and 7.5 mg/kg. This effect was apparent whether the drug was administered before, or after inflammation. In contrast, neither compound was effective in attenuating the intravesical plasma extravasation induced by turpentine. The data therefore suggest that the VVH and tissue inflammation responses are mediated via different mechanisms. In addition, the turpentine-induced VVH appears to be mediated, at least partially, by the B2 receptor in the early phase, with the B1 receptor only becoming important later.

Adrenergic beta-Antagonists↗
PubMed1998

The host factor polyhedrin promoter binding protein (PPBP) is involved in transcription from the baculovirus polyhedrin gene promoter.

Hypertranscription and temporal expression from the Autographa californica nuclear polyhedrosis (AcNPV) baculovirus polyhedrin promoter involves an alpha-amanitin-resistant RNA polymerase and requires a trans-acting viral factor(s). We previously reported that a 30-kDa host factor, polyhedrin promoter binding protein (PPBP), binds with unusual affinity, specificity, and stability to the transcriptionally important motif AATAAATAAGTATT within the polyhedrin (polh) initiator promoter and also displays coding strand-specific single-stranded DNA (ssDNA)-binding activity (S. Burma, B. Mukherjee, A. Jain, S. Habib, and S. E. Hasnain, J. Biol. Chem. 269:2750-2757, 1994; B. Mukherjee, S. Burma, and S. E. Hasnain, J. Biol. Chem. 270:4405-4411, 1995). We now present evidence which indicates that an additional factor(s) is involved in stabilizing PPBP-duplex promoter and PPBP-ssDNA interactions. TBP (TATA box binding protein) present in Spodoptera frugiperda (Sf9) cells is characteristically distinct from PPBP and does not interact directly with the polh promoter. Replacement of PPBP cognate sequences within the polh promoter with random nucleotides abolished PPBP binding in vitro and also failed to express the luciferase reporter gene in vivo. Phosphocellulose fractions of total nuclear extract from virus-infected cells which support in vitro transcription from the polh promoter contain PPBP activity. When PPBP was sequestered by the presence of oligonucleotides containing PPBP cognate sequence motifs, in vitro transcription of a C-free reporter cassette was affected but was restored by the exogenous addition of nuclear extract containing PPBP. When PPBP was mopped out in vivo by a plasmid carrying PPBP cognate sequence present in trans, polh promoter-driven expression of the luciferase reporter was abolished, demonstrating that binding of PPBP to the polh promoter is essential for transcription.

Animals↗
PubMed1997

Involvement of host factors in transcription from baculovirus very late promoters -- a review.

The baculovirus expression vector system has emerged as the system of choice for the expression of a number of heterologous genes of both prokaryotic and eukaryotic origin. This system utilizes the baculovirus very late, hyperactive polyhedrin and p10 promoters to drive the transcription of foreign genes. Regulation of transcription from these promoters is presently not well understood even though a number of viral gene products that may be important for transcription have been identified. Fresh insight into host-virus interactions during baculovirus pathogenesis is now offered by the identification of insect host factors that interact with transcriptionally essential motifs of these promoters as well as cis-acting enhancer-like elements upstream from the promoter.

Base Sequence↗
PubMed1997

Development of a functional assay for Ca2+ release activity of IP3R and expression of an IP3R gene fragment in the baculovirus-insect cell system.

Receptor-stimulated phosphoinositide (PI) hydrolysis is an important and ubiquitous mechanism of intracellular signaling. Inositol 1,4,5-trisphosphate (IP3), generated by phosphoinositide (PI) hydrolysis, binds to and gates an intracellular Ca2+ channel, the IP3 receptor (IP3R), which is therefore a central component of this signaling cascade. Here we describe the development of a baculovirus (BV)/Sf (S. frugiperda) cell system that can be used to look at IP3R function. Agonist-evoked changes in intracellular Ca2+ levels [Ca2+]i were measured (using Fura2) in Sf cells expressing the gene encoding the muscarinic acetylcholine receptor (vm1AchR). Furthermore, we have constructed a recombinant BV (vIP3R), with the core of the IP3R ligand-binding domain from the Drosophila IP3R, under the polyhedrin promoter. The recombinant protein from such a virus was expected to act as a large ligand sink for IP3, generated by stimulation of vm1AchR. Cells coinfected with recombinant BV carrying the potential dominant-negative vIP3R construct and vm1AchR have been used to assay the modulation of IP3R-mediated Ca2+ release, by the ligand sink.

Animals↗
PubMed1996

A 38-kDa host factor interacts with functionally important motifs within the Autographa californica multinucleocapsid nuclear polyhedrosis virus homologous region (hr1) DNA sequence.

We recently demonstrated that the Autographa californica multinucleocapsid nuclear polyhedrosis virus homologous region (hr1) enhances transcription from the viral polyhedrin promoter and also functions as a putative origin of replication (ori). Hr1, carrying five 28-base pair core palindrome units, has also been mapped with respect to its enhancer and ori functions (Habib, S., Pandey, S., Chatterji, U., Burma, S., Ahmad, R., Jain, A., and Hasnain, S. E. (1996) DNA Cell Biol. 15, 737-747). A 38-kDa host factor termed hr1-binding protein (hr1-BP) binds with high specificity and affinity (Kd approximately 6.5 x 10(-11) M) to functionally important motifs within hr1. The core palindrome as well as sequences immediately flanking it are required for this interaction. Divalent cations are not essential, and ionic interactions play only a minor role in complex formation. hr1-BP binds through the minor groove of the double helix to multiple sites within hr1, and binding occurs as a function of the number of modules within hr1. Phosphorylation of hr1-BP is important for host factor-hr1 interaction. Hr1-BP differs in several respects from the other host factor, polyhedrin promoter-binding protein, described previously (Burma, S., Mukherjee, B., Jain, A., Habib, S., and Hasnain, S. E. (1994) J. Biol. Chem. 269, 2750-2757). When hr1-BP was sequestered out, in vivo, by a plasmid carrying hr1 alone, the hr1-mediated enhancement of reporter expression was abolished, demonstrating that the binding of hr1-BP may be crucial for the enhancer activity of the dual function hr1 element.

Base Sequence↗
PubMed1996

Expression and functional characterisation of the clpC gene of Mycobacterium leprae: ClpC protein elicits human antibody response.

This paper reports the expression of a previously described gene [Nath and Laal, Nucleic Acids Res. 18 (1990) 4935], currently identified as the clpC gene of Mycobacterium leprae, using an in vitro rabbit reticulocyte lysate-coupled transcription/translation system. The produced protein moved as a 95-kDa band on SDS-PAGE. An additional band of 79 kDa was seen which may have resulted from a GTG codon downstream to the initiating ATG in the clpC sequence. A threefold increase in synthesis of the 95-kDa protein was achieved by altering the translation codon context sequence of the ATG start codon. The ClpC (caseinolytic protease C) amino acid sequence, which contained two nucleotide-binding sites, exhibited in vitro ATP binding. Of functional significance was its immunoreactivity in human subjects with mycobacterial infection. Leprosy and tuberculosis patients with active disease had antibodies which recognised ClpC in dot ELISA.

Adenosine Triphosphate↗
PubMed1996

Bifunctionality of the AcMNPV homologous region sequence (hr1): enhancer and ori functions have different sequence requirements.

The Autographa californica multinucleocapsid nuclear polyhedrosis virus (AcMNPV) homologous region sequence hr1 is a putative origin of replication (ori) sequence and can also function as a transcriptional enhancer for delayed-early genes. We demonstrate that this 750-bp sequence, carrying five 28-bp core palin-dromes, enhances expression from the very late polyhedrin promoter up to 11-fold in a classical enhancer fashion in transient expression assays. Enhancement is at the level of transcription, as evident from RNase protection assay analysis. It is mediated by an alpha-amanitin-insensitive RNA polymerase from the authentic polyhedrin promoter transcription start site and follows the temporal activation profile characteristic of the polyhedrin promoter. Three lines of evidence conclusively demonstrated that hr1 acts typically as an enhancer of polyhedrin gene transcription independent of its role as an ori: (i) linearized hr1-reporter plasmids, incapable of replicating in the host cell, could enhance transcription from the promoter; (ii) reporter plasmid copy number was not affected by the presence of aphidicolin during transfection; (iii) reporter plasmid DNA recovered from Sf9 cells was sensitive to Dpn I confirming its unreplicated state in the transfection regime followed by us. Molecular dissection of the hr1 sequence elements revealed that a core palindrome alone can function as an ori sequence whereas a palindrome along with flanking sequences is essential for the enhancer activity. Enhancement of luciferase expression from the polyhedrin promoter is a function of the number of core palindromes and flanking sequences. Our results demonstrate that hr1, which has several motifs for enhancer binding proteins and transcription factors, has a dual role associated with both DNA replication and transcriptional enhancement.

Animals↗