Biomedical subjects

J Yoon

Publications and source records attributed to J Yoon.

At least 19 recordsLinked to original sources

PubMed1999

The Ets factors PU.1 and Spi-B regulate the transcription in vivo of P2Y10, a lymphoid restricted heptahelical receptor.

To investigate the in vivo functions of PU.1 and Spi-B, two highly related Ets transcription factors, we previously generated PU. 1(+/+)Spi-B(-/-) and PU.1(+/-)Spi-B(-/-) mice and demonstrated a significant decrease in B-cell receptor (BCR) signaling in mutants. Major components of BCR signaling appear to be expressed at normal levels in these mice, implying that PU.1 and Spi-B cooperate in the transcription of additional target genes important for antigen receptor signaling. We used subtractive hybridization to identify novel in vivo PU.1/Spi-B target genes and determined that the expression of a heptahelical receptor, P2Y10, is dramatically reduced in PU.1(+/-)Spi-B(-/-) B-cells. Further analysis shows that P2Y10 expression is restricted to lymphoid cells and parallels that of Spi-B in B-lymphocytes. Lastly, the P2Y10 promoter contains a PU. 1/Spi-B binding site functionally required for efficient transcription in B-cells. Thus, P2Y10 is likely to be a direct in vivo transcriptional target for PU.1 and Spi-B and provides a unique model to explore transcriptional regulation by this Ets factor subfamily. Furthermore, P2Y10 suggests an intriguing connection between heterotrimeric G-proteins and BCR signaling.

3T3 Cells↗

PubMed1999

Structural analysis of phylogenetically conserved J domain protein gene.

Novel cDNAs encoding evolutionarily conserved J Domain Proteins (JDPs) were investigated from Drosophila and mouse. Each of the full coding sequences potentially encodes a conserved J domain, but lacks additional characteristic structures present in DnaJ family proteins. The expression was restricted to head in Drosophila. However, ubiquitous expression was observed in mice with the highest level in kidney.

Amino Acid Sequence↗

PubMed1999

Characterization of the Shank family of synaptic proteins. Multiple genes, alternative splicing, and differential expression in brain and development.

Shank1, Shank2, and Shank3 constitute a family of proteins that may function as molecular scaffolds in the postsynaptic density (PSD). Shank directly interacts with GKAP and Homer, thus potentially bridging the N-methyl-D-aspartate receptor-PSD-95-GKAP complex and the mGluR-Homer complex in synapses (Naisbitt, S., Kim, E., Tu, J. C. , Xiao, B., Sala, S., Valtschanoff, J., Weinberg, R. J., Worley, P. F., and Sheng, M. (1999) Neuron 23, 569-582; Tu, J. C., Xiao, B., Naisbitt, S., Yuan, J. P., Petralia, R. S., Brakeman, P., Doan, A., Aakalu, V. K., Lanahan, A. A., Sheng, M., and Worley, P. F. (1999) Neuron 23, 583-592). Shank contains multiple domains for protein-protein interaction including ankyrin repeats, an SH3 domain, a PSD-95/Dlg/ZO-1 domain, a sterile alpha motif domain, and a proline-rich region. By characterizing Shank cDNA clones and RT-PCR products, we found that there are four sites for alternative splicing in Shank1 and another four sites in Shank2, some of which result in deletion of specific domains of the Shank protein. In addition, the expression of the splice variants is differentially regulated in different regions of rat brain during development. Immunoblot analysis of Shank proteins in rat brain using five different Shank antibodies reveals marked heterogeneity in size (120-240 kDa) and differential spatiotemporal expression. Shank1 immunoreactivity is concentrated at excitatory synaptic sites in adult brain, and the punctate staining of Shank1 is seen in developing rat brains as early as postnatal day 7. These results suggest that alternative splicing in the Shank family may be a mechanism that regulates the molecular structure of Shank and the spectrum of Shank-interacting proteins in the PSDs of adult and developing brain.

Adaptor Proteins, Signal Transducing↗

PubMed1999

SPI-B activates transcription via a unique proline, serine, and threonine domain and exhibits DNA binding affinity differences from PU.1.

SPI-B is a B lymphocyte-specific Ets transcription factor that shares a high degree of similarity with PU.1/SPI-1. In direct contrast to PU.1(-/-) mice that die in utero and lack monocytes, neutrophils, B cells, and T cells, Spi-B-/- mice are viable and exhibit a severe B cell proliferation defect. Since PU.1 is expressed at wild type levels in Spi-B-/- B cells, the mutant mice provide genetic evidence that SPI-B and PU.1 have at least some non-redundant roles in B lymphocytes. To begin to understand the molecular basis for these defects, we delineated functional domains of SPI-B for comparison to those of PU.1. By using a heterologous co-transfection system, we identified two independent transactivation domains in the N terminus of SPI-B. Interestingly, only one of these domains (amino acids 31-61), a proline/serine/threonine-rich region, unique among Ets proteins, is necessary for transactivation of the immunoglobulin lambda light chain enhancer. This transactivation motif is in marked contrast to PU.1, which contains acidic and glutamine-rich domains. In addition, we describe a functional PU.1 site within the c-FES promoter which SPI-B fails to bind efficiently and transactivate. Finally, we show that SPI-B interacts with the PU.1 cofactors Pip, TBP, c-Jun and with lower affinity to nuclear factor interleukin-6beta and retinoblastoma. Taken together, these data suggest that SPI-B binds DNA with a different affinity for certain sites than PU.1 and harbors different transactivation domains. We conclude that SPI-B may activate unique target genes in B lymphocytes and interact with unique, although currently unidentified, cofactors.

Amino Acid Sequence↗

PubMed1999

The characterization of macrophages and osteoclasts in tissues harvested from revised total hip prostheses.

The differentiation and maturation of macrophages and osteoclasts at the prosthetic interface in cases of implant loosening are poorly understood. Using histochemical and immunohistochemical staining methods, we compare macrophage differentiation in tissues from revised hip replacements in patients with specific clinical-radiological appearances. Periprosthetic tissues were harvested from 12 cemented acetabular and 12 cemented femoral components in 24 patients undergoing revision hip replacement. The prostheses were all radiographically and clinically loose. Six acetabular and six femoral components demonstrated radiographic ballooning osteolysis. Serial 6 microm frozen sections of the periprosthetic tissues were processed with hematoxylin and eosin for general tissue morphology, and analyzed for the presence of tartrate resistant acid phosphatase (TRAP, an osteoclast marker). Immunoperoxidase staining using monoclonal antibodies to CD68 (macrophages and osteoclasts) and CD51 (the alpha chain of the vitronectin receptor, an osteoclast marker) was also performed. Approximately 8-30% of the total cells in the tissues were positive for TRAP and the vitronectin receptor, and comprised a subset of the CD68 positive macrophages and macrophage polykaryons. However, there were no statistically significant differences between specific groups (femoral vs. acetabular, osteolysis vs. no osteolysis) for the numbers or percentages of macrophages or osteoclast-like cells. Once prosthetic loosening has occurred, few differences in the macrophage-osteoclast profile of tissues from different periprosthetic locations, with and without osteolysis, are noted. This suggests a final common biologic pathway for periprosthetic bone resorption, once implant loosening has transpired.

Acid Phosphatase↗

PubMed1999

Identification of the enteropathogens Campylobacter jejuni and Campylobacter coli based on the cadF virulence gene and its product.

Campylobacter jejuni and Campylobacter coli are common causes of gastroenteritis in humans. Infection with C. jejuni or C. coli is commonly acquired by eating undercooked chicken. The goal of this study was to develop specific detection assays for C. jejuni and C. coli isolates based on the cadF virulence gene and its product. The cadF gene from C. jejuni and C. coli encodes a 37-kDa outer membrane protein that promotes the binding of these pathogens to intestinal epithelial cells. A fragment of approximately 400 bp was amplified from 38 of 40 (95%) C. jejuni isolates and 5 of 6 (83.3%) C. coli isolates with primers designed to amplify an internal fragment of the cadF gene. PCR was then used to amplify Campylobacter DNA from store-bought chickens. A 400-bp band was amplified from 26 of the 27 chicken carcasses tested by the PCR-based assay. The CadF protein was detected in every C. jejuni and C. coli isolate tested, as judged by immunoblot analysis with a rabbit anti-C. jejuni 37-kDa serum. In addition, methanol-fixed samples of whole-cell C. jejuni and C. coli were detected with the rabbit anti-37-kDa serum by using an indirect-immunofluorescence microscopy assay. These findings indicate that the cadF gene and its product are conserved among C. jejuni and C. coli isolates and that a PCR assay based on the cadF gene may be useful for the detection of Campylobacter organisms in food products.

Animals↗

PubMed1999

A precipitation reaction found in patients with hepatitis C as a marker for the purification of virus-like particles.

Even after the molecular cloning of the hepatitis C virus (HCV), an HCV-specific precipitation reaction has not yet been identified. We attempted to develop a precipitation system exclusively for anti-HCV-positive sera as a first step in finding an HCV-specific antigen and HCV-associated particles. In some patients being in a final stage of different liver diseases, we found sera (179/132,761; designated 'a-CK') which specifically precipitated with anti-HCV- and HCV-RNA-positive sera (designated 'CK'). When CK-positive sera were searched for in patients with various liver diseases using standard a-CK-positive plasma, CK was detected in 420 (57.9%) of 726 anti-HCV-positive sera and in none of the 1,630 anti-HCV-negative ones. The nature of CK and a-CK has not been fully clarified yet; CK demonstrated inter-betagamma mobility, whereas a-CK showed beta-globulin mobility; CK was not detected in cryoprecipitate, but HCV RNA was present in precipitates of CK-positive plasma incubated with one that was a-CK positive. Transmission electron microscopy revealed two size ranges of particles in the precipitate of CK- and a-CK-positive plasmas, 23-38 nm and 48-65 nm. We have found a novel precipitation system which is exclusive to anti-HCV-positive sera and which specifically precipitates an HCV-RNA-containing serum fraction and particles. This system can be useful for the purification and characterization of the circulating particles. Furthermore, it may be a new approach to the nature of HCV-RNA-carrying material.

Centrifugation↗

PubMed1998

An analysis of two tandem promoters of the Drosophila purple gene.

We have analyzed two tandem promoters, separated by only about 400 bp, of the purple (pr) gene of Drosophila melanogaster, by fusing them to the firefly luciferase reporter gene and employing a transient expression assay with Drosophila S2 cells. Both the distal promoter and the proximal promoter were found to function in S2 cells and an about 700 bp long region (-270 to +421), containing both promoters, was sufficient to effect maximal promoter activity. When the two promoters were analyzed separately, the distal promoter was found to be much stronger in its function than the proximal promoter. At least three different kinds of cis elements near the transcription start site appear to play crucial roles in driving constitutive expression from the distal promoter. On the other hand, only a single cis element, which may play a role in tissue-specific expression, appears to be important for the activity of the proximal promoter in S2 cells. We propose that the clustering of important cis elements near the transcription start sites may be responsible for the selective regulation of the two tandem promoters.

Animals↗

PubMed1998

Molecular characterization of a novel Drosophila gene which is expressed in the central nervous system.

We have isolated and characterized a novel Drosophila melanogaster gene (noe) that is specifically and abundantly expressed in the central nervous system (CNS). The gene, which maps to 74B on the left arm of third chromosome, encodes a protein of 74 amino acids with no significant similarity to known protein sequences. The deduced amino acid sequence of the gene product is rich in basic amino acids, especially the lysine, and contains five potential phosphorylation sites. The noe gene lacks introns and seems to produce two transcripts by alternative polyadenylations. The promoter region deduced from 5'-RACE analysis contains a sequence similar to the TATA-box consensus sequence. RNA blot analysis detected 1.0 kb noe transcripts that are expressed from the third-instar larval stage to the adult stage and which are predominantly found in the adult heads. In situ hybridizations to tissue sections showed that the gene is abundantly expressed in neuronal cell bodies as well as in the neurophiles of adult and larval CNS (brain, optic lobe, and thoracic ganglia of adults and larval brain).

Amino Acid Sequence↗

PubMed1998

Promoter analysis of the Drosophila genes encoding TFIIB and TATA box-binding protein.

We have analyzed the 5'-flanking regions of the Drosophila genes encoding the TFIIB and TATA box-binding proteins (TBP) and mapped the regulatory regions required for their efficient expression. We found that the 500 bp long region (-439 to +60) and the 401 bp long region (-261 to +138) are required for the efficient expression of TFIIB and TBP genes, respectively. In the TFIIB promoter region, the upstream DNA between -439 and -280 and the downstream DNA between +8 and +60 are necessary for the stimulation of promoter activity. The upstream DNA between -439 and -280 stimulates transcription in an orientation dependent manner. In the TBP promoter region, the upstream DNA between -261 and -207, and the downstream DNA between +15 and +138 are necessary for the stimulation of promoter activity. The upstream DNA (-261 to -207) required for TBP promoter activity contains a 11 bp long palindromic sequence and a DNA replication-related element sequence. Particularly, we could find that the downstream promoter regions of TFIIB and TBP genes contain the conserved nucleotide sequences, suggesting the presence of a common regulatory mechanism for the expression of these two genes.

Animals↗

PubMed1998

Isolation and characterization of the Drosophila melanogaster cDNA encoding the sepiapterin reductase.

We have isolated and characterized the cDNA encoding Drosophila melanogaster sepiapterin reductase (SR). The amino acid sequence deduced from the cDNA sequence was 29% identical to those of mammalian SRs. The active site residues proposed from the three-dimensional structure of mouse SR are well conserved in Drosophila SR. The protein-coding region of the cDNA was expressed in Escherichia coli as a histidine fusion protein, and the resulting recombinant protein proved to have SR activity. The SR activity of the recombinant protein was inhibited by two indoleamines, N-acetyl serotonin and melatonin. Southern analysis suggests that the Drosophila SR gene is encoded by a single copy gene. RNA blot analysis revealed that the gene expresses 1.5 kb mRNA in both adult heads and bodies.

Alcohol Oxidoreductases↗

PubMed1998

Molecular characterization of the Drosophila melanogaster gene encoding the pterin 4alpha-carbinolamine dehydratase.

We have isolated and characterized the cDNA and the genomic DNA encoding Drosophila melanogaster pterin 4alpha-carbinolamine dehydratase (PCD). The amino acid sequence deduced from the cDNA sequence was very similar to those of PCDs previously reported in other species (19-57% identity). The protein coding region of the cDNA was expressed in E. coli as a histidine fusion protein, and the expressed protein proved to have PCD activity. The characterization of the Drosophila genomic clone revealed that the Drosophila PCD gene is interrupted by two introns. The potential promoter region, deduced from the determination of the transcription start point (tsp), lacks the distinct TATAAA box consensus sequence.

Amino Acid Sequence↗

PubMed1998

Characterization of bovine and human cDNAs encoding NAP-22 (22 kDa neuronal tissue-enriched acidic protein) homologs.

We have characterized the bovine and the human cDNAs encoding the NAP-22 (22 kDa neuronal tissue-enriched acidic protein) homologs. Both bovine and human cDNAs encode proteins of 227 amino acids. The deduced amino acid sequences of the bovine and the human proteins are 63% and 65% identical, respectively, to that of rat NAP-22 protein, strongly suggesting that both the cDNAs characterized encode NAP-22 proteins. They also share 45% and 41% amino acid sequence identities with chicken CAP-23 (23 kDa cytoskeleton associated protein). Several important protein motifs, including myristoylation and phosphorylation sites, are well conserved in sequences and positions in all three mammalian NAP-22 proteins and chicken CAP-23 proteins. The bovine cDNA was characterized further. Southern analysis of the bovine genomic DNA suggests that the bovine NAP-22 protein is encoded by a single-copy gene. RNA blot analysis revealed that the bovine gene for NAP-22 protein encodes a 1.7 kb transcript that is present only in the brain. Our data suggest that the four proteins, bovine and human NAP-22 homologs, rat NAP-22, and chicken CAP-23, have homologous functions in different organisms.

3' Untranslated Regions↗

PubMed1998

YAC/STS map of 15Mb of Xp21.3-p11.3, at 100kb resolution, with refined comparisons of genetic distances and DMD structure.

The 15 Mb region between DXS997 and DXS8054 in Xp21.3-p11.3 has been mapped at seven-fold average coverage in yeast artificial chromosomes (YACs) and 100 kb inter-sequence tagged site (STS) distance. YACs from six different collections show self-consistent maps. The STSs include 18 (CA) repeat and one tetranucleotide repeat marker that detect polymorphism, as well as eight well-studied genes, a second site for MXS1 sequences, and three expressed sequence tags (ESTs). One of the ESTs maps to intron 7 of Duchenne muscular dystrophy, and seems to be a processed intronic sequence with a poly(A) tail.

Base Sequence↗

PubMed1998

Investigations of azapeptides as mimetics of Leu-enkephalin.

Solution syntheses of azapeptide pentamers 2, 3, and 4 were accomplished. The binding affinity of these azapeptides and azatide 1 were examined in the context of monoclonal antibody 3-E7 known to strongly bind the [Leu5]enkephalin sequence.

Enkephalin, Leucine↗

PubMed1998

Environmental Management of Human Waste Disposal for Recreational Boating Activities

/ A methodology to estimate the number of pump-out facilities and dump stations required to service human waste disposal for recreational power boating activities in Pennsylvania during the 1994 boating season is described. Study results suggest that a total of 39 additional pump-out stations and 13 dump stations may be required on seven major waterbodies: The Three Rivers Area, Lake Erie/Presque Isle Bay, Raystown Lake, the Susquehanna River, the Delaware River, Lake Wallenpaupack, and the Kinzua Reservoir. Suggestions for improving the methodology are provided. KEY WORDS: Human waste; Recreation; Power boating; Waste facilities; Waste disposal; Pennsylvania

Journal Article↗

PubMed1998

Pulsed wave and color Doppler echocardiography and cardiac catheterization findings in bilateral pulmonary vein stenosis.

Pulmonary vein stenosis is a rare condition that usually is congenital in origin; it is almost invariably fatal in its bilateral and severe forms. It is often overlooked, however, during clinical examination, routine echocardiography, and even at cardiac catheterization. This report describes the pulsed Doppler, color flow echocardiography, and hemodynamic findings of bilateral pulmonary vein stenosis that accompany ventricular septal defect and atrial septal defect corrected by surgery.

Adult↗

PubMed1997

Promoter region of the Drosophila melanogaster norpA gene.

We have sequenced the 5'-flanking region of the Drosophila melanogaster norpA gene and characterized its promoter. The potential promoter region, which was deduced from the determination of the transcription start point (tsp), lacks a distinct TATA box sequence. Deletion analysis of the promoter region suggests that the minimal promoter necessary for efficient expression of the gene is located between -138 (PstI) and +278 relative to the tsp. Within this minimal promoter region, at least two downstream regulatory elements responsible for the stimulation of gene expression seem to exist in the DNA fragments between +44 and +121 and between +214 and +278. Among these, the DNA fragment between +44 and +121 affects promoter activity more dramatically (about 6-7 fold). This DNA fragment contains the consensus promoter element previously reported to be important for photoreceptor cell-specific expression, and this promoter element seems to be working in the norpA gene expression.

Animals↗