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S Acharya

Publications and source records attributed to S Acharya.

At least 55 records · Page 3Linked to original sources
PubMed1997

The human mismatch recognition complex hMSH2-hMSH6 functions as a novel molecular switch.

The mechanism of DNA mismatch repair has been modeled upon biochemical studies of the E. coli DNA adenine methylation-instructed pathway where the initial recognition of mismatched nucleotides is performed by the MutS protein. MutS homologs (MSH) have been identified based on a highly conserved region containing a Walker-A adenine nucleotide binding motif. Here we show that adenine nucleotide binding and hydrolysis by the human mismatch recognition complex hMSH2-hMSH6 functions as a novel molecular switch. The hMSH2-hMSH6 complex is ON (binds mismatched nucleotides) in the ADP-bound form and OFF in the ATP-bound form. These results suggest a new model for the function of MutS proteins during mismatch repair in which the switch determines the timing of downstream events.

Adenosine Diphosphate↗
PubMed1997

Prenatally diagnosed neuroblastoma.

BACKGROUND: Prenatally diagnosed neuroblastomas have been reported in increasing numbers over the past several years, and there are now a few reviews based on up to 21 cases. The purpose of this article is to review the clinical and biologic features of prenatally diagnosed neuroblastoma based on a review of 55 cases. METHODS: A review was conducted of 3 cases seen at the study institution and 52 other cases reported thus far in the literature. RESULTS: Prenatal diagnosis was made usually after 32 weeks of gestation. Approximately 93% of the tumors were adrenal in origin, and 44% of these were cystic. Thirty-seven patients (67%) had Stage I disease, 12 (22%) had Stage IV-S disease, and only 3 (5%) had Stage IV disease. The DNA index was favorable (> 1) in 14 of 16 patients studied. None of these 16 patients studied had amplification of the N-myc oncogene. Catecholamines were elevated in only 33% of the patients. The liver was the most common site of dissemination, which was observed in 25% of patients; bone involvement was not observed in any patient. Ultrasonography failed to detect existing hepatic metastasis in three patients. Primary surgical resection was performed in 47 patients (85%). Chemotherapy was given to five patients and radiotherapy to three. Of the 50 patients for whom follow-up information was available, 45 (90%) were alive at a range of 2-120 months from diagnosis. CONCLUSIONS: Prenatally diagnosed neuroblastomas are predominantly adrenal in origin and frequently cystic. The liver is the most common site of dissemination and bone involvement is notably absent. The vast majority of these infants have a favorable stage of disease (I, II, and IV-S) and favorable biologic features, and consequently have an excellent prognosis. Although surgery alone is curative for most patients, a period of observation may avoid surgery in some individuals who may achieve spontaneous regression.

Adrenal Gland Neoplasms↗
PubMed1997

Transducin-alpha C-terminal peptide binding site consists of C-D and E-F loops of rhodopsin.

The binding of heterotrimeric GTP-binding proteins (G-proteins) to serpentine receptors involves several independent contacts. We have deduced the points of interaction between mutant bovine rhodopsins and alphat-(340-350), a peptide corresponding to the C terminus of the alpha subunit (alphat) of bovine retinal G-protein, transducin. Direct binding of alphat-(340-350) to rhodopsin stabilizes the activated metarhodopsin II state (M II), consequently uncoupling the rhodopsin-transducin interaction. This peptide action requires two segments on the cytoplasmic domain of rhodopsin: the Tyr136-Val137-Val138-Val139 sequence on the C-D loop and the Glu247-Lys248-Glu249-Val250-Thr251 sequence on the E-F loop. We propose that a tertiary interaction of these two loop regions forms a pocket for binding the alphat C terminus of the transducin during light transduction in vivo. In most G-proteins, the C termini of alpha subunits are important for interaction with receptors, and, in several serpentine receptors, regions similar to those in rhodopsin are essential for G-protein activation, indicating that the interaction described here may be a generally applicable mode of G-protein binding in signal transduction.

Amino Acid Sequence↗
PubMed1997

Role of proofreading and mismatch repair in maintaining the stability of nucleotide repeats in DNA.

The role of the proofreading exonuclease in maintaining the stability of multiply repeated units in DNA was studied in Escherichia coli. Reversion of plasmids in which the beta-galactosidase alpha complementing sequence was moved +2 out of frame by inserts containing (CA)14, (CA)5, (CA)2 or (TA)6 or +1 by creating a run of 8 C was compared in mutS and mutSdnaQ strains. Proofreading corrects at least half of the frameshift errors for all the plasmids and at least 99% of the errors in the (CA)2 plasmid. The (CA)2 plasmid reverts mostly by +1 frameshifts in the restriction sites flanking the insert. With the (CA)14, (TA)6, (CA)5 and 8C plasmids, reversion is mainly by loss of a repeat unit. The data support the hypothesis that the dnaQgene product recognizes frameshifts close to the DNA growing point. Frameshifts distal to the growing point are mainly corrected by mismatch repair. We speculate that mismatches in mononucleotide repeats are susceptible to proofreading because they can either migrate to a point where they are recognized by the exonuclease or, alternatively, because single nucleotide distortions are more readily detected than dinucleotides.

Bacteriophage M13↗
PubMed1997

Cell cycle regulation of the human DNA mismatch repair genes hMSH2, hMLH1, and hPMS2.

Hereditary nonpolyposis colorectal cancer is a cancer susceptibility syndrome that has been found to be caused by mutations in any of several genes involved in DNA mismatch repair, including hMSH2, hMLH1, or hPMS2. Recent reports have suggested that hMSH2 and hMLH1 have a role in the regulation of the cell cycle. To determine if these genes are cell cycle regulated, we examined their mRNA and protein levels throughout the cell cycle in IMR-90 normal human lung fibroblasts. We demonstrate that the levels of hMSH2 mRNA and protein do not change appreciably throughout the cell cycle. Although hMLH1 mRNA levels remained constant, there was a modest (approximately 50%) increase in its protein levels during late G1 and S phase. The levels of hPMS2 mRNA fluctuated (decreasing 50% in G1 and increasing 50% in S phase), whereas hPMS2 protein levels increased 50% in late G1 and S phase. Our data indicate that, at least in normal cells, the machinery responsible for the detection and repair of mismatched DNA bases is present throughout the cell cycle.

Adenosine Triphosphatases↗
PubMed1997

A histopathological study of liver and kidney in male Wistar rats treated with subtoxic doses of t-butyl alcohol and trichloroacetic acid.

Tertiary butyl alcohol and trichloroacetic acid are known to be contaminants in drinking water. In order to evaluate the interactive toxicity of t-butyl alcohol with trichloroacetic acid, young male Wistar rats were dosed through water at a dose level of t-butyl alcohol (TBA)-0.5% (v/v), trichloroacetic acid (TCA)-25 ppm and a combined dose of TBA + TCA (0.5% v/v TBA-25 ppm TCA) for a period of 10 weeks ad libitum and were maintained on normal diet. The control animals received plain water and normal diet. The liver and kidney histology was undertaken to see whether subtoxic administration of TBA and TCA individually as well as combined administration for a period of 10 weeks would bring about any histological alterations. It was observed that TBA, TCA and TBA + TCA caused histological alterations in the liver such as centrilobular necrosis, vacuolation in hepatocytes and loss of hepatic architecture. TBA and TBA + TCA caused periportal proliferation and lymphocytic infiltration. Hypertrophy of hepatocytes in the periportal area was a characteristic feature in the liver of TCA treated rats. Moreover, in the histology of the kidney, in the three treated groups, degeneration of renal tubules, with syncitial arrangements of the nucleus of renal tubular epithelial cells was evident. In addition to this, degeneration of the basement membrane of the Bowmans capsule, diffused glomeruli and vacuolation of glomeruli was also evident in the three treated rat kidneys. Renal tubular proliferation in certain areas was also evident in certain areas of the kidney in TCA treated rats. The results indicate that, TBA and TCA do bring about alterations in histology of liver and kidney, but on combined administration, do not show enhanced toxicity in the form of increased hepatic and renal injury.

Animals↗
PubMed1997

Human MSH2 binds to trinucleotide repeat DNA structures associated with neurodegenerative diseases.

The expansion of trinucleotide repeat sequences is associated with several neurodegenerative diseases. The mechanism of this expansion is unknown but may involve slipped-strand structures where adjacent rather than perfect complementary sequences of a trinucleotide repeat become paired. Here, we have studied the interaction of the human mismatch repair protein MSH2 with slipped-strand structures formed from a triplet repeat sequence in order to address the possible role of MSH2 in trinucleotide expansion. Genomic clones of the myotonic dystrophy locus containing disease-relevant lengths of (CTG)n x (CAG)n triplet repeats were examined. We have constructed two types of slipped-strand structures by annealing complementary strands of DNA containing: (i) equal numbers of trinucleotide repeats (homoduplex slipped structures or S-DNA) or (ii) different numbers of repeats (heteroduplex slipped intermediates or SI-DNA). SI-DNAs having an excess of either CTG or CAG repeats were structurally distinct and could be separated electrophoretically and studied individually. Using a band-shift assay, the MSH2 was shown to bind to both S-DNA and SI-DNA in a structure-specific manner. The affinity of MSH2 increased with the length of the repeat sequence. Furthermore, MSH2 bound preferentially to looped-out CAG repeat sequences, implicating a strand asymmetry in MSH2 recognition. Our results are consistent with the idea that MSH2 may participate in trinucleotide repeat expansion via its role in repair and/or recombination.

Binding, Competitive↗
PubMed1997

Factors affecting the pharmacokinetics of parenteral chloramphenicol in enteric fever.

Chloramphenicol pharmacokinetics were studied in 29 Nepalese adults diagnosed with uncomplicated enteric fever and randomized to receive succinate ester 30 mg/kg i.v. or i.m. Serial plasma concentrations of chloramphenicol, and iothalamate (to estimate glomerular filtration rate), antipyrine (hepatocellular function) and Indocyanine Green (liver blood flow) were measured by HPLC and kinetic parameters estimated by non-compartmental analysis. In culture-positive patients (n = 16), mean residence times (MRTs) and steady-state volumes of distribution (V(d)ss) for i.v. chloramphenicol (mean +/- S.D.; 4.9 +/- 0.9 h and 1.9 +/- 0.8 L/kg; n = 7) were less than after i.m. chloramphenicol (12.3 +/- 7.3 h and 3.7 +/- 2.5 L/kg; n = 9; P < 0.05), with a higher peak plasma concentration after i.v. (16.2 +/- 9.1 versus 7.8 +/- 3.6 mg/L; P < 0.05); plasma clearance (Cl(p)) was similar in the two groups (368 +/- 172 and 310 +/- 224 mL/kg/min after i.v. and i.m. respectively). In 17 patients examined during convalescence, MRT and Vdss were less than in acute illness regardless of route chloramphenicol administration. There were similar changes in chloramphenicol kinetic parameters in culture-negative patients. Antipyrine Cl(p) and liver blood flow correlated weakly with chloramphenicol Cl(p) in culture-positive patients (P < 0.1) and were higher in convalescence; no such associations were seen for iothalamate Cl(p). These data indicate that i.v. chloramphenicol produces peak plasma concentrations which are on average twice those after i.m. injection of the same dose, due principally to a smaller V(d)ss. Cl(p) is uninfluenced by route of administration and is determined more by hepatic metabolism than renal excretion. Intramuscular treatment may result in sub-therapeutic chloramphenicol concentrations initially, but continued regular i.v. dosing is more likely to produce levels at which bone marrow toxicity occurs.

Adolescent↗
PubMed1997

In vivo lipid peroxidation responses of tissues in lead-treated Swiss mice.

A single intraperitoneal injection of lead acetate (200 mg/kg body weight) increased the lipid peroxidation potential (LPP) measured as thiobarbituric acid-reactive substance (TBA-RS) in different tissues of Swiss mice. All the tissues taken for experimentation, generated significantly higher amount of TBA-RS in lead-treated mice when compared with the respective control value. However, none of the tissues could correspond to the control value after the lapse of four weeks post-treatment. Possibilities of differential responsiveness of tissues to generate lipid peroxides in lead-treated mice have been discussed.

Animals↗
PubMed1996

hMSH2 forms specific mispair-binding complexes with hMSH3 and hMSH6.

The genetic and biochemical properties of three human MutS homologues, hMSH2, hMSH3, and hMSH6, have been examined. The full-length hMSH6 cDNA and genomic locus were isolated and characterized, and it was demonstrated that the hMSH6 gene consisted of 10 exons and mapped to chromosome 2p15-16. The hMSH3 cDNA was in some cases found to contain a 27-bp deletion resulting in a loss of nine amino acids, depending on the individual from which the cDNA was isolated. hMSH2, hMSH3, and hMSH6 all showed similar tissue-specific expression patterns. hMSH2 protein formed a complex with both hMSH3 and hMSH6 proteins, similar to protein complexes demonstrated by studies of the Saccharomyces cerevisiae MSH2, MSH3, and MSH6. hMSH2 was also found to form a homomultimer complex, but neither hMSH3 nor hMSH6 appear to interact with themselves or each other. Analysis of the mismatched nucleotide-binding specificity of the hMSH2-hMSH3 and hMSH2-hMSH6 protein complexes showed that they have overlapping but not identical binding specificity. These results help to explain the distribution of mutations in different mismatch-repair genes seen in hereditary nonpolyposis colon cancer.

Base Sequence↗
PubMed1996

Characterisation of a DNA pairing activity copurifying with DNA ligase in a partially purified extract from rat testis.

Rat testicular nuclear extracts were fractionated sequentially on phosphocellulose, heparin-agarose and ssDNA-cellulose columns, in order to isolate and characterise a strand-transfer activity from a mammalian meiotic tissue. A partially purified fraction, eluting at 0.6 M KCl from ssDNA-cellulose column, catalyzed the formation of two classes of products migrating slowly on an agarose gel. The formation of one of these classes of products-the aggregates-was dependent on the presence of both the substrates (M13mp19 RF III and M13mp19 ssDNA) and on homology. The presence of ATP was essential for the formation of aggregates, though its hydrolysis was not required. EM analysis of the products indicated the presence of structures which resembled paired DNA molecules: duplex-duplex paired (Y-shaped and ds-ds paired structures) and ss-ds paired (duplex DNA paired with the single-stranded DNA) structures, indicating the presence of a pairing protein in the fraction. However, alpha- and sigma-structures were not observed. The other class of products, seen as discrete bands, were identified biochemically and by electron microscopy as ligated products. A DNA ligase-adenylate adduct of molecular weight 100 kDa was formed by the fraction. Both 5' to 3' and 3' to 5' exonucleases were absent and hence did not contribute to the formation of the products.

Adenosine Triphosphate↗
PubMed1996

Modulation of GDP release from transducin by the conserved Glu134-Arg135 sequence in rhodopsin.

A superfamily of seven-transmembrane helix receptors catalyzes GDP release from heterotrimeric guanine nucleotide-binding proteins (G proteins) to initiate the intracellular signaling cascade. The photoreceptor rhodopsin is a prototypical member of the superfamily that activates the retinal G protein transducin (Gt). The cytoplasmic domain of rhodopsin binds and activates Gt, but residues that stimulate GDP release from Gt have not been identified until now. We show here that the abnormal signal transduction phenotypes of several different mutations affecting the highly conserved Glu134-Arg135 charge pair result from alteration of the GDP release step in the Gt activation cascade. We propose that Glu134 and Arg135 constitute the site that directly provides the signal from rhodopsin to activate GDP release from Gt. Because the Glu/Asp-Arg sequence occurs at a topologically identical location in most of the seven-transmembrane helix receptors, we propose that these residues constitute a switch for signal transfer.

Amino Acid Sequence↗
PubMed1996

Bilateral fractures of the capitulum. A case report.

We report a case of isolated displaced fracture of the capitulum in both elbows. Treatment was by closed manipulation and immobilisation for 3 weeks which achieved a satisfactory result, both elbows regaining 100 degrees of flexion.

Female↗
PubMed1996

The mismatch-repair protein hMSH2 binds selectively to DNA adducts of the anticancer drug cisplatin.

BACKGROUND: The antitumor drug cis-diamminedichloroplatinum(II) (cis-DDP or cisplatin) exerts its cytotoxic effects through the formation of covalent DNA adducts. A family of proteins possessing a common HMG box motif that binds specifically to cisplatin DNA adducts has been previously suggested to be important in the clinical efficacy of the drug. RESULTS: We have shown that the human mismatch-repair protein, hMSH2, also binds specifically to DNA containing cisplatin adducts and displays selectivity for the DNA adducts of therapeutically active platinum complexes. Moreover, hMSH2 is overexpressed in testicular and ovarian tissue; tumors in these tissues are most effectively treated by cisplatin. CONCLUSIONS: Our results suggest a role for hMSH2 in mediating cisplatin toxicity. Supporting this view, previous studies in Escherichia coli dam- strains demonstrate that mutations in mismatch-repair proteins confer resistance to cisplatin toxicity. Mismatch-repair deficiency is also correlated with tolerance to O6-methylguanine, a cytotoxic DNA lesion formed by methylating agents. A current model ascribes O6-methylguanine toxicity to unsuccessful attempts at repair of this lesion by mismatch-repair proteins, resulting in a futile cycle of incision and synthesis, leading ultimately to lethal DNA-strand breaks. We propose that mismatch repair may contribute to cisplatin toxicity by a similar mechanism. Alternatively, hMSH2 may shield cisplatin adducts from repair, allowing adducts to persist, thus enhancing lethality.

Antineoplastic Agents↗
PubMed1995

Administration of subtoxic doses of t-butyl alcohol and trichloroacetic acid to male Wistar rats to study the interactive toxicity.

Tertiary butyl alcohol and trichloroacetic acid (TCA) are known to be contaminants in drinking water. In order to evaluate the interactive toxicity of t-butyl alcohol (TBA) with TCA, young male Wistar rats were dosed through water at a dose level of TBA (0.5% v/v), 25 ppm TCA and a combined dose of TBA+TCA (0.5% v/v TBA, 25 ppm TCA) for a period of 10 weeks ad libitum and were maintained on normal diet. The control animals received plain water and normal diet. There was remarkable loss of body weight and significantly decreased liver triglycerides in the treatment groups in the order of TBA+TCA, TCA, TBA and increased liver weights were observed. Serum succinate dehydrogenase (SDH) levels were significantly increased in TCA- and TBA+TCA-treated groups. There was no significant change in serum alanine (GPT), aspartate (GOT) aminotransferase, serum alkaline (ALP) and acid (ACP) phosphatase levels as well as liver glutathione (GSH) and liver and serum cholesterol levels in the treated groups. But serum triglycerides, liver glycogen, serum glucose (only in TBA- and TCA-treated animals) were significantly high in the treated groups. Lipid peroxidation measured by diene conjugation was significant in TBA+TCA-treated group and kidney GSH levels were significantly low in the treated groups. These results show that interaction of TBA+TCA does bring about alteration in biochemical parameters which may play a pivotal role in toxic responses on long-term exposure.

Alanine Transaminase↗
PubMed1995

Efficacy of dexamethasone pulse therapy in progressive systemic sclerosis.

BACKGROUND: Systemic sclerosis is a disease of unknown etiology for which no specific treatment is effective. Pulse therapy with corticosteroids has been tried for various autoimmune disorders with minimal side effects. We undertook this study to determine the efficacy of dexamethasone pulse therapy in progressive systemic sclerosis (PSS). METHODS: Five women with PSS between the ages 30 and 60 years, received 100 mg dexamethasone in 500 mL of 5% dextrose by slow intravenous infusion over 3 hours for 3 consecutive days, once a month. RESULTS: All patients had symptomatic and clinical improvement. The vital capacity improved in three and posttreatment histopathologic regression was seen in two patients. CONCLUSIONS: Dexamethasone pulse therapy may provide an additional option for treating systemic sclerosis.

Adult↗