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S Barik

Publications and source records attributed to S Barik.

At least 37 records · Page 2Linked to original sources
PubMed1998

Role of cellular actin in the gene expression and morphogenesis of human respiratory syncytial virus.

Cytoskeletal protein actin and nonactin cellular proteins were essential for human respiratory syncytial virus (RSV) gene expression. In vitro, specific antibodies against actin inhibited RSV transcription, whereas antibodies against other cytoskeletal proteins had little or no effect. Affinity purified cellular actin or bacterially expressed recombinant actin activated RSV transcription. However, optimal transcription required additional cellular protein(s) that appeared to function as accessory factor(s) for actin. In the absence of actin, these proteins did not activate viral transcription. Purified viral nucleocapsids contained actin, but no cytokeratin, tubulin, or vimentin. Cytochalasin D or DNasel--agents that destabilize actin polymers--had little effect on RSV transcription. RSV infection itself seemed to alter the structure of the cellular actin filaments. Treatment of infected cells with cytochalasin D produced a more severe disruption of the filaments and drastically reduced the production of infectious virus particles but still had little effect on intracellular synthesis of viral macromolecules. Thus actin seems to serve a dual role in RSV life cycle: its monomeric form as well as polymeric form activate viral transcription, while only the microfilament form may take part in viral morphogenesis and/or budding.

Actin Cytoskeleton↗
PubMed1998

Inhibitor-1 interaction domain that mediates the inhibition of protein phosphatase-1.

Inhibitor-1 (I-1), a cyclic AMP-regulated phosphoprotein, inhibits protein phosphatase-1 (PP1) activity in response to hormones. The molecular mechanism for PP1 inhibition by I-1 remains unknown. Mutation of nine acidic residues lining a proposed I-1-binding channel in rabbit PP1alpha yielded one mutant (E256A) slightly impaired in its inhibition by I-1, with the IC50 increased by 3-fold, and one mutant (E275R) located in the beta12-beta13 loop that showed 4-fold enhanced inhibition by I-1. Substituting Tyr-272, a proposed binding site for the toxins okadaic acid and microcystin-LR, in the beta12-beta13 loop with Trp, Phe, Asp, Arg, or Ala impaired PP1alpha inhibition by I-1 by 8-10-fold. Chemical mutagenesis of the Saccharomyces cerevisiae PP1 gene (GLC7) yielded 20 point mutations in the PP1 coding region. Two-hybrid analyses and biochemical assays of these yeast enzymes identified four additional residues in the beta12-beta13 loop that were required for PP1 binding and inhibition by I-1. Ten-fold higher concentrations of I-1 were required to inhibit these mutants. Finally, deletion of the beta12-beta13 loop from PP1alpha maintained full enzyme activity, but attenuated inhibition by I-1 by >100-fold. These data identified the beta12-beta13 loop in the PP1 catalytic subunit as a domain that mediates binding and enzyme inhibition by I-1.

Binding Sites↗
PubMed1998

Hypothermic effects of dopamine D3 receptor agonists in the island of Calleja Magna. Potentiation by D1 activation.

The selective functions of D3 receptors in the brain are still poorly understood, mainly because all the ligands active at dopamine D3 receptors have also a high affinity for the D2 receptors. However, it is possible to study selectively D3 receptor function because some brain structures, such as the islands of Calleja, contain D3 and not D2 receptors. The position of the island of Calleja Magna in the rat brain makes it possible to inject dopamine D3 ligands into the vicinity of these D3 receptors, and to study their behavioral role, with no concomitant action on D2 receptors. We studied the effects on body temperature and on locomotion of unilateral microinjections of D2/D3 receptors ligands into the island of Calleja Magna and into the adjacent nucleus accumbens. The results show that D3 agonists injected into the island of Calleja Magna decrease body temperature and that this effect is potentiated by simultaneous injection of the D1 agonist SKF 38393. D3 agonists have no effect on locomotor activity in the island of Calleja Magna. In the nucleus accumbens, the D3 agonists have only weak effects on body temperature, but, when associated with a D1 agonist, strongly stimulate locomotor activity. The effects on body temperature of unilateral microinjections of dopamine agonists into unilaterally dopamine-depleted animals are the same as those in nondepleted ones. This indicates that the D3 receptors are localized postsynaptically in the island of Calleja Magna.

Animals↗
PubMed1998

Time course of hypo-osmotic swellings of human spermatozoa: evidence of ordered transition between swelling subtypes.

The hypo-osmotic swelling test (HOST or HOS test) usually takes into consideration the total HOS response value with no emphasis either on the value of the response subtypes or the response evaluation time. This study investigated the time course of HOS responses and analysed their physiological relevance. Raw semen spermatozoa and Percoll washed spermatozoa were used in the experiment. The morphological changes in the sperm tail were monitored by incubating the spermatozoa in the hypo-osmotic solution for 16 different time periods. The HOS reactive spermatozoa and the type of HOS reaction (swelling subtypes) of the samples subjected to different duration of treatment were identified under a phase contrast microscope. Also the fate of individual spermatozoa in a hypo-osmotic environment were monitored for 30 min. In spermatozoa exposed to a hypo-osmotic solution, the motility lasted usually less than 2 min and motility characteristics were uniquely different from that of the spermatozoa under iso-osmotic conditions. The HOS response development was permanent but the motility loss due to hypo-osmotic shock was reversible up to 1 min of incubation. There was an indication of ordered transition among the HOS swelling subtypes apparently initiating with subtype b destined to c, d, e, f and g. Further, the subtypes a and g showed gradual decrease and increase, respectively, while subtype b showed abrupt initial increase and then gradual decrease. Transition from b to g could be direct or via one or more than one subtypes. Ultrastructure based analysis indicated that HOS response subtypes are the apparent reflection of the differences in the cytoskeletal assembly of the sperm tail and thus may be identifying different physiological variants in the sperm population. These results indicate that shorter incubation is essential to document the kinetics of various HOS responses but the conventional HOS test misses these important HOS features because of lengthy incubation. Since the time course of ordered transition of HOS responses will vary more than the total HOS response in semen of different aetiologies, the importance of HOS response subtypes and response evaluation time should be taken into consideration when applying HOS test.

Cell Separation↗
PubMed1998

Flexible outpatient hysteroscopy without anaesthesia: a safe, successful and well tolerated procedure.

The objective of this study was to assess the feasibility and tolerance of diagnostic outpatient flexible hysteroscopy without anaesthesia. Records from 554 consecutive patients were analysed retrospectively. Success rate, reasons for failure, adverse reactions and level of pain were the main outcome measures. Hysteroscopy was successful in 90.5% of patients and well tolerated in 93.3%; 5.4% experienced moderate to severe pain. Inability to negotiate the cervical canal accounted for 47% of failed procedures and poor view for 42%. These results suggest that flexible outpatient hysteroscopy without anaesthesia is a successful and well tolerated procedure.

Adult↗
PubMed1998

Persistent activation of RelA by respiratory syncytial virus involves protein kinase C, underphosphorylated IkappaBbeta, and sequestration of protein phosphatase 2A by the viral phosphoprotein.

Respiratory syncytial virus (RSV) activated the RelA (p65) subunit of nuclear factor kappa B (NF-kappaB) over many hours postinfection. The initial activation coincided with phosphorylation and degradation of IkappaBalpha, the cytoplasmic inhibitor of RelA. During persistent activation of NF-kappaB at later times in infection, syntheses of inhibitors IkappaBalpha as well as IkappaBbeta were restored. However, the resynthesized IkappaBbeta was in an underphosphorylated state, which apparently prevented inhibition of NF-kappaB. Use of specific inhibitors suggested that the pathway leading to the persistent-but not the initial-activation of NF-kappaB involved signaling through protein kinase C (PKC) and reactive oxygen intermediates of nonmitochondrial origin, whereas phospholipase C or D played little or no role. Thus, RSV infection led to the activation of NF-kappaB by a biphasic mechanism: a transient or early activation involving phosphorylation of the inhibitor IkappaB polypeptides, and a persistent or long-term activation requiring PKC and the generation of hypophosphorylated IkappaBbeta. At least a part of the activation was through a novel mechanism in which the viral phosphoprotein P associated with but was not dephosphorylated by protein phosphatase 2A and thus sequestered and inhibited the latter. We postulate that this led to a net increase in the phosphorylation state of signaling proteins that are responsible for RelA activation.

Amino Acid Sequence↗
PubMed1997

Identification, cloning, and mutational analysis of the casein kinase 1 cDNA of the malaria parasite, Plasmodium falciparum. Stage-specific expression of the gene.

The cDNA for casein kinase 1 (CK1) of Plasmodium falciparum was cloned, sequenced, and expressed in bacteria. The single major open reading frame of the 1.2-kilobase pair cDNA coded for a 324-amino acid polypeptide of approximately 37 kDa, the predicted sequence of which showed strong identity with known CK1 isoforms. The purified recombinant enzyme exhibited properties characteristic of CK1, such as inhibition by CK1-7, the ability to phosphorylate a highly specific peptide substrate, and a strong preference for ATP over GTP. A casein kinase activity, partially purified from soluble extracts of P. falciparum by affinity chromatography through CK1-7 columns displayed identical properties. The activity showed a stage-specific expression in the parasite, in the order trophozoite > ring >> schizont. Northern analysis indicated the existence of two major CK1 mRNAs, 2.4 and 3.2 kilobase pairs long, the levels of which were in the order ring > schizont > trophozoite. Mutagenesis of recombinant CK1 defined important amino acid residues and their potential role in the conformation of the enzyme. The malarial CK1 appeared to be the one of the smallest and perhaps the most primitive CK1 enzymes known, containing little sequence information beyond the minimal catalytic domain.

Amino Acid Sequence↗
PubMed1997

Transcriptional induction of multiple cytokines by human respiratory syncytial virus requires activation of NF-kappa B and is inhibited by sodium salicylate and aspirin.

Infection of the lung epithelial cell line A549 by respiratory syncytial virus (RSV) resulted in the elevated synthesis of multiple cellular cytokines, including a number of interleukins (ILs). Detailed studies of IL-11 induction revealed that it required infection by viable virus and involved a net increase in the steady state level of IL-11 mRNA. Nuclear run-on assays showed a direct effect of RSV on IL-11 gene transcription. Mutational analysis of the IL-11 promoter fused to a reporter luciferase gene demonstrated the requirement of a region 720 nucleotides upstream of the mRNA start site in the transcriptional induction of IL-11 by RSV. Two nearly identical 10-nucleotide-long sequences GGGGTCTCCC and GGGTCTCCCC in this region resembled the NF-kappa B consensus motif. Mutation of either sequence greatly reduced RSV-mediated induction of IL-11 promoter activity. NF-kappa B sites in IL-1 alpha, IL-6, and IL-8 promoters were also required for RSV-mediated induction of transcription of these promoters. Immunological studies and use of reporter gene constructs provided direct evidence for the activation and nuclear translocation of NF-kappa B by RSV. Sodium salicylate and aspirin, inhibitors of NF-kappa B activation, abolished transcriptional induction of all these cytokines by RSV. Together, these studies demonstrated an essential role of NF-kappa B in RSV-mediated transcription of multiple cytokines genes and suggested a possible use of salicylates in managing airway inflammation and viral pathogenesis during RSV infection.

Anti-Inflammatory Agents, Non-Steroidal↗
PubMed1997

Extractability and nutritional value of leaf protein from tropical aquatic plants.

In a study conducted on the extraction of protein from the leaves of 30 freshwater aquatic plants, the highest standing crop fresh yield was found in Typha latifolia (2650 g/m2). The Bio-Medical Data Processing (BMDP) K-means clustering program with K = 2 showed that 11 of the 30 plants had a high protein nitrogen extractability as well as a high nitrogen content of the extracted protein. Among these, leaf protein from Allmania nodiflora had the highest content of crude protein (62.7%) and beta-carotene (782.4 micrograms/g). Leaf protein prepared from Hygrophila spinosa, Ottelia alismoides and Polygonum barbatum had low in-vitro digestibility. The levels of alkaloids and polyphenols were lower in the extracted protein compared to that present in the original leaf sample.

Dietary Proteins↗
PubMed1997

Phosphorylation of the hepatitis delta virus antigens.

We used two-dimensional electrophoresis (nonequilibrium pH gradient electrophoresis followed by sodium dodecyl sulfate-10% polyacrylamide gel electrophoresis) coupled with 32P labeling and immunoblotting detection with 125I-protein A to detect and quantitate phosphorylation of the large and small forms of the delta antigen (deltaAg-L and deltaAg-S, respectively). Analysis of deltaAg species from the serum and liver of an infected woodchuck as well as deltaAg species expressed in and secreted from transfected Huh7 cells revealed the following. (i) No detectable phosphorylation of deltaAg-S occurred. (ii) In virions from the serum of an infected animal and in the particles secreted from cotransfected cells, none of the deltaAg-L was phosphorylated. (iii) Only in the infected liver and in transfected cells was any phosphorylation detected; it corresponded to a monophosphorylated form of deltaAg-L. Given these results, we carried out serine-to-alanine mutagenesis of the deltaAg-L to determine whether the monophosphorylation was predominantly at a specific site on the unique 19-amino-acid (aa) extension. We mutated each of the two serines, aa 207 and 210, on this extension and also the serine at aa 177. These three mutations had no significant effect on phosphorylation. In contrast, mutagenesis to alanine of the cysteine at aa 211, which normally acts as the acceptor for farnesylation, completely inhibited phosphorylation. Our interpretation is that the site(s) of phosphorylation is probably not in the 19-aa extension unique to deltaAg-L and that phosphorylation of deltaAg-L may depend upon prior farnesylation. The possible significance of the intracellular phosphorylated forms of deltaAg-L is discussed.

Animals↗
PubMed1997

Relationship between opacity of transformed E. coli colonies and over-expression of the recombinant transcript.

Following transformation with recombinant plasmid clones, E. coli BL21(DE3) often produced extremely opaque colonies on a standard semisolid agar plate, as compared with the translucent colonies produced by normal, untransformed bacteria A standard BL21(DE3) culture consisted of two kinds of cells one kind produced translucent colonies and the other produced opaque colonies upon transformation by recombinant plasmids. The translucent-generating phenotype often switched to the opaque-generating phenotype, which was irreversible. Opacity in the BL21(DE3) background was correlated to a higher preinduced level of T7 RNA polymerase, presumably through a stable and inheritable genetic change. In all E. coli strains tested, a robust transcription of the recombinant gene from the plasmid clone was found to be an essential prerequisite for every high opacity; translation of the RNA was not required. The degree of opacity was also determined by the nature of the insert in a given strain background. Colony, opacity generally, but not invariably, correlated with a smaller colony size on semisolid agar and a reduced growth rate in liquid culture.

Bacteriophage T7↗
PubMed1997

Inherited factor X deficiency in two brothers.

Two brothers born to same parents were diagnosed with inherited factor X deficiency of severe type. Clinical presentation in both the cases were haemarthrosis. The elder brother was diagnosed in the year 1991 when he was four and half years old. Recently the youngest child in the family also presented with haemarthrosis at age of one and half years. Diagnosis was made by abnormal results of Coagulation factors screening mainly Prothrombin time, Activated partial thromboplastin time, Russell's viper venom test, mixing tests factor X assay. Both the brothers had Factor X activity less than one percent.

Child, Preschool↗
PubMed1996

Evidence for a functional role of the dopamine D3 receptors in the cerebellum.

It has recently been shown that the lobules 9 and 10 of the cerebellum contain high densities of dopamine D3 receptors (and almost no D2 receptors [Bouthenet, M.L., Souil, E., Martres, M.P., Sokoloff, P., Giros, B. and Schwartz, J.C., Brain Res., 564 (1991) 203-219; Sokoloff, P., Giros, B., Martres, M.P., Bouthenet, M.L. and Schwartz, J.C., Nature, 347 (1990) 146-151]), and the functional role of this cerebellar dopamine system is unknown. In this study, using microinjections of dopamine receptor ligands into the lobules 9 and 10 of the cerebellum and into the nucleus accumbens, we show that the cerebellar dopamine D3 receptors have a functional role in the regulation of locomotor activity. When microinjected into the lobules 9 and 10 of the cerebellum, amisulpride (a dopamine D2 and D3 antagonist) and nafadotride (a preferential D3 antagonist) dose-dependently alter locomotor activity. At low doses, both agents stimulate locomotor activity, while inhibition is observed at higher doses. Haloperidol (a D2 antagonist) and apomorphine (a dopamine agonist) have no effects at low doses, but decrease locomotor activity at high doses. Similar effects are found in the nucleus accumbens, however the effects are stronger in the nucleus accumbens than in the cerebellum. Therefore, the dopamine D3 receptor system in lobules 9 and 10 of the cerebellum has a functional role, similar but weaker than the D3 receptor system in the nucleus accumbens.

Amisulpride↗