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G An

Publications and source records attributed to G An.

At least 109 records · Page 6Linked to original sources
PubMed1992

Effect of deletions 5' to the translation initiation sequence on the expression of an mRNA in animal cells.

To learn if an mRNA.18S rRNA interaction or a special secondary structure in the mRNA start region is essential for translation in eukaryotic cells, we constructed recombinant plasmids with the SV40 early promoter 5' to part of the Escherichia coli tufB-lacZ gene. Deletion of bases potentially complementary to the 18S rRNA highly increased the transient beta-galactosidase expressed in transfected CHO cells. Deletion of bases that fostered formation of potential hairpins with the mRNA 5'-terminus or altered the structure of the coding region reduced beta-galactosidase activity suggesting that these features of the mRNA secondary structure may be essential for initiation of translation. Computer aided analysis of the potential structure of 290 mRNAs suggests these are conserved features of the initiation region.

Animals↗
PubMed1992

Pollen-specific expression of the Arabidopsis thaliana alpha 1-tubulin promoter assayed by beta-glucuronidase, chloramphenicol acetyltransferase and diphtheria toxin reporter genes.

We have characterized the promoter specificity of the Arabidopsis thaliana alpha 1-tubulin (alpha 1-tub) gene by studying expression patterns of gene fusions between the 2.2 kbp 5' upstream region of the alpha 1-tub gene and each of three different reporters: chloramphenical acetyltransferase, beta-glucuronidase or the diphtheria toxin chain A gene. Analysis of transgenic tobacco and Arabidopsis plants carrying the transgene showed that the chloramphenicol acetyltransferase and beta-glucuronidase activities were not detected in any vegetative or reproductive organs except mature pollen. Transgenic tobacco plants carrying the diphtheria toxin chain A gene under the control of the alpha 1-tub promoter were of normal phenotype but seed fertility was drastically reduced. Furthermore, the transgene could not be transmitted to the next generation through pollen, supporting the observation that the alpha 1-tub promoter is active only in pollen. It was observed that the promoter activity was most active in mature pollen and decreased significantly during in vitro pollen germination, indicating that the promoter is inactive or subdued in germinating pollen. The promoter activity was not affected by various plant growth hormones during pollen maturation.

Arabidopsis↗
PubMed1992

Inhibition of Sucrose Enhancer Effect of the Potato Proteinase Inhibitor II Promoter by Salicylic Acid.

Effect of salicylic acid (SA) on the expression of the potato proteinase inhibitor (PI) II promoter was studied with transgenic tobacco plants (Nicotiana tabacum) carrying a gene fusion between the PI-II promoter and the chloramphenicol acetyltransferase (cat) reporter. As previously observed, the PI-II promoter was inducible by wounding and the promoter activity was further enhanced by sucrose. Addition of SA did not influence the wound induction of the PI-II promoter but significantly inhibited the sucrose response. The 5'-deletion mutant -573 was unable to respond to wounding but did respond to sucrose and SA. The 3'-deletion analysis indicated the presence of a sucrose-responsive element between -574 and -520. A study of the insertion mutants revealed the function of another sucrose-responsive element between -522 and -500. Enhancer effects of these sucrose-responsive elements were inhibited by SA. These studies suggest that SA inhibits PI-II promoter activity by decreasing the sucrose response. Analysis of SA-related chemicals revealed that only acetyl-SA showed a similar inhibitory effect, and other hydroxybenzoic acids had little or no effect on the sucrose enhancer activity. Therefore, it seems that the interaction between SA and the receptor molecule is specific.

Journal Article↗
PubMed1992

Identification of G-Box Sequence as an Essential Element for Methyl Jasmonate Response of Potato Proteinase Inhibitor II Promoter.

The potato proteinase inhibitor II promoter was studied to identify cis-acting regulatory sequences involved in methyl jasmonate (MJ) response using transgenic tobacco plants carrying various lengths of the promoter fused to a chloramphenicol acetyltransferase reporter gene. An internal fragment between -625 and -520 was sufficient to confer a response to MJ, wounding, or sucrose when it was placed upstream of the nos promoter -101, which contains the CAAT-TATA region. Deletion of the proteinase inhibitor II promoter sequence upstream of -611 did not affect the MJ response, but a further deletion to -573 eliminated the response. The 3'-deletion study showed that the DNA sequence downstream from -520 is dispensable. However, 3'-deletion mutant -574 did not respond to the MJ treatment. These results indicated that an element essential for the MJ response is located at the -574/-573 region where the G-box sequence (CACGTGG) is located. The G-box sequence was not required for the sucrose enhancer effect, suggesting that the MJ response mechanism is different from that of sucrose.

Journal Article↗
PubMed1992

An unusual expression of a squamous cell marker, small proline-rich protein gene, in tracheobronchial epithelium: differential regulation and gene mapping.

An unusual expression of a putative squamous cell marker, small proline-rich protein (spr1), in mucociliary epithelial cells of conducting airways was demonstrated in a serum-free culture system. A cDNA clone was isolated from the cDNA library of monkey tracheobronchial epithelial (TBE) cells by differential hybridization. This cDNA clone, MT5, exhibited 98% homology to a DNA sequence obtained from human keratinocytes treated with either UV light or phorbol esters (T. Kartasova et al., 1988, Mol. Cell. Biol. 8:2195-2230). The predicted peptide of MT5 is unusual for its high content of proline (29%), glutamine (18%), and cysteine (9%) and its repeated PKVPEPC units. The level of spr1 mRNA in cultured cells was inhibited more than 90% by vitamin A. In contrast, phorbol 12-myristate 13-acetate (PMA) stimulated the level of spr1 mRNA by 3- to 8-fold. This differential regulation coincided with the effects of these chemicals on the cornification of cultured TBE cells. Using MT5 as a probe, we have localized the tracheal spr1 gene on the human chromosome 1 by a Southern blot analysis using a panel of human-rodent somatic cell hybrid DNAs. The gene was further sublocalized to bands q22-23 by in situ hybridization.

Amino Acid Sequence↗
PubMed1991

Structural and functional analysis of promoter from gliadin, an endosperm-specific storage protein gene of Triticum aestivum L.

To identify cis-regulatory elements of the gliadin gene, a study of the gliadin gene promoter was conducted by transient expression analysis of plasmid DNAs which were introduced into plant protoplasts by electroporation. The promoter region (-592 bp to +18 bp from the translational start) of this developmentally regulated gene, when fused upstream to the chloramphenicol acetyl transferase (CAT) reporter cassette was unable to direct significant CAT expression in wheat or tobacco suspension cells. Because this monocot gene promoter appeared to be under stringent tissue-specific control, a hybrid promoter approach using a nopaline synthase (nos) promoter was employed. A series of 3' deletions of the gliadin promoter were placed upstream of either a nonfunctional -101 nos or a nearly wild-type -155 nos promoter fused in turn to a CAT reporter gene cassette. Transient expression analysis of these plasmid DNAs in tobacco cells showed that the gliadin fragment could either restore the activity of the non-functional nos promoter (series I) or enhance the activity of the functional nos promoter (series II). The degree of restoration of the promoter function conferred by gliadin fragments of the first series was proportional to the enhancing effect of the same fragments in the second series of constructs. The transcriptional activity of the gliadin (-592 bp to -77 bp) -nos hybrid promoter was reduced by 26% upon 3' deletion of sequences in the region -141 bp to -77 bp, which contains both the TATA and CCAAT boxes.(ABSTRACT TRUNCATED AT 250 WORDS)

Base Sequence↗
PubMed1991

Expression of DNA coding for diphtheria toxin chain a is toxic to plant cells.

DNA coding for the enzymatically active subunit A of diphtheria toxin was placed under the control of the cauliflower mosaic virus 35S promoter and the Agrobacterium left transfer-DNA gene 7 polyadenylation signal. Agrobacteria carrying a binary plant vector with the chimeric diphtheria toxin A gene had very low transforming activity in tobacco (Nicotiana tabacum L.), and greatly diminished the recovery of stable transformants when mixed together with agrobacteria which alone transformed plant cells well. The introduction of this chimeric molecule into tobacco cells by electroporation lowered the level of the transient expression of the coelectroporated chloramphenicol acetyltransferase reporter gene indicating that expression of diphtheria toxin chain A in plant cells is toxic. We have developed a binary vector pGA987 which can be used for probing a variety of plant promoters.

Journal Article↗
PubMed1991

[Transcutaneous PO2 monitoring during cervico-thoracic epidural analgesia].

The measurement of transcutaneous oxygen tension (PtcO2) in 19 patients during cervico-thoracic epidural block with 1 percent lidocaine is reported. The data demonstrated close correlations between PtcO2 and PaO2 levels. PtcO2 in about 50 percent of the patients showed significant decreases approximately 25 minutes after epidural administration of lidocaine. Complaints of dyspnea were unrelated to PtcO2 and PaO2 levels. The authors suggest that cervico-thoracic epidural block even under low concentrations of local anesthetic (1 percent lidocaine) may depress the respiratory function after a transient interval. Great attention should be paid to the management of respiratory function during this period of time.

Adolescent↗
PubMed1990

Induction in the expression of an unusual proline-rich protein by pig tracheal surface epithelial cells maintained in primary culture.

Primary cell culture is a valuable tool for studying the regulation of gene expression since many differentiated traits are conserved. Cells cultured from the epithelial lining of pig trachea were selected as a model system for mucin synthesis. RNAs were isolated from pig trachea epithelial linings and from pig trachea surface epithelial cells cultured in serum-free media. Cell free translations showed an unusually high incorporation of [3H]proline into a relatively small protein (about 20 kDa), but only with RNA from the cells in culture. RNA prepared from pig trachea cells immediately before placing the cells in culture (day 0) did not contain mRNA encoding this unusual proline-rich protein. However, the expression of this protein was dramatically induced within 2 days of maintaining the cells in culture.

Animals↗
PubMed1990

Production of a foreign protein product with genetically modified plant cells.

Plant cells (Nicotiana tabacum) were genetically engineered to produce a foreign protein, chloramphenicol acetyltransferase (CAT), and the CAT production from suspension cultures was investigated. Suspension cultures were grown in a shake flask, a stirred fermenter, and a bubble-column fermenter. The CAT production was growth related and the maximum activity was reached during the early stationary phase. A 41-day, semicontinuous stirred fermenter run, consisting of five sequential batch runs, demonstrated long-term CAT production. Continuous CAT production was also accomplished in a bubble-column fermenter at a medium flow rate of 3.1 ml h-1, which was equivalent to a dilution rate of 0.25 day-1.

Cells, Cultured↗
PubMed1990

Wound-inducible nuclear protein binds DNA fragments that regulate a proteinase inhibitor II gene from potato.

Deletion analysis from the 3' to the 5' end of the promoter region of the wound-inducible potato proteinase inhibitor IIK gene has identified a 421-base sequence at -136 to -557 that is necessary for expression. Utilizing DNA band-shift assays, a 10-base sequence within the 421-base region was found to bind a nuclear protein from wounded tomato leaves. This 10-base sequence is adjacent to an 8-base consensus sequence at -147 to -155 that is present in the promoter region of several elicitor-inducible genes from various other plants. The evidence suggests that a complex set of cis- and trans-acting elements within the -136 to -165 region of the potato IIK gene may be involved with the signaling mechanisms that regulate the inducibility of this gene in response to pest and pathogen attacks.

Base Sequence↗
PubMed1990

Nopaline synthase promoter is wound inducible and auxin inducible.

The activity of the nopaline synthase (nos) promoter is differentially regulated in several plant organs. In this article we demonstrate that the nos promoter is wound inducible in both vegetative and reproductive organs. The induction of the nos promoter was observed in leaves, stems, cotyledons, and various reproductive organs, suggesting that the response is not organ specific. The wound response was further enhanced by addition of auxins. Other growth substances had no effect on the wound-inducible nos promoter activity. Deletion analysis of the nos promoter indicated that the 10-base pair (GCACATACGT) Z element located between -123 and -114 or an element overlapping with this sequence is essential for the wound and auxin responses.

Amino Acid Oxidoreductases↗
PubMed1989

Three distinct regulatory elements comprise the upstream promoter region of the nopaline synthase gene.

Fine deletion mutants were generated in the upstream control region of the nopaline synthase (nos) promoter to define the position and role of upstream regulatory elements. The results indicated that the 8 bp sequence (CAGAAACC) at -106/-113 and its inverted repeat (GGTTTCTG) at -140/-147 are important for promoter function. The downstream element appears more important than the upstream element since deletion of the former reduced promoter activity more significantly than deletion of the latter. Deletion of the element alone, however, did not abolish promoter function, whereas, deletion of the 10 bp potential Z-DNA-forming (Z) element located between the repeat elements nullified promoter activity. Therefore, it appears that the Z element is an essential upstream regulator and the repeated elements are upstream modulators of the nos promoter. These elements are functionally distinct since alteration of stereospecificity or insertion of short oligonucleotides between the elements did not significantly influence promoter activity. These regulatory elements were unable to function from 200 bp upstream of the CCAAT-TATA box region.

Amino Acid Oxidoreductases↗
PubMed1989

Expression of proteinase inhibitors I and II in transgenic tobacco plants: effects on natural defense against Manduca sexta larvae.

Genes containing the cauliflower mosaic virus 35S promoter fused to open reading frames coding for tomato proteinase inhibitor I, tomato inhibitor II, and potato inhibitor II were expressed in transgenic tobacco plants. Inhibitor I and II proteins were identified by immunoblotting and quantified by immunoradial diffusion. Both inhibitors exhibited the molecular weights found for the native proteins in their natural environments. Extracts of leaves from transformed plants contained inhibitory activities against trypsin and chymotrypsin that reflected the levels of inhibitor I or II protein present. The results demonstrate that in tobacco leaves the introns of both inhibitor I and inhibitor II genes were excised correctly and that pre and prepro inhibitor I and II proteins were correctly processed. Growth of Manduca sexta larvae (tobacco hornworms) feeding on leaves of transgenic plants containing inhibitor II, a powerful inhibitor of both trypsin and chymotrypsin, was significantly retarded, compared to growth of larvae fed untransformed leaves. Levels of inhibitor II protein as low as 50 micrograms/g of tissue moderately affected larval growth, whereas levels above 100 micrograms/g severely reduced growth. The presence of tomato inhibitor I protein, a potent inhibitor of chymotrypsin but a weak inhibitor of trypsin, in transgenic tobacco leaves had little effect on the growth of the larvae. These experiments indicated that trypsin inhibitory activity, but not chymotrypsin inhibitory activity, was mainly responsible for the inhibition of larval growth.

Animals↗
PubMed1989

Functional analysis of the 3' control region of the potato wound-inducible proteinase inhibitor II gene.

Proteinase inhibitor genes are expressed strongly in specific plant tissues under both developmental and environmental regulation. We have studied the role of the 3' control region of the potato proteinase inhibitor II gene (PI-II) that is inducible in leaves in response to herbivore attacks or other severe wounding. Comparison of the terminator from the PI-II gene with two different terminators from the 6b and 7 genes, driven by a common PI-II promoter-cat fusion molecule, indicated that the PI-II terminator provided the most efficient expression of cat. The PI-II terminator also caused a significantly elevated cat gene expression driven by the cauliflower mosaic virus 35S promoter. The increase in the level of expression is probably not due to the presence of an enhancer element in the PI-II terminator region, but to cis-acting elements involved in mRNA processing or stability. Both transient and stable transformation analyses of the deletion mutants in the 3'-flanking sequence indicated that about a 100-base pair DNA fragment surrounding the polyadenylation site is essential for the efficient gene expression. This region seems to consist of several regulatory elements, including the conserved sequence, CGTGTCTT, which is located 9 bases downstream from the polyadenylation site. The elements appear to contribute to the increased stability of mRNAs containing the PI-II terminator.

Amino Acid Sequence↗