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Biomedical subjects

C Guha

Publications and source records attributed to C Guha.

26 records · Page 2Linked to original sources
PubMed1995

Regulation of human Gc (vitamin D--binding) protein levels: hormonal and cytokine control of gene expression in vitro.

Studies were performed in Hep3B hepatocytes to better elucidate the mechanisms regulating circulating levels of human group-specific component (Gc). We measured changes in Gc messenger RNA (mRNA) synthesis and levels of secreted protein resulting from treatment of hepatocytes with cytokines and hormones known to influence synthesis of other proteins of hepatic origin. We particularly focused on compounds known to be prototypic stimulants during the acute phase response. Interleukin-6 (IL-6) and dexamethasone were shown to increase Gc mRNA approximately twofold while transforming growth factor beta (TGF beta) decreased Gc mRNA in a dose-dependent fashion by up to fivefold. The effects on secreted Gc protein levels were similar. These results indicate that Gc protein appears to be regulated differently than the other members of this gene family, albumin and alpha-fetoprotein (AFP), which are negative acute phase reactants. In addition, these contrasting effects on Gc synthesis of IL-6 and dexamethasone and of TGF beta suggest that high basal levels of Gc synthesis may be maintained during the acute phase response.

Actins↗
PubMed1993

The vitamin D-binding protein gene: quantitation of amplified nucleic acids by ELISA.

We have utilized enzyme-linked immunosorbent assay (ELISA) to quantitate PCR-amplified DNA. This method was used to measure mRNA for the vitamin D-binding protein (Gc), beta-actin and the transferrin receptor (TR) gene in the Hep3B cell line. Total RNA from Hep3B cells was reverse transcribed to obtain cDNA, which was amplified in the presence of digoxigenin-dUTP by PCR. The PCR products were then hybridized in liquid phase to a biotinylated, nested capture probe for the respective sequences. The hybridized products were bound to a streptavidin-coated ELISA plate and were detected by an alkaline-phosphatase-conjugated antibody to digoxigenin. ELISA standard curves for Gc and control genes, beta-actin and TR, were obtained after PCR amplification of serial dilutions of Hep3B total RNA. As an external standard, an ELISA standard curve for Gc was obtained after PCR amplification of serial dilutions of a full-length Gc cDNA insert obtained from a recombinant plasmid. Thus, we were able to develop a non-isotopic quantitation assay for PCR-amplified DNA that is highly sensitive and has the specificity of hybridization-based methods.

Actins↗
PubMed1981

RNA synthesis and degradation during preferential inhibition of protein synthesis by cobalt chloride in Escherichia coli K-12.

It has been observed that growth of Escherichia coli cells are inhibited when treated with cobalt chloride (300 microM). It has also been shown that CoCl2 preferentially inhibits translation without inhibiting the process of transcription (1, 2, 8). We report here, that during treatment of E. coli cells with CoCl2, both messenger RNA and stable RNA synthesis is slowed down about 2.5 folds. The rate of degradation of mRNA also decreases and both chemical and functional half-life of mRNA increases about 2.5 folds in Co-treated cells. This clearly shows that the process of transcription is also affected while translation is preferentially inhibited during CoCl2 treatment.

Cobalt↗
PubMed1981

A ribonuclease-resistant cytoplasmic 10 S ribonucleoprotein of chick embryonic muscle. A potent inhibitor of cell-free protein synthesis.

A ribonucleoprotein (RNP) particle sedimenting at 10 S in sucrose gradients had been isolated from the post-polysomal fraction of homogenates of 14-day-old chick embryonic leg and breast muscle by sucrose gradient fractionation and gel filtration. The 10 S RNP contains a 4 S RNA species (base composition: AMP, .3%; GMP, 22.2%; CMP, 24.2%; and UMP, 23.2%), and shows three major bands in the 70-90-nucleotide size range by polyacrylamide gel electrophoresis in 99% formamide. The 4 S RNA does not contain oligo(U)- and oligo(A)-rich tracts. The RNP has a characteristic buoyant density of 1.410 g/ml, which corresponds to an RNA/protein ratio of about 1:4. The UV absorption spectra of the RNP is very distinct from that of its RNA component. Both 4 S RNA and the 10 S RNP are potent inhibitors of translation of a variety of mRNAs such as chick muscle poly(A)+ mRNA, rabbit globin mRNA, EMC virus RNA, and poly(A)- and mRNA of rat liver in micrococcal nuclease-treated rabbit reticulocyte lysate. The inhibitory action of the RNA and the RNP on mRNA translation appears to involve the initiation process. The RNA and RNP do not have a nuclease activity associated with them. The hyperchromicity profile of the inhibitory RNA with increasing temperature indicates that it does not contain a significant amount of double-stranded structure. This is also supported by the complete loss of biological activity of the RNA by treatment with pancreatic RNase. In contrast, the inhibitory activity of the RNP was resistant to RNase. Electrophoresis of the protein moieties of the inhibitory RNP using both one- and two-dimensional gel techniques in the presence of sodium dodecyl sulfate shows a complex pattern of polypeptides of Mr = 12,000-150,000. The protein pattern of the 10 S particle is quite different from those of free and polysomal mRNP and poly(A)-protein complexes of chick embryonic muscles, indicating that most, if not all of the mRNA-associated proteins, are absent in the 19 S RNP. The properties of the inhibitory RNA indicate that it is different from the various low molecular weight RNA species which are involved in the modulation of protein synthesis in cell-free systems. It is concluded that the 10 S particle represents a novel class of RNP, which may be involved in posttranscriptional regulation of protein synthesis in embryonic muscles.

Animals↗
PubMed1980

Non-coordinate regulation of RNA synthesis in Escherichia coli exposed to 0 degrees C.

A non-coordinate mode of regulation of RNA synthesis is observed in Escherichia coli cells during exposure to 0 degrees C. The stable RNA synthesis is preferentially inhibited with simultaneous accumulation of messenger RNA. The species of RNA synthesized at 0 degrees C was determined by several criteria such as sedimentation value in sucrose gradients, DNA-RNA hybridization, half life measurements, protein synthesizing capacity and its functional rate of decay. The mode of regulation of RNA synthesis at 0 degrees C is unique and is distinct from the non-coordinate regulation observed during amino acid starvation under stringent control.

Bacterial Proteins↗
PubMed1979

A model of translational control involving mRNA-associated proteins in chick embryonic muscles.

Highly purified poly(A)-containing free and polysomal mRNP particles have been isolated by chromatography of subcellular fractions of chick embryonic muscles on oligo-dT-cellulose and elution with low salt buffer at 45 degrees. The free and polysomal mRNP represent two distinct classes of macromolecules, the free particles having a more complex nucleoprotein organization than the polysomal particles. Comparison of the protein moieties of three classes of poly(A)-containing cytoplasmic mRNP -- those released from nuclei after in vitro transcription and processing (transported mRNP), the free, and polysomal mRNP -- strongly suggests that the majority of the mRNA-associated proteins are exchanged in the cytoplasm during the various functional states of mRNA. A model of translational control involving the participation of mRNA-associated proteins in chick embryonic muscles and by analogy in other differentiated eukaryotic cells is proposed.

Animals↗