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B Chatterjee

Publications and source records attributed to B Chatterjee.

At least 55 records · Page 3Linked to original sources
PubMed1995

Nuclear factor kappa B functions as a negative regulator for the rat androgen receptor gene and NF-kappa B activity increases during the age-dependent desensitization of the liver.

Transcriptional regulation of the steroid hormone receptor genes plays a central role in temporal changes of target cell sensitivity during development, maturation, and aging. Sequence-specific DNA-protein interactions mediate these regulatory functions. Progressive 5' deletion of the rat androgen receptor (rAR) gene immediately beyond the -572 base pair (bp) region causes a marked increase in its promoter activity. DNase I footprinting with nuclear proteins revealed a protected area encompassing -574- to -554-bp positions that begins with a perfectly palindromic nuclear factor kappa B (NF-kappa B) motif. Electrophoretic mobility shift analyses (EMSA) showed that the decameric rAR NF-kappa B site at positions -574 to -565 cross-competes with the authentic kappa immunoglobulin light chain enhancer for specific protein binding. Supershift with specific antibodies to NF-kappa B subunits confirmed that the two retarded bands observed in the EMSA with the labeled rAR probe are due to p50/p65 and p50/p50 dimers of the NF-kappa B/Rel proteins. Fragments of rAR promoter with either deletion or point mutation of the NF-kappa B site are found to be about 2- to 3-fold more effective as compared to the wild type control in driving a heterologous reporter gene in cellulo. Thus, unlike most other known cases, NF-kappa B acts as a negative regulator for the rAR gene. The physiological relevance of this repressor function is evident from a 10-fold increase in the p50/p50 form of the NF-kappa B activity in the liver of aged rats exhibiting hepatic androgen desensitization. The newly identified repressor element is a rare example of a naturally occurring perfect palindromic binding motif for the NF-kappa B/Rel family of transcription factors. This repressor factor and the positively acting age-dependent factor, ADF, described earlier (Supakar, P. C., Song, C. S., Jung, M. H., Slomczynska, M. A., Kim, J.-M., Vellanoweth, R. L., Chatterjee, B. & Roy, A. K. (1993) J. Biol. Chem. 268, 26400-26408) function to coordinate the tissue-specific down-regulation of the rAR gene during aging.

Aging↗
PubMed1995

Direct detection and characterization of rotavirus into subgroups by dot blot hybridization and correlation with 'long' and 'short' electropherotypes.

BACKGROUND: Enzyme-linked immunosorbent assay (ELISA) and polyacrylamide gel electrophoresis (PAGE) of viral RNA are well-established methods for detection of rotavirus in stool samples. Dot-blot hybridization has also been found to be a sensitive and specific technique for detection and characterization of rotaviruses. OBJECTIVES: To compare the performance of dot blot hybridization with ELISA and PAGE for detection of rotavirus in stool samples. To assess the use of dot blot hybridization for characterization of rotaviruses into subgroups. STUDY DESIGN: Stool samples were collected from 214 children presenting to the hospital with acute diarrhoea. These were assayed for rotavirus by ELISA and PAGE. Dot-blot hybridization was done with full length cloned radiolabelled c-DNA probes of gene segment 6 of SA-11 (subgroup I) and Wa (subgroup II) rotaviruses. RESULTS: Out of 214 stool samples 134 were found to be positive for rotavirus by one of the three methods. Among these 134 positive specimens 114 were positive by dot blot hybridization, this included 18 specimens which were positive only by dot blot assay. One-hundred-and-twelve of these 114 specimens could be subgrouped. Fifteen of these were classified as subgroup I, 97 as subgroup II and two had a dual subgroup specificity. Three subgroup 1 strains had a 'long' RNA pattern, whereas one subgroup II strain had a 'short' RNA pattern which has not been reported earlier for human rotaviruses. CONCLUSION: Dot blot hybridization as described here is a sensitive and specific assay for detection and subgrouping of rotaviruses. However, as there is a considerable genomic diversity among rotaviruses, the panel should include probes from all the genotypes of gene segment 6.

Journal Article↗
PubMed1995

Androgen action.

Androgens are C-19 steroids that provide major regulatory influences on male reproductive function. Testosterone, the principal androgenic steroid, is secreted by the Leydig cells of the testes. Both testosterone and its 5 alpha reduced derivative 5 alpha-dihydrotestosterone (DHT) are physiological ligands for the androgen receptor (AR). Ligand-activated AR acts as a nuclear transcription factor and mediates androgen action. AR, along with receptors for a number of C-21 steroids such as glucocorticoid, mineralocorticoid, and progesterone, share the same 15 base pair consensus element composed of 5'-GGA/TACAnnnTGTTCT-3'. Despite this cross-reactivity at the level of the DNA, physiologically, androgens regulate their target genes with a high degree of receptor specificity. Such a regulatory specificity appears to be due to multiphasic interactions involving enzymatic activation/inactivation of the steroid ligand, interaction with specific receptor-associated nuclear factors on or around the hormone response element, and differential regulation of the receptor gene expression. Conversion of testosterone to 5 alpha-dihydrotestosterone in target cells is a widespread activation mechanism that amplifies the androgenic signal. Unlike the testosterone-AR complex, DHT-activated AR has a longer half-life, and thus prolongs androgen action. Oxido-reduction of androgens by 17 beta-hydroxysteroid dehydrogenase and sulfurylation by androgen sulfotransferase are two major pathways of androgen inactivation in target cells. Prenatal deprivation of androgen action, due to mutations in either the AR or the 5 alpha-reductase gene, results in developmental abnormalities of male reproductive tissues and also cause partial or complete androgen-insensitivity syndromes. Elucidation of various molecular steps in androgen action is allowing development of improved therapeutic agents for the management of disorders of androgen action such as the prostatic hypertrophy and neoplasia.

Androgens↗
PubMed1994

Molecular analysis of four lactate dehydrogenase-A mutants in the mouse.

Four electrophoretic and/or enzyme-activity variants of murine LDH-A subunit (Ldhla-m1Neu, Ldhla-m5Neu, Ldhla-m6Neu, Ldhla-m9Neu), induced by procarbazine hydrochloride or ethylnitrosourea (ENU), were analyzed at the DNA level. The exons of the Ldhl gene from homozygous mutants were amplified by PCR and sequenced. Three mutations resulted from nucleotide substitutions in exon 5: the transitions A-->G at codons 216 (Ldhla-m5Neu) and 225 (Ldhla-m6Neu), and the transversion G-->C (Ldhla-m1Neu) at codon 222. The mutations resulted in the replacements of Glu by Gly (Ldhla-m5Neu), Gln by Arg (Ldhla-m6Neu) and Asp by His (Ldhla-m1Neu). The fourth base substitution, the transition T-->C (Ldhla-m9Neu), has been found at the GT donor splice site following the first exon; this mutation affected the efficiency of transcription. All ENU-induced mutations were A/T-->G/C transitions. The mutation events could be correlated with the biochemical and physiological alterations observed in affected mice.

Amino Acid Sequence↗
PubMed1994

Androgen and estrogen sulfotransferases of the rat liver: physiological function, molecular cloning, and in vitro expression.

Cloning of the androgen and estrogen sulfotransferases has allowed studies on their spatiotemporal regulation and physiological function. These two enzymes appear to be involved in regulating target cell sensitivity for the androgenic and estrogenic steroids. Recombinant androgen sulfotransferase produced in insect Sf9 cells through a baculoviral expression vector is capable of transferring the sulfate from PAPS (phosphoadenosine phosphosulfate) to 3 alpha, 3 beta, 17 alpha, and 17 beta hydroxyl groups of a number of steroid substrates indicating a broad range of substrate specificity.

Aging↗
PubMed1994

Superhelical density of goat mitochondrial DNA: fluorimetric studies.

EtBr binding to DNA has been used to determine the superhelical density of Goat mtDNA. The differential binding of closed circular and nicked circular DNA is estimated by simple fluorescence measurements, which is a function of state of DNA supercoiling. The method is simple and quick over the cesium chloride density gradient centrifugation studies.

Animals↗
PubMed1993

A novel regulatory element associated with age-dependent expression of the rat androgen receptor gene.

A large body of evidence indicates that the genetic program of aging has co-evolved with the sexual mode of reproduction (Partridge, L., and Barton, N. H. (1993) Nature 362, 305-311). Age-dependent changes in target cell sensitivity to reproductive hormones can be considered part of this evolutionary linkage. Here we describe a novel regulatory element in the rat androgen receptor (AR) gene promoter associated with its age-dependent expression in the liver. This element consists of two (19 and 25 base pairs) contiguous sites, one specifically binding an Age-dependent Factor (ADF) and the other an Associated Factor (AF). Both deletion and point mutations of the ADF site result in about a 5-fold decline in the AR promoter function. Unlike AF, which is relatively tissue specific, ADF appears to be ubiquitous. The ubiquitous and evolutionarily conserved nature of ADF suggests a fundamental role of this novel transcription factor in programmed gene expression.

Aging↗
PubMed1993

A distal activation domain is critical in the regulation of the rat androgen receptor gene promoter.

The far upstream region of the rat androgen receptor (AR) gene has been cloned, and the nucleotide sequence up to -2656 bp established. Nested deletion mutants of rat AR 5' flanking sequences were ligated to the luciferase reporter gene, and their promoter activities were examined in transfected COS1 cells. Results show a critical cis-acting domain located between positions -960 and -940. Deletion of this cis element resulted in a greater than 90% decrease in the promoter activity. A nuclear protein that specifically binds to this 21-nucleotide sequence was identified by gel mobility shift analysis. The -960/-940 cis element has no identify to the binding sequence of any known transcription factor. Furthermore, the cognate binding protein is present in both rat and human (HeLa) cell nuclear extracts. We conclude that a novel trans-activator interacting at the -960/-940 region plays a critical role in the regulation of AR gene expression.

Animals↗
PubMed1993

Temporal variation in the distribution of rotavirus electropherotypes in Delhi, India.

The present study was carried out to investigate the varying pattern of rotavirus electropherotypes in Delhi, India. During a 25-month study period, rotavirus was detected in 104 of 990 (10.5%) samples collected from children with acute gastroenteritis, except in the months of June 1988, June 1989, October 1989, January 1990 and March 1990. Viral RNA migration was studied by PAGE in 83 rotavirus positive samples, and 69 (83.1%) of them gave a discernible RNA pattern. A "long" RNA pattern was observed in 58 (84%) and a "short" RNA pattern in 11 (16%) samples. There were three major electropherotypes among the "long" RNA types and two major electropherotypes among the "short" RNA RNA types. Sequential appearance of electropherotypes was seen with every seasonal peak of infection.

Acute Disease↗
PubMed1992

Starch digestion and adsorption by beta-amylase of Emericella nidulans (Aspergillus nidulans).

A mutant strain of Emericella nidulans MNU 82 was isolated by multistep mutation. The beta-amylase produced by the mutant was able to digest raw starch. It was readily and strongly adsorbed onto raw starch at pH 5.0. The enzyme to starch ratio was 1950 U/g starch. The enzyme showed no correlation between the capacity of raw starch digestion and adsorption of the enzyme.

Adsorption↗
PubMed1992

Spatio-temporal expression of estrogen sulfotransferase within the hepatic lobule of male rats: implication of in situ estrogen inactivation in androgen action.

Estrogen sulfotransferase (EST) catalyzes transfer of the sulfate group from phosphoadenosine phosphosulfate to estrogenic steroids. Since estrogen sulfates do not bind to the estrogen receptor with high affinity, EST can control the intracellular level of the receptor-active estrogens. Androgen action in the rat liver, as indicated by the androgenic induction of alpha 2u-globulin, is inhibited by low levels of estrogens. Thus, in situ estrogen inactivation by EST is expected to increase hepatic androgen sensitivity. During the lifespan of the animal, rat liver undergoes three distinct phases of androgen sensitivity, i.e. prepubertal androgen insensitivity, androgen sensitivity after approximately 40 days of age, and androgen insensitivity during senescence (greater than 750 days). EST in the liver is expressed only after puberty, when the liver becomes androgen sensitive. Furthermore, localization of EST and its corresponding mRNA within the lobular unit of the liver demonstrates that only androgen-responsive hepatocytes located around the central vein contain immunoreactive EST and its corresponding mRNA. These temporal and spatial correlations of EST expression and hepatic androgen sensitivity support the concept that steroid-inactivating enzymes play important roles in sex hormone action.

Aging↗
PubMed1992

Estrogen sulfotransferase of the rat liver: complementary DNA cloning and age- and sex-specific regulation of messenger RNA.

Mammalian estrogen sulfotransferase (EST; EC 2.8.2.4) sulfurylates the hydroxyl group of estrogenic steroids by transferring the sulfate from a cosubstrate adenosine 3'-phosphate-5'-phosphosulfate. Sulfurylated steroids do not bind to the estrogen receptor with high affinity and, therefore, are hormonally inactive. We have purified rat liver EST and developed monoclonal antibody to this enzyme. By immunoscreening a lambda gt-11 expression library constructed from male rat liver cDNAs, the cDNA clone corresponding to EST was identified and isolated. A recombinant expression plasmid (pCMV5) containing this cDNA insert when transfected into COS-7 cells generated both immunologically and enzymatically active EST. With the help of this cDNA probe, we have explored the regulation of the EST mRNA in the liver and the possible role of this enzyme in sex hormone action. During the lifespan of male rats, only the young adult animals show hepatic androgen responsiveness. Also, estrogenic hormones strongly antagonize androgen action in the rat liver. Northern blot analysis of liver RNA derived from male rats of different ages shows that the androgen sensitivity of young adult animals is associated with a high expression of EST mRNA. During the same period, mRNA corresponding to dehydroepiandrosterone sulfotransferase is markedly (approximately 10-fold) down-regulated. Such a correlation is in concordance with the role of these enzymes in the maintenance of hepatic androgen sensitivity during young adult life by inactivating the estrogenic and sparing the androgenic steroids. Furthermore, the increase in the hepatic androgen sensitivity of androgen-treated female rats is also associated with the induction of EST.(ABSTRACT TRUNCATED AT 250 WORDS)

Age Factors↗
PubMed1991

Age-dependent reversal of the lobular distribution of androgen-inducible alpha 2u globulin and androgen-repressible SMP-2 in rat liver.

Hepatocytes situated at pericentral and periportal zones of the liver lobule show differences in the expression of several liver-specific genes, such as androgen-inducible alpha 2u globulin and androgen-repressible senescence marker protein-2 (SMP-2). A marked temporal difference in the expression of these two androgen-regulated genes has also been observed. The liver of the pre-pubertal male rat is insensitive to androgen, and during this period hepatocytes synthesize only SMP-2. During young adult life (greater than 40 days), the liver becomes androgen sensitive and concomitant synthesis of alpha 2u globulin and repression of SMP-2 occur. In the senescent male rat (greater than 750 days), the liver again becomes androgen insensitive when the decline in alpha 2u globulin is accompanied by an increase in SMP-2 synthesis. In this article we present results to show a correlation between the temporal and spatial (intralobular) changes in the expression of the androgen-inducible alpha 2u globulin and the androgen-repressible SMP-2 in rat hepatocytes. Results indicate that the temporal changes in hepatic androgen sensitivity are dictated by the intralobular location of the hepatocytes. Hepatocytes located around the central vein (pericentral/perivenous) may benefit from a paracrine advantage for the expression of a subset of genes, including the gene for the androgen receptor.

Aging↗
PubMed1991

Androgen receptor messenger ribonucleic acid (mRNA) in the rat liver: changes in mRNA levels during maturation, aging, and calorie restriction.

By means of RNAase protection assay with an antisense cRNA probe, we have shown that the liver of the young adult male rat contains androgen receptor (AR) mRNA to a level of 4% compared to the prostate. Steady state levels of AR mRNA in the liver show both sex and age specificity. Compared to that of the male, the female liver contains a markedly reduced amount of AR mRNA. AR mRNA is almost undetectable in livers of prepubertal male (less than 35 days old) and senescent male (greater than 750 days old) rats. Both prepubertal and senescent animals are relatively insensitive to the androgenic induction of alpha 2u-globulin, a hepatic secretory protein. The age-dependent decline in hepatic androgen sensitivity and AR mRNA level can be delayed considerably by a 40% reduction in the dietary calorie intake. Analysis of poly(A)-containing RNA from two liver cell populations, hepatocytes and nonhepatocytes, revealed that only the hepatocytes that express alpha 2u-globulin gene contain AR mRNA. From these results and our earlier observation of in vitro induction of alpha 2u-globulin in isolated rat liver, we conclude 1) that androgen can act directly on hepatocytes to promote alpha 2u-globulin synthesis; 2) that changes in the hepatic androgen sensitivity during maturation and aging are reflections of the age-dependent expression of the receptor gene; and 3) that retardation of the age-dependent loss of androgen sensitivity by calorie restriction is due to a concomitant delay in the decline of the hepatic AR mRNA level.

Aging↗
PubMed1990

Changes in hepatic androgen sensitivity and gene expression during aging.

The hepatic tissue of the male rat exhibits a gradual decline and ultimate loss in androgen responsiveness during in vivo aging. Appearance of the age-associated androgen insensitivity can be delayed by dietary calorie restriction, an effective means for life-span extension. The androgen receptor mRNA is detectable in the liver only in its androgen-responsive state. Pubertal appearance of hepatic androgen sensitivity is remarkably correlated with the concomitant appearance of a cytoplasmic androgen binding (CAB) protein. Androgen resistance during senescence is associated with the loss of hepatic CAB activity as well. We are investigating the molecular basis for the temporal modulation of this hormone sensitivity through studies on the differential expression of two androgen-responsive marker genes. These are the androgen-repressible SMP-2, and the androgen-inducible alpha 2u-globulin. Androgen resistance of hepatocytes during aging results in repression of the alpha 2u-globulin gene, and derepression of the SMP-2 gene. The structural organizations for both of these genes have been characterized. The role of nuclear transcription factors (androgen receptor and any other transacting factor(s) which may be involved) in the coordinate regulation of alpha 2u-globulin and SMP-2 during aging and nutritional manipulation is being explored to establish the molecular mechanism of andropause in the liver.

Aging↗
PubMed1990

The senescence marker protein (SMP-2) of the rat liver: purification, immunochemical characterization and age-dependent regulation.

In vitro translation of total rat hepatic mRNAs has identified a 31 kilodalton senescence marker protein (SMP-2) which is present in higher amounts in prepubertal and senescent males than in the post-pubertal adult male (more than 10-fold). SMP-2 is an androgen-repressible protein. The negative regulation of the SMP-2 gene activity by androgen accounts for its increased expression during the androgen insensitive states of the prepubertal and senescent livers, and its constitutive expression in the female liver. A combination of separation procedures including salt fractionation, chromatofocusing, ion-exchange chromatography and preparative gel electrophoresis have led to the purification of SMP-2 to apparent homogeneity. The purified protein showed the same electrophoretic mobility as the sex- and age-specific in vitro translation product of hepatic mRNAs. The polyclonal antibody to SMP-2 was produced in the rabbit. The antibody selectively reacted with the 31 kDa sex- and age-specific translation product of hepatic mRNAs. Western blot analysis of the liver cytosol confirms monospecificity of the antiserum, as well as age- and sex-dependent changes in the tissue level of SMP-2. Histochemical staining of liver sections with the antiserum reveals a preferential periportal localization of SMP-2 in the hepatocytes. This finding is in marked contrast to the androgen-inducible alpha 2u globulin which is preferentially synthesized and localized in the pericentral hepatocytes. Thus, the zonal distribution of SMP-2 correlates with polarized androgen sensitivity of the hepatocytes within the liver lobule.

Aging↗