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L Gao

Publications and source records attributed to L Gao.

At least 19 recordsLinked to original sources
PubMed1998

Mutations in the Exo III motif of the herpes simplex virus DNA polymerase gene can confer altered drug sensitivities.

Two herpes simplex virus mutants containing mutated residues within the conserved Exo III motif of the polymerase gene were previously shown to be defective in 3'-5' exonuclease activity and exhibited extremely high mutation frequencies. In this study, we have shown that these mutants also exhibited higher resistance to phosphonoacetic acid and sensitivity to aphidicolin and all nucleoside analogs tested, including acyclovir and gangciclovir, compared to wild-type virus. Marker transfer experiments and sequencing analyses demonstrated that these altered phenotypes were the result of mutations within the Exo III motif. The data indicate that, aside from leading to exonuclease deficiency, mutations in the Exo III motif may also affect interaction of nucleoside triphosphates with the catalytic sites of polymerase activity.

Amino Acid Sequence↗
PubMed1998

A novel response to dioxin. Induction of ecto-ATPase gene expression.

We used differential display to discover a new gene that the environmental contaminant 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) regulates in mouse hepatoma cells. Its predicted amino acid sequence suggests that the gene encodes an ecto-ATPase that contains multiple glycosylation sites, conserved cysteine residues, and apyrase conserved regions. cDNA expression experiments in mouse hepatoma cells confirm that the new gene encodes an ecto-ATPase. Wild-type mouse hepatoma cells contain both constitutive and TCDD-inducible ecto-ATPase activity. Induction of ecto-ATPase gene expression by TCDD is direct and occurs at the transcriptional level. Studies in mutant hepatoma cells indicate that induction requires both the aromatic hydrocarbon receptor (AhR) and the AhR nuclear translocator (Arnt). Furthermore, induction requires AhR's transactivation domain, but not that of Arnt. Our findings reveal new aspects of dioxin's biological effects and TCDD-dependent gene regulation.

Adenosine Triphosphatases↗
PubMed1998

Isolation and functional analysis of the promoter of the rat CMP-NeuAc:GM3 alpha2,8 sialyltransferase gene 1.

A 2.1-kb 5'-flanking fragment of the rat CMP-NeuAc:GM3 alpha2,8 sialyltransferase (GD3-synthase) gene was cloned by the genomic walking procedure. The promoter activity of the fragment was assessed in F-11 cells by transient transfection and the locations for the basal and maximal promoter activities were defined. Primer extension analysis identified a transcription start site approximately 98 bp upstream of the ATG start codon. DNA sequence analysis of the promoter revealed a number of consensus binding sites for known transcription factors such as SP1, AP1, NFkappaB, C/EBP and TFIID, and a repeat GC-GT sequence motif seen for the formation of Z-type DNA. Both TATA and CCAAT boxes were not found in the promoter. Our results from deletion constructs suggested that both positive and negative cis-acting regulatory regions were present in this TATA-less promoter of the rat GD3-synthase gene.

Animals↗
PubMed1998

Identification of a two EF-hand Ca2+ binding domain in lobster skeletal muscle ryanodine receptor/Ca2+ release channel.

The lobster skeletal muscle Ca2+ release channel, known also as the ryanodine receptor, is composed of four polypeptides of approximately 5000 amino acids each, like its mammalian counterparts. Clones encoding the carboxy-terminal region of the lobster ryanodine receptor were isolated from a lobster skeletal muscle cDNA library. Analysis of the deduced 1513 carboxy-terminal amino acid sequence suggests a cytoplasmic Ca2+ binding domain consisting of two EF-hand Ca2+ binding motifs (amino acid residues 594-656). The Ca2+ binding properties of this domain were assessed by preparing bacterial fusion proteins with sequences from the lobster Ca2+ binding domain and the corresponding sequences of the rabbit cardiac and skeletal muscle ryanodine receptors. The lobster skeletal muscle fusion protein bound 45Ca2+ in Ca2+ overlays, and bound two Ca2+ under equilibrium binding conditions with a Hill dissociation constant (KH) of 0.9 mM and coefficient (nH) of 1.4. Rabbit skeletal and cardiac fusion proteins bound two Ca2+ with KHs of 3.7 and 3.8 mM and nHs of 1.1 and 1.3, respectively. Similar to results previously reported for the mammalian RyRs, the lobster RyR was activated by micromolar Ca2+ and inhibited by millimolar Ca2+, as determined in single-channel and [3H]ryanodine binding measurements. These results suggest that the two EF-hand Ca2+ binding domain of the lobster Ca2+ release channel as well as the corresponding regions of the mammalian channels may play a role in Ca2+ inactivation of sarcoplasmic reticulum Ca2+ release.

Amino Acid Sequence↗
PubMed1998

Characterization and biological significance of immunosuppressive peptide D2702.75-84(E --> V) binding protein. Isolation of heme oxygenase-1.

This is the first report on peptidic inhibitors of heme oxygenase. Such peptides were originally developed from the immunomodulatory peptide 2702.75-84 which corresponds to amino acid residues 75 to 84 of the alpha1-helix of HLA-B2702 (2702.75-84) and has been shown to be immunosuppressive in vitro and in vivo. In vitro, 2702.75-84 inhibited cytotoxic T- and natural killer cell- mediated target cell lysis, and in vivo peptide therapy resulted in prolongation of heart and skin allograft survival in mice. The peptide was also shown to bind to heat shock protein 70. However, D-enantiomers of 2702.75-84 and derivatives thereof, while still being immunosuppressive, did not bind to heat shock protein 70. This study was designed to identify proteins binding to peptide D2702.75-84(E --> V) (rvnlrialry) consisting of D-amino acids. Compared with 2702.75-84 (RENLRIALRY), glutamic acid residue 76 (E) was replaced with valine (V). Affinity chromatography using immobilized D2702.75-84(E --> V) and mouse and human cell extracts, resulted in the isolation of heme oxygenase-1 (HO-1). Peptide D2702.75-84 inhibited HO activity in vitro in a dose dependent manner. Similar to what has been observed with other inhibitors of HO, administration of peptide into mice resulted in an up-regulation of HO-1 mRNA and protein, as well as enzyme activity in liver, spleen and kidney. Other peptides derived from 2702.75-84 with similar immunomodulatory activity displayed similar effects. In contrast, inactive derivatives of 2702.75-84 had no effect on HO activity. Therefore, the immunosuppressive effects of the described immunomodulatory peptides are similar to those of cobalt-protoporphyrin, a known up-regulator of HO-1. Our results suggest that HO-1 modulation may be a novel mechanism of immunomodulation.

Adjuvants, Immunologic↗
PubMed1998

Stress protein-induced immunosuppression: inhibition of cellular immune effector functions following overexpression of haem oxygenase (HSP 32).

This is the first report on suppression of immune effector functions following upregulation of heat shock protein 32 (HSP 32), known as haem oxygenase (HO-1). Here we evaluated the effect of cobalt-protoporphyrin (CoPP)-induced HO-1 expression on cell-mediated immune responses. Administration of CoPP to CBA mice resulted in overexpression of HO-1 in the spleen, liver and kidneys. In vitro measurements of T cell-mediated and NK-cell-mediated cytotoxicity in spleens from CoPP-treated animals demonstrated a severe suppression of their effector functions while administration of Zn-PP or vitamin B12 had no effect. Furthermore, CoPP therapy decreased the lymphoproliferative alloresponse and differentiation of cytotoxic T cells. Inhibition of proliferation appeared to be due to cell growth arrest with an increased number of cells staying in G0/G1 phase. Despite the suppressed proliferative response, IL-2 production in the MLR was not inhibited. In contrast, CoPP decreased the production of IL-10, IFN-gamma and TNF-alpha. In vivo, CoPP prolonged the survival of heterotopic heart allografts in mice. The immunosuppressive effects following CoPP-mediated upregulation of HO-1 were similar to those observed after peptide-mediated upregulation of HO-1. The results indicate that overexpression of HO results in the inhibition of several immune effector functions and thus provides an explanation for stress-induced immunosuppression.

Animals↗
PubMed1998

Abdominal image segmentation using three-dimensional deformable models.

RATIONALE AND OBJECTIVES: The authors develop a three-dimensional (3-D) deformable surface model-based segmentation scheme for abdominal computed tomography (CT) image segmentation. METHODS: A parameterized 3-D surface model was developed to represent the human abdominal organs. An energy function defined on the direction of the image gradient and the surface normal of the deformable model was introduced to measure the match between the model and image data. A conjugate gradient algorithm was adapted to the minimization of the energy function. RESULTS: Test results for synthetic images showed that the incorporation of surface directional information improved the results over those using only the magnitude of the image gradient. The algorithm was tested on 21 CT datasets. Of the 21 cases tested, 11 were evaluated visually by a radiologist and the results were judged to be without noticeable error. The other 10 were evaluated over a distance function. The average distance was less than 1 voxel. CONCLUSIONS: The deformable model-based segmentation scheme produces robust and acceptable outputs on abdominal CT images.

Algorithms↗
PubMed1998

Regions of human immunodeficiency virus type 1 nef required for function in vivo.

In vivo studies in monkeys and humans have indicated that immunodeficiency viruses with Nef deleted are nonpathogenic in immunocompetent hosts, and this has motivated a search for live attenuated vaccine candidates. However, the mechanisms of action of Nef remain elusive. To define the regions of human immunodeficiency virus type 1 (HIV-1) Nef which mediate in vivo pathogenicity, a series of mutated isogenic viruses were inoculated into human thymic implants in SCID-hu mice. Mutation of several regions, including the myristoylation site at the second glycine and a region encompassing amino acids 41 through 49 of Nef, profoundly affected pathogenicity. Surprisingly, mutations of prolines in either of the two distant PXXP SH3 binding domains did not affect pathogenicity, indicating that these regions are not required for Nef activity in developing T-lineage cells. These data suggest that some functions of Nef described in vitro may not be relevant for in vivo pathogenicity.

Animals↗
PubMed1998

gamma-glutamylcysteine synthetase: mRNA stabilization and independent subunit transcription by 4-hydroxy-2-nonenal.

gamma-Glutamylcysteine synthetase (GCS), the rate-limiting enzyme in de novo glutathione (GSH) synthesis, is composed of one catalytic (heavy) and one regulatory (light) subunit. Although both subunits are increased at the mRNA level by oxidants, it is not clear whether they are regulated through the same mechanism. 4-Hydroxy-2-nonenal (4HNE), a lipid peroxidation product, may act as a mediator for the induction of gene expression by oxidants. In the present study, 4HNE was used to study the mechanism of induction of the two GCS subunits in rat lung epithelial L2 cells. 4HNE increased both the transcription rates and the stability of mRNA for both GCS subunits, resulting in an increased mRNA content for both subunits. Both GCS subunit proteins and enzymatic activities also increased. Emetine, a protein synthesis inhibitor, blocked the increase in GCS light subunit mRNA but not the increase in GCS heavy subunit mRNA. This suggested that although 4HNE increased transcription and stabilization of both GCS subunit mRNAs, the signaling pathways involved in the induction of the two GCS subunits differed.

Aldehydes↗
PubMed1997

Human genes encoding U3 snRNA associate with coiled bodies in interphase cells and are clustered on chromosome 17p11.2 in a complex inverted repeat structure.

Coiled bodies (CBs) are nuclear organelles whose morphological structure and molecular composition have been conserved from plants to animals. Furthermore, CBs are often found to co-localize with specific DNA loci in both mammalian somatic nuclei and amphibian oocytes. Much as rDNA sequences are called nucleolus organizers, we term these coiled body-associated sequences 'coiled body organizers' (CBORs). The only sequences that have been shown to be CBORs in human cells are the U1, U2 and histone gene loci. We wanted to determine whether other snRNA genes might also act as CBORs. In this paper we show that human U3 genes (the RNU3 locus) preferentially associate with CBs in interphase cells. In addition, we have analyzed the genomic organization of the RNU3 locus by constructing a BAC and P1 clone contig. We found that, unlike the RNU1 and RNU2 loci, U3 genes are not tandemly repeated. Rather, U3 genes are clustered on human chromosome 17p11.2, with evidence for large inverted duplications within the cluster. Thus all of the CBORs identified to date are composed of either tandemly repeated or tightly clustered genes. The evolutionary and cell biological consequences of this type of organization are discussed.

Base Sequence↗
PubMed1997

Immunosuppression by D-isomers of HLA class I heavy chain (amino acid 75 to 84)-derived peptides is independent of binding to HSC70.

BACKGROUND: Peptides derived from the class I heavy chain were shown to modulate immune responses in vitro and in vivo. A peptide derived from HLA-B2702 (2702.75-84) inhibited differentiation of cytotoxic T cells as well as T cell and natural killer cell-mediated cytotoxicity in vitro. Peptide-mediated immunomodulation seemed to be independent of the MHC proteins expressed by responder and stimulator cells. In vivo studies in rodents demonstrated prolongation of heart and skin allograft survival after peptide therapy. Here, the correlation between the peptide's biological activity and its amino acid sequence was analyzed using peptides derived from amino acid 75-84 of several mouse, rat, and human MHC class I proteins as well as peptides with single amino acid substitutions in the 2702.75-84 sequence. METHODS: Peptides consisting of both L- and D-amino acids were tested for inhibition of murine and human T cell-mediated and lymphokine-activated killer cell-mediated cytotoxicity, binding to hsc70, and prolongation of heart allograft survival in vivo. RESULTS: Replacement of glutamic acid residue (E) at position 75 with valine (V) resulted in a peptide [2702.75-84(E>V)] with increased in vitro and in vivo activity but unchanged affinity for hsc70. Surprisingly, both L- and D-isomers of 2702.75-84 and 2702.75-84(E>V) inhibited cytotoxic cells in vitro and prolonged heart allograft survival in vivo. However, as expected, the peptides consisting of D-amino acids did not bind to hsc70. CONCLUSION: Assuming that both D- and L-isomers modulate immune responses by similar mechanisms, these results suggest that the peptides' effect is independent of binding to hsc70.

Adenosine Triphosphatases↗
PubMed1997

Evidence for a role of C-terminal amino acid residues in skeletal muscle Ca2+ release channel (ryanodine receptor) function.

The effects of deleting 1, 3 and 15 amino acid residues from the highly conserved C-terminus of the tetrameric skeletal muscle ryanodine receptor (RyR) complex were determined. Immunoblot analysis indicated similar expression levels in HEK293 cells for full-length and mutant proteins. Full-length and RyR lacking the last amino acid showed [3H]ryanodine binding and single channel activities typical of native receptors. Deletion of 3 amino acids resulted in decreased activities, whereas deletion of 15 amino acids yielded an inactive RyR. These results suggest that the most 15 C-terminal amino acids are important for the expression of a functional RyR complex.

Animals↗
PubMed1997

Cloning of the cDNA coding for rat brain CMP-NeuAc:GD3 alpha2-8 sialyltransferase.

The cDNA coding for GT3-synthase has been cloned from a rat fetal brain cDNA library. The complete nucleotide sequence and the deduced amino acid sequence of the rat GT3-synthase cDNA were highly homologous to those of the mouse GT3-synthase. Quantitative RT-PCR showed that, as a key enzyme for the synthesis of 'c'-series gangliosides, the expression of GT3-synthase was developmentally regulated in embryonic rat brains.

Amino Acid Sequence↗
PubMed1997

Ghost artifact reduction for echo planar imaging using image phase correction.

An algorithm is described for reducing ghost artifacts in echo planar imaging (EPI) using phase corrections derived from images reconstructed using only even or odd k-space lines. The N/2 ghost, that arises principally from time-reversal of alternate k-space lines, was significantly reduced by this algorithm without the need for a calibration scan. In images obtained in eight subjects undergoing EPI for auditory functional MRI (fMRI) experiments, N/2 ghost intensity was reduced from 10.3% +/- 2.1% (range: 7.9-14.1%) to 4.5% +/- 0.2% (range: 4.1-4.9%) of parent image intensity, corresponding to a percent reduction in ghost intensity of 54% +/- 9% (range: 43-65%), and the algorithm restored this intensity to the parent image. It provided a significant improvement in image appearance, and increased the correlation coefficients related to neural activation in functional MRI studies. The algorithm provided reduction of artifacts from all polynomial orders of spatial phase errors in both spatial directions. The algorithm did not eliminate N/2 ghost intensity contributed by field inhomogeneities, susceptibility, or chemical shift.

Algorithms↗