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K Su

Publications and source records attributed to K Su.

At least 37 records · Page 2Linked to original sources
PubMed1998

Cloning and characterization of a novel human chemokine receptor.

The present study reports the identification of a human gene, HCR, which encodes a novel human chemokine receptor. The partial sequence of the HCR gene was first found in a human neutrophil cDNA library. With the use of an expressed sequence tag (EST) probe from the neutrophil library, the full length HCR cDNA was isolated. The open reading frame of HCR cDNA predicts a protein of 345 amino acids with seven transmembrane domain topography. The HCR gene exhibits good homology to human MIP-1a receptor with 43.1% amino acid identity and 64.4% amino acid similarity and also shows considerable sequence homology to other human chemokine receptors such as the MCP-3 receptor, MCP-5 receptor, and MCP-1 receptor. Northern blot analysis suggests that HCR gene is expressed abundantly in immunal tissues such as spleen, fetal liver, lymph node, and bone marrow. Strong expression was also found in human lung and heart. A chromosome mapping study indicated that HCR gene is positioned within human chromosome band Xq13. Our result suggests that HCR gene is a novel putative chemokine receptor.

Amino Acid Sequence↗
PubMed1998

Cloning of a putative human neurotransmitter receptor expressed in skeletal muscle and brain.

With the use of the degenerated nucleotides that contain the conserved sequence of G protein-coupled receptor, we have identified a 648-bp clone (HDGRC02) from human genomic DNA with significant sequence homology to human neurotransmitter receptors. HDGRC02 was then used as a probe for the screening of full length gene. From human Lambda DASH II genomic library, a 1.6 Kb clone encoded a full length gene was isolated and named putative neurotransmitter receptor (PNR). PNR has a single open reading frame which predicts a 38.3 KD protein of 338 amino acids with seven transmembrane domain topography. The amino acid sequence of PNR exhibits considerable homology to the rat 5-HR1D receptor with 35% amino acid identity and 56% amino acid similarity. PNR also shows significant sequence homology to the 5-HT1D receptor from Japanese puffer fish fugu, to the 5-HT4L receptor from mouse, to the alpha-2 adrenergic receptor and to the D2 dopamine receptor. Northern blot analysis indicates that PNR is expressed in skeletal muscle and selected areas of the brain. A chromosome mapping study located the PNR gene with human chromosome band of 6q23. The findings in the present study demonstrate that PNR is a putative neurotransmitter receptor.

Amino Acid Sequence↗
PubMed1998

Cloning, characterization, and mapping of human homolog of mouse T-cell death-associated gene.

To establish immunologic autotolerance, self-reactive immature thymocytes are eliminated by negative selection during T-cell development in the thymus. Self-reactive clones undergo apoptosis after stimulation via the T-cell receptor (TCR). The process of cell selection is determined by the dedication of the TCR for tolerogenic antigen/major histocompatibility complex. We have cloned a novel human gene that is highly homologous in the transmembrane and G protein-coupling domains to mouse T-cell death-associated gene 8 (TDAG8). The gene, human TDAG8 (hTDAG8), which belongs to the G protein-couple receptor superfamily, encodes a protein of 337 amino acids. An expressed sequence tag (EST) corresponding to hTDAG8 was identified from a human thyroid cDNA library and subsequently used to isolate a full-length genomic clone. Northern blot analysis revealed that the hTDAG8 gene is expressed predominantly in lymphoid tissues, including peripheral blood leukocytes, spleen, lymph nodes, and thymus. Stably transfected mammalian CHO cells were generated, and heterologous expression of hTDAG8 was confirmed by Northern blot analysis. Fluorescent in situ hybridization (FISH) revealed that hTDAG8 maps to human chromosome 14q31-32.1, a region in which abnormalities associated with human T-cell lymphoma or leukemia are found. Taken together, these data implicate the hTDAG8 gene in T-cell-associated diseases in humans, but its actual physiological and pathological role in the human immune system needs further investigation.

Amino Acid Sequence↗
PubMed1998

A binding site for multiple transcriptional activators in the fushi tarazu proximal enhancer is essential for gene expression in vivo.

The Drosophila homeobox gene fushi tarazu (ftz) is expressed in a highly dynamic striped pattern in early embryos. A key regulatory element that controls the ftz pattern is the ftz proximal enhancer, which mediates positive autoregulation via multiple binding sites for the Ftz protein. In addition, the enhancer is necessary for stripe establishment prior to the onset of autoregulation. We previously identified nine binding sites for multiple Drosophila nuclear proteins in a core 323-bp region of the enhancer. Three of these nine sites interact with the same cohort of nuclear proteins in vitro. We showed previously that the nuclear receptor Ftz-F1 interacts with this repeated module. Here we purified additional proteins interacting with this module from Drosophila nuclear extracts. Peptide sequences of the zinc finger protein Ttk and the transcription factor Adf-1 were obtained. While Ttk is thought to be a repressor of ftz stripes, we have shown that both Adf-1 and Ftz-F1 activate transcription in a binding site-dependent fashion. These two proteins are expressed ubiquitously at the time ftz is expressed in stripes, suggesting that either may activate striped expression alone or in combination with the Ftz protein. The roles of the nine nuclear factor binding sites were tested in vivo, by site-directed mutagenesis of individual and multiple sites. The three Ftz-F1-Adf-1-Ttk binding sites were found to be functionally redundant and essential for stripe expression in transgenic embryos. Thus, a biochemical analysis identified cis-acting regulatory modules that are required for gene expression in vivo. The finding of repeated binding sites for multiple nuclear proteins underscores the high degree of redundancy built into embryonic gene regulatory networks.

Amino Acid Sequence↗
PubMed1998

[Determination of antimicrobial preservative dimethyl fumarate by high performance liquid chromatography].

This paper described a HPLC method for determining dimethyl fumarate (DMF), a new antimicrobial preservative. Chromatographic conditions were as follows: Micropak MGH 5 mu cap column, 4.00 mm x 300.0 mm EtOH-H2O (90:10, V/V) eluent, and UV detector at 216 nm. In the range of 0.036-2.96 micrograms, the sample size and peak area showed a linear relationship with regression equation, Y = 3.215 x 10(-6)X - 0.04797 and r = 0.9996. Standard deviation was 3.2 x 10(-4).

Chromatography, High Pressure Liquid↗
PubMed1997

Isolation, characterization, and mapping of two human potassium channels.

Two novel human genes encoding putative potassium channels, kH1 and kH2, were identified from a human fetal brain cDNA library. Sequence analysis showed that kH1 and kH2 are homologous to rat IK8 and rat K13, respectively. The kH1 encodes a polypeptide of 495 amino acids, which shares 88% and 95% identity to IK8 at the nucleotide and amino acid level, respectively. The kH2 encodes a polypeptide of 515 amino acids with 86% and 92% identity to K13 at the nucleotide and amino acid level, respectively. Northern blot studies revealed that one mRNA species, approximately 5kb, of the kH1 was expressed abundantly in tissues examined, including the heart, skeletal muscle, and less abundant in the brain, liver, kidney, and pancreas. Interestingly, an alternative spliced form of 2.4 kb mRNA species of the kH1 was also found in the brain. Unlike kH1, 2.4 kb of kH2 was expressed predominantly in the brain, placenta, and the skeletal muscle where it shared a differently spliced form of the kH2 mRNA, approximately 2.0 kb. Fluorescence in situ hybridization localized kH1 to the human chromosome 2p25 and kH2 to the human chromosome 20q13.

Amino Acid Sequence↗
PubMed1997

A 2.5-Mb transcript map of a tumor-suppressing subchromosomal transferable fragment from 11p15.5, and isolation and sequence analysis of three novel genes.

11p15.5 is an important tumor-suppressor gene region, showing loss of heterozygosity in Wilms tumor, rhabdomyosarcoma, adrenocortical carcinoma, and lung, ovarian, and breast cancer. We previously mapped directly by genetic complementation a subtransferable fragment (STF) harboring an embryonal tumor-suppressor gene and spanning about 2.5 Mb. We have now mapped the centromeric end of this STF between D11S988 and D11S12 and its telomeric end between D11S1318 and TH. We have isolated a complete contig of PAC, P1, BAC, and cosmid genomic clones spanning the entire 2.5-Mb region defined by this STF, as well as more than 200 exons from these genomic clones using exon trapping. We have isolated genes in this region by directly screening DNA libraries as well as by database searching for ESTs. Nine of these genes have been reported previously by us and by others. However, the initial mapping of most of those genes was based on FISH or somatic cell hybrid analysis, and here we precisely define their physical location. These genes include RRM1, GOK (D11S4896E), Nup98, CARS, hNAP2 (NAP1L4), p57KIP2 (CDKN1C), KVLQT1 (KCNA9), TAPA-1, and ASCL2. In addition, we have identified several novel genes in this region, three of which, termed TSSC1, TSSC2, and TSSC3, are reported here. TSSC1 shows homology to Rb-associated protein p48 and chromatin assembly factor CAF1, and it is located between GOK and Nup98. TSSC2 is homologous to Caenorhabditis elegans beta-mannosyl transferase, and it lies between Nup98 and CARS. TSSC3 shows homology to mouse TDAG51, which is implicated in FasL-mediated apoptosis, and it is located between hNAP2 and p57KIP2. Thus, these genes may play a role in malignancies that involve this region.

Amino Acid Sequence↗
PubMed1997

Cloning, mapping, and tissue distribution of a human homologue of the mouse jerky gene product.

Inactivation of the jerky gene by insertion of a transgene into the mouse genome results in epileptic seizures in transgenic mice. This finding indicates that the jerky gene plays an important role in inducing epilepsy syndromes in mice. We report here our efforts in cloning, chromosomal mapping, and analysis of tissue distribution of a novel human gene, the HHMJG, a homologue to the mouse jerky gene product. We have successfully identified a full length cDNA clone encoding a novel human protein homologous to the mouse jerky gene product. The finding was based on the result of an analysis of EST (expressed sequence tag) sequences of a clone from a human tonsil cDNA library. A 4.0 kb mRNA species of the HHMJG is abundantly expressed in the majority of human tissues examined, including brain and skeletal muscle. However, in the testes, two mRNA species of the HHMJG, approximately 2.0 and 4.0 kb, are abundantly expressed. Sequence analysis of the HHMJG cDNA indicates that it encodes a putative protein of 51 kD, which shares significant sequence homology to not only the mouse jerky gene product but also some nuclear regulatory proteins, such as centromere binding protein-B. The predicted nuclear localization of the HHMJG product suggests that this protein may function as a nuclear regulatory protein. The result of human chromosomal mapping shows that the HHMJG is located on human chromosome 11q21. Our identification of the HHMJG cDNA provides a potential gene candidate to further investigate the biological significance and clinical implications of the HHMJG in human epilepsy.

Amino Acid Sequence↗
PubMed1997

A novel endothelin receptor type-B-like gene enriched in the brain.

We report here our effort of cloning and characterization of a novel human gene, which encodes a putative human endothelin receptor type B like protein (hET(B)R-LP), from a human hippocampus tissue cDNA library. hET(B)R-LP consists of 614 amino acids with seven putative transmembrane domains. The deduced amino acid sequence of hET(B)R-LP is 52% similar and 26.7% identical to human endothelin type B receptor. A 4.0 kb mRNA of hET(B)R-LP is abundantly expressed in the human brain. The results of in situ hybridization and reverse transcriptase in situ gene amplification reveal tissue distribution and cellular localization of signals of hET(B)R-LP mRNA in the neuronal cells, particularly concentrated in Purkinje cells of the cerebellum, and neuronal cells of the hippocampus of human brain, including pyramidal cells of Ammon's horn and granule cells of the dentate gyrus. A 4.0 kb mRNA of hET(B)R-LP is also less abundantly expressed in the liver and the placenta. Expression of recombinant protein, hET(B)R-LP/HA, in cells of COS7 and HEK293 transfected with plasmid DNA, hET(B)R-LP/HA/pcDNA1/Amp, was confirmed by Northern blot analysis and by immunofluorescence staining of cells with anti-HA antibody. Specific binding of radiolabeled ET-1 and ET-3 to membrane preparations and to intact cells expressing recombinant protein of hET(B)R-LP/HA did not show any significant difference of binding properties between cells transfected with plasmid DNA, hET(B)R-LP/HA/pcDNA1/Amp, and cells untransfected, including both COS7 cells and HEK293 cells. The results of assays of measuring Ca++ mobilization and cAMP production in HEK293 cells indicate that ET-1, ET-3, bombesin and neuropeptide Y are unable to produce any kind of significant difference of Ca++ mobilization and cAMP production between HEK293 cells expressing recombinant protein and HEK293 cells untransfected or HEK293 cells transfected with vector DNA only (pcDNA1/Amp) in functional assays performed. Therefore, its ligand and physiological significance of hET(B)R-LP remains to be discovered.

Amino Acid Sequence↗
PubMed1997

Cloning and partial characterization of the mouse glutamine:fructose-6-phosphate amidotransferase (GFAT) gene promoter.

Glutamine:fructose-6-phosphate amidotransferase (GFAT) is the enzyme that is rate limiting in the synthesis of glucosamine and hexosamines. Glucosamine has been proposed to contribute to the glucotoxicity of diabetes. Evidence that the gene encoding GFAT is transcriptionally regulated prompted us to clone and characterize its promoter. The position of the mouse GFAT promoter relative to the translational start site was located by primer extension and found to be 149 bp upstream of the translational start site. A 1.9 kb SacI fragment of the GFAT gene was found to contain the promoter and 88 bp of sequence downstream of the transcriptional start site. This promoter segment could drive expression of a luciferase reporter gene, could confer correct transcriptional initiation to the reporter and could confer the EGF-responsiveness previously observed in the native gene. The mouse GFAT promoter lacks a canonical TATA box and has several GC boxes within a highly GC-rich region. Deletional analysis of the promoter indicated that a proximal element extending to -120 relative to the transcriptional start site could confer reporter expression at a level of 57% of the 1.9 kb construct. Detailed analysis of this proximal region by DNase I footprinting, electrophoretic mobility shift assays and site-directed mutagenesis indicated that Sp1 binds to three elements in this proximal promoter segment and plays a vital role in regulation of transcription from this gene.

3T3 Cells↗
PubMed1997

Cloning and characterization of a novel human DNase.

The therapeutic significance of recombinant human DNase I in treating the patients with cystic fibrosis has risen our interests in identifying other human DNase I-like enzymes to study their biological significance. Here we described our work of cloning and characterization of a novel gene, which encodes a human protein homologous to human DNase I. A full length cDNA clone of this gene consists of 1290 bp, encoding a polypeptide of 306 amino acids. The deduced amino acid sequence of this novel human DNase (nhDNase) is 45% identical to that of human DNase I. Among sixteen human tissues examined by Northern Blot, high level expression of nhDNase was found in human liver and spleen. Recombinant protein of nhDNase was produced in a Baculovirus expression system and purified by chromatography and reverse-phase HPLC. Purified recombinant nhDNase migrated as a single band of about 33 kD molecular weight analyzed by SDS-PAGE. The DNase activity of nhDNase was demonstrated by assay of hydrolysis of S.S.DNA. Its activity was dependent upon the presence of divalent metal irons, calcium and magnesium. However, unlike bovine pancreas DNase I, nhDNase was not inhibited by G-actin of bovine muscle, which indicates the physiological significance of this enzyme in clinical implication.

Actins↗
PubMed1997

The nuclear hormone receptor Ftz-F1 is a cofactor for the Drosophila homeodomain protein Ftz.

Homeobox genes specify cell fate and positional identity in embryos throughout the animal kingdom. Paradoxically, although each has a specific function in vivo, the in vitro DNA-binding specificities of homeodomain proteins are overlapping and relatively weak. A current model is that homeodomain proteins interact with cofactors that increase specificity in vivo. Here we use a native binding site for the homeodomain protein Fushi tarazu (Ftz) to isolate Ftz-F1, a protein of the nuclear hormone-receptor superfamily and a new Ftz cofactor. Ftz and Ftz-F1 are present in a complex in Drosophila embryos. Ftz-F1 facilitates the binding of Ftz to DNA, allowing interactions with weak-affinity sites at concentrations of Ftz that alone bind only high-affinity sites. Embryos lacking Ftz-F1 display ftz-like pair-rule cuticular defects. This phenotype is a result of abnormal ftz function because it is expressed but fails to activate downstream target genes. Cooperative interaction between homeodomain proteins and cofactors of different classes may serve as a general mechanism to increase HOX protein specificity and to broaden the range of target sites they regulate.

Animals↗
PubMed1997

O glycosylation of an Sp1-derived peptide blocks known Sp1 protein interactions.

The O-linked N-acetylglucosamine (O-GlcNAc) modification of proteins is dynamic and abundant in the nucleus and cytosol. Several transcription factors, including Sp1, have been shown to contain this modification; however, the functional role of O-GlcNAc in these proteins has not been determined. In this paper we describe the use of the previously characterized glutamine-rich transactivation domain of Sp1 (B-c) as a model to investigate the role of O-GlcNAc in Sp1's transcriptionally relevant protein-to-protein interactions with the TATA-binding-protein-associated factor (TAF110) and holo-Sp1. When the model Sp1 peptide was overexpressed in primate cells, this 97-amino-acid domain of Sp1 was found to contain a dominant O-GlcNAc residue at high stoichiometry, which allowed the mapping and mutagenesis of this glycosylation site. In vitro interaction studies between this segment of Sp1 and Drosophila TAF110 or holo-Sp1 indicate that the O-GlcNAc modification functions to inhibit the largely hydrophobic interactions between these proteins. In HeLa cells, the mutation at the mapped glycosylation site was permissive for transcriptional activation. We propose the hypothesis that the removal of O-GlcNAc from an interaction domain can be a signal for protein association. O-GlcNAc may thereby prevent untimely and ectopic interactions.

Amino Acid Sequence↗
PubMed1996

Molecular cloning and characterization of the human anaphylatoxin C3a receptor.

In a human neutrophil cDNA library, an orphan G-protein-coupled receptor, HNFAG09, with 37% nucleotide identity to the C5a receptor (C5a-R, CD88) was identified. A novel feature of this gene, unlike C5a-R and other G-protein-coupled receptors, is the presence of an extraordinarily large predicted extracellular loop comprised of in excess of 160 amino acid residues between transmembrane domains 4 and 5. Northern blot analysis revealed that expression of mRNA for this receptor in human tissues, while similar, was distinct from C5a-R expression. Although there were differences in expression, transcripts for both receptors were detected in tissues throughout the body and the central nervous system. Mammalian cells stably expressing HNFAG09 specifically bound 125I-C3a and responded to a C3a carboxyl-terminal analogue synthetic peptide and to human C3a but not to rC5a with a robust calcium mobilization response. HNFAG09 encodes the human anaphylatoxin C3a receptor.

Amino Acid Sequence↗
PubMed1995

Human interleukin-3 receptor modulates bcl-2 mRNA and protein levels through protein kinase C in TF-1 cells.

Upon withdrawal of interleukin-3 (IL-3) from human factor-dependent erythroleukemic cell line TF-1, bcl-2 mRNA and protein levels decrease within 8 to 24 hours. Accompanying this decrease is the onset of apoptosis as determined by flow cytometric analysis of DNA degradation. By 8 to 18 hours of deprivation approximately 70% to 80% of the cells have entered apoptosis. Downregulation of protein kinase (PK) by a 24-hour incubation in 100 nmol/L 12-O-tetradecanoyl-phorbol-13-acetate (TPA) in the presence of IL-3 dramatically reduced bcl-2 mRNA levels, and induced apoptosis in the presence of IL-3. We have also found that even in the presence of IL-3, two inhibitors of PKC, light-activated calphostin and H-7, substantially reduced the levels of bcl-2 mRNA between 8 and 24 hours as measured by a semi-quantitative reverse transcriptase/polymerase chain reaction assay method; however, the cyclic nucleotide-dependent PK inhibitor HA 1004, that is a structural analog of H-7 but a poor inhibitor of PKC, did not reduce bcl-2 levels in the presence of IL-3. This decrease in bcl-2 mRNA was accompanied by a decline in bcl-2 protein levels by 8 to 24 hours after addition of light-activated calphostin. In addition to interfering with the maintenance of bcl-2 mRNA levels, inhibition of PKC with H-7 inhibited the induction of bcl-2 mRNA in factor-deprived TF-1 cells restimulated with IL-3. The cyclic nucleotide-dependent PK inhibitor HA 1004 did not inhibit IL-3-induced bcl-2 mRNA. Studies with actinomycin D showed that transcription plays a major role in maintaining bcl-2 levels in TF-1 cells, and it is therefore likely that IL-3 plays a role in maintaining bcl-2 transcription through activation of PKC in these cells.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗
PubMed1993

Pre-Golgi degradation of yeast prepro-alpha-factor expressed in a mammalian cell. Influence of cell type-specific oligosaccharide processing on intracellular fate.

We demonstrate that expression of yeast prepro-alpha-factor in GH3 rat pituitary cells results in its degradation in an endoplasmic reticulum (ER) or early Golgi compartment, suggesting that this wild-type prohormone is recognized as abnormal or misfolded in the context of a mammalian ER. In GH3 cells, as in yeast, the nascent polypeptide is efficiently targeted to the ER, where it undergoes cleavage of its amino-terminal signal peptide and core glycosylation to form glycosylated pro-alpha-factor (gp alpha f). Subsequently, however, this species disappears from cells with a half-time of 25-30 min (including a 10-20-min lag), and no alpha-factor- or proregion-derived products can be detected. Localization of the degradative process to the ER is suggested by its occurrence in the presence of brefeldin A or chloroquine and by the endoglycosidase H susceptibility of the substrate. We present evidence that Asn-linked oligosaccharide processing, which differs in extent between yeast and mammalian cells, may be an important factor in determining degradation in this heterologous circumstance. When GH3 cells are treated with deoxymannojirimycin, an inhibitor of ER alpha-mannosidases, gp alpha f is essentially stable, suggesting that trimming of core oligosaccharides below Man8 (a process that does not occur in yeast) strongly promotes proteolysis. Inhibition of ER glucosidase activity by treatment with deoxynojirimycin, by contrast, considerably accelerates the disappearance of gp alpha f (t1/2 = 8-10 min). These data indicate that cell type-specific post-translational modifications of a secretory glycoprotein can substantially modify its recognition by the mammalian ER degradative apparatus.

1-Deoxynojirimycin↗
PubMed1989

A preliminary dose-ranging trial of proglumide for the treatment of refractory schizophrenics.

A dose-finding study of proglumide added to neuroleptics for the treatment of schizophrenic patients who were relatively refractory to their ongoing neuroleptic regimen was performed. Initially, four patients were open-label treated using a regimen of progressively increasing doses (proglumide 0.5-1024 mg/day) for 4 weeks. Afterwards, seven patients were given low doses (0.5 mg/day) followed by higher doses (500 mg/day) for a total period of 8 weeks. Overall, no improvement was seen in these refractory patients as a group at any dose. In individual patients, modest improvement or worsening of psychotic symptoms was observed. The results suggest that more potent cholecystokinin (CCK) antagonists and a greater knowledge of CCK pharmacology are needed before novel treatments exploiting the interaction of CCK and dopamine in the brain can be developed.

Antipsychotic Agents↗