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PubMed1988

Nucleotide sequence of cloned cDNAs encoding chicken preproparathyroid hormone.

In order to characterize an avian parathyroid hormone gene, a lambda gt10 cDNA library constructed from chicken parathyroid gland mRNA was screened with a human preproparathyroid hormone (preproPTH) cDNA probe. Nucleotide sequence analysis of three independent clones confirmed that they encoded chicken preproPTH. This analysis, complemented by primer extension and Northern blot analysis of mRNA, demonstrated a 5'-untranslated region for chicken preproPTH of 127 nucleotides, a coding region of 357 nucleotides, and a 3'-untranslated region of approximately 2500 nucleotides. The coding sequence predicts a mature chicken PTH of 88 amino acids in contrast to the 84 amino acids of the mammalian hormones. Comparison of the avian and the mammalian hormones shows striking homology in the region of amino acids 1-32. The middle and carboxyl-terminal portions of chicken PTH, however, differ considerably from the mammalian hormones and include deletions of sequences conserved in mammalian PTH and insertions of novel peptide sequences. Comparison of the avian and mammalian structures suggests potential alterations of the mammalian sequences that may lead to altered bioactivity and/or hormone metabolism.

Amino Acid Sequence↗
PubMed1995

Cloning and expression of the alpha subunit of succinyl-CoA synthetase from Dictyostelium discoideum.

We have isolated and characterized a genomic fragment and a cDNA clone for the alpha subunit of succinyl-CoA synthetase (SCS) from Dictyostelium discoideum. The coding region predicts a protein of 310 amino acids. Direct amino acid sequence data indicate that the first 16 amino acids encoded by the cDNA comprise a mitochondrial targeting sequence. The predicted molecular mass of the mature protein is 31 kDa on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The deduced amino acid sequence of the alpha subunit from D. discoideum shows 75% identity to that from rat liver, which represents a GTP-utilizing form of SCS, and 62% identity to the Escherichia coli protein, which is an ATP-utilizing form. We have previously shown that the enzyme purified from growing D. discoideum is a GTP-utilizing enzyme. In D. discoideum, the alpha subunit is a single copy gene that is developmentally regulated. Highest levels of its mRNA are seen in growing cells, which is followed by a relatively dramatic decline at the time cells complete their aggregation program. In contrast, the level of the protein, as determined by Western blots, remains unchanged throughout development. This indicates that any developmental changes in SCS activity reflect changes in the availability of its substrates or effectors and not changes in the level of enzyme.

Amino Acid Sequence↗
PubMed1997

Molecular cloning of a bovine renal G-protein coupled receptor gene (bRGR): regulation of bRGR mRNA levels by amino acid availability.

A cDNA of 3.2 kb, encoding a putative G protein-coupled receptor and hence called bRGR1, has been isolated from a cDNA library generated from the bovine renal epithelial cell line NBL-1. This cDNA consisted of 41 base pairs of 5'-untranslated sequence, an open reading frame of 1083 base pairs, and a 2.07 kb fragment of 3'-untranslated sequence that includes a poly(dA) tail. The coding sequence predicts a protein of 361 residues. The ligand of the bRGR1 protein may be of low molecular weight, as deduced from the analysis of the predicted primary structure of the receptor protein and the comparison with other subtypes of the G protein-coupled receptor family. The amounts of bRGR1 mRNA significantly increase when NBL-1 cells are cultured in an amino acid-depleted medium. This effect can not be caused by a decrease in protein synthesis because cycloheximide did not mimic the increase in bRGR1 mRNA levels triggered by amino acid starvation. These data suggest that bRGR1 may be an amino acid-regulated gene.

Amino Acid Sequence↗
PubMed1998

Molecular cloning and expression of a cDNA encoding a human thioredoxin-like protein.

This report describes the cloning of a human cDNA that encodes a new protein (Txl, Thioredoxin-like) that belongs to the expanding family of thioredoxins based on sequence comparison of the deduced amino acid sequence. This cDNA, with a total length of 1,278 bp, consists of 205 bp of 5'-untranslated sequence (including an in frame stop codon), an open reading frame of 870 bp and a 203 bp fragment of 3'-untranslated sequence. The coding sequence predicts a protein of 289 amino acids with two distinct domains: an N-terminal domain of 105 residues homologous to the rest of mammalian thioredoxins containing the conserved active site (CGPC) and a C-terminal domain of 184 residues with no homology with any other protein in the database. Northern blot analysis indicates that the txl probe hybridizes to a 1.3 Kb mRNA and is ubiquitously expressed in human tissues with the highest expression in stomach, testis and bone marrow.

Amino Acid Sequence↗
PubMed1996

Gonadotropins beta-GtHI and beta-GtHII from the gilthead seabream, Sparus aurata.

Full-length cDNA sequences encoding the beta-subunits of the gonadotrophins GtHI and GtHII were isolated, cloned, and sequenced from a single gilthead seabream (Sparus aurata) pituitary using RACE PCR. Beta-GtHI and beta-GtHII degenerate PCR primers were designed according to regions of high amino acid sequence homology between the chum salmon and the bonito beta-GtHI or beta-GtHII. DNA sequence analysis of the cloned PCR products confirmed the presence of the predicted complete coding region as well as 5' and 3' untranslated sequences. The deduced amino acid sequences of beta-GtHI and beta-GtHII from the gilthead seabream were compared to GtH sequences from a number of teleosts. Seabream beta-GtHI shows the highest homology to the bonito beta-GtHI (76%) and the striped bass (74%), while seabream beta-GtHII is 98% homologous to the yellow fin porgy, 87% homologous to the bonito, and 93% homologous to the striped bass beta-GtHII. Northern blot analysis showed the transcripts of the gilthead seabream beta-GtHI and beta-GtHII to be about 700 and 740 bases, respectively. During the spawning season, beta-GtHI is expressed at higher levels in males than in females while beta-GtHII is expressed at similar levels in both sexes.

Amino Acid Sequence↗
PubMed1999

Duality of gonadotropin in a primitive teleost, Japanese eel (Anguilla japonica).

The duality of gonadotropin (GTH) is well-established in relatively modern teleosts. In primitive teleosts such as eel and catfish, however, only a single GTH (GTH-II) has been isolated and characterized. Therefore, the objective of this study was to clarify the duality of GTH, particularly the presence of GTH-I in primitive teleosts. We attempted to obtain a cDNA encoding the beta subunit of GTH-I from Japanese eel, Anguilla japonica, as a representative primitive teleost species. Rapid amplification of cDNA ends (RACE) polymerase chain reaction (PCR) was used to amplify GTH-Ibeta cDNA prepared from immature male Japanese eel pituitaries, and the obtained PCR products were subcloned and sequenced. A degenerate PCR primer was designed based on a highly conserved region between teleost GTH-Ibeta and mammalian FSHbeta. DNA sequence analysis of the cloned PCR products confirmed the presence of the predicted complete coding region as well as the 5' and 3' untranslated regions. The deduced amino acid sequence from these clones showed high homology to goldfish GTH-Ibeta (60%), whereas the identity between Japanese eel GTH-Ibeta and GTH-IIbeta was lower (42%). Phylogenetic analysis confirmed that Japanese eel GTH-Ibeta belonged to the teleost GTH-Ibeta group. These results provide a definitive proof of the presence of two types of GTHs (GTH-I and GTH-II) in Japanese eel, as has been shown in other teleosts. The duality of GTHs is applicable for teleosts in general. Northern blot analysis showed the transcripts of Japanese eel GTH-Ibeta and GTH-IIbeta to be 1200 and 750 bases, respectively. GTH-Ibeta gene was expressed in immature fish, while GTH-IIbeta gene was expressed in spermiating males and ovulated females, suggesting that two GTHs are differentially expressed at different sexual stages and may play separate roles in the reproductive process in Japanese eel.

Amino Acid Sequence↗
PubMed1996

Molecular cloning, tissue distribution, and chromosomal localization of human cationic amino acid transporter 2 (HCAT2).

Human cationic amino acid transporter 2 (HCAT2) was isolated from a human intestine cDNA library. The nucleotide sequence of the coding region predicts a 658-amino-acid protein with a calculated molecular weight of 71,669. As 91% of the residues are identical with those of the mouse cationic amino acid transporter 2 (MCAT2), HCAT2 seems to be a human counterpart of MCAT2. We found no isoform as was present in MCAT2. In Northern blot analysis, a single (9.0 kb) HCAT2 mRNA transcript was present in various human tissues. The highest level of expression was observed in skeletal muscle and the lowest level in the kidney. Hydropathy plots indicated that the translated protein is predicted to have 14 transmembrane domains with three potential N-glycosylation sites. Two patients with lysinuric protein intolerance (MIM No. 222700) were analyzed for HCAT2 cDNA but no mutation was detected. The HCAT2 gene was assigned to human chromosome 8p21.3-p22.

Amino Acid Metabolism, Inborn Errors↗
PubMed1998

Multiple transcriptional variants and RNA editing in C18orf1, a novel gene with LDLRA and transmembrane domains on 18p11.2.

C18orf1 is a novel brain-expressed transcript, mapping to 18p11.2. Upon further characterization, we found multiple and differentially expressed transcriptional variants. C18orf1 alpha 1, an 8.5-kb transcript, was predicted to code for a 306-amino-acid protein and a 7.1-kb 3'-untranslated region (UTR). This variant was encoded by at least six exons. Alternative transcripts included alpha 2, identical to alpha 1 but missing 18 residues, and N-terminal-truncated variants termed beta 1 and beta 2. A motif search suggested the presence of a transmembrane domain in both alpha and beta and a low-density lipoprotein receptor class A (LDLRA) domain in the alpha-specific N-terminal. In LDLR, LDLRA has been shown to be involved in binding Ca2+ and LDL, raising the possibility that C18orf1 might bind Ca2+ and an unknown ligand. We also present evidence of RNA editing in the 5'-UTR of beta 2, the first demonstration of this phenomenon in 5'-UTR.

Adult↗
PubMed2001

Molecular cloning and mapping of the human nicotinic acetylcholine receptor alpha10 (CHRNA10).

We report the isolation and initial characterization of a new member of the human nicotinic acetylcholine receptor (nAChR) subunit family, alpha10 (CHRNA10), from both inner-ear neuroepithelium and lymphoid tissue. The cDNA is 1959 nucleotides in length, with a coding region predicting a protein of 451 amino acids that is 90% identical to rat alpha10. The alpha10 gene was localized to chromosome 11p15.5. Human alpha10 was detected in human inner-ear tissue, tonsil, immortalized B-cells, cultured T-cells and peripheral blood lymphocytes using reverse transcriptase-polymerase chain reaction, Northern blot hybridization, and immunohistochemistry. We also detected the expression of the human nAChR alpha9 (CHRNA9) mRNA in these same tissues using RT-PCR and Northern blot hybridization.

Amino Acid Sequence↗
PubMed1995

The DNA sequence of equine herpesvirus 2.

The complete DNA sequence of equine herpesvirus 2 (EHV-2) strain 86/67 was determined. The genome is 184,427 bp in size and has a base composition of 57.5% G + C. Unusually for a herpesvirus, about a third of the sequence distributed in several large blocks appears not to encode proteins. The 79 open reading frames that were identified as probably polypeptide-coding are predicted to encode 77 distinct proteins. Amino acid sequence comparisons confirmed that EHV-2 is a gamma-herpesvirus that is genetically collinear with herpesvirus saimiri (HVS; a gamma 2-herpesvirus) and Epstein-Barr virus (EBV; a gamma 1-herpesvirus), with a closer relationship to the former. Moreover, EHV-2 specifies eight proteins that have counterparts in HVS but not in EBV and only a single protein that has a homologue in EBV but not in HVS (EBV BCRF1, which encodes an interleukin 10-like protein). EHV-2 also encodes three potential G protein-coupled receptors, one with a counterpart in HVS that is specific for alpha chemokines, another with a counterpart in human cytomegalovirus (a beta-herpesvirus), which is specific for beta chemokines, and a third that is assigned more tentatively and lacks detectable counterparts in other herpesviruses.

Amino Acid Sequence↗
PubMed1998

The identification and characterization of microsatellites in the compact genome of the Japanese pufferfish, Fugu rubripes: perspectives in functional and comparative genomic analyses.

Fugu rubripes (Fugu) has one of the smallest recorded vertebrate genomes and is an economic tool for comparative DNA sequence analysis. Initial characterization of 128 kb of Fugu DNA attributed the compactness of this genome, in part, to a sparseness of repetitive DNA sequence compared with mammalian genomic sequences. This paper describes a new and comprehensive analysis in which 501 theoretically possible microsatellites with a repeat unit of one to six bases were used to query two orders of magnitude more Fugu DNA (i.e. 11.338 Mb). A total of 6042 microsatellites were identified and categorized. In decreasing order, the 20 most frequently occurring microsatellites are AC, A, C, AGG, AG, AGC, AAT, AAAT, ACAG, ACGC, ATCC, AAC, ATC, AGGG, AAAG, AAG, AAAC, AT, CCG and TTAGGG. The 20 most frequently occurring microsatellites represent 81.79% of all microsatellites identified. Our results indicate that one microsatellite occurs every 1.876 kb of DNA in Fugu, 11.55% of the microsatellites are detected in open reading frames that are predicted protein coding regions. With respect to the proportion of microsatellites present in open reading frames and the total abundance (bp) of all microsatellites, the genome of Fugu is similar to the genome of many other vertebrate species. Previous estimates performed indicate that approximately 1% of many vertebrate genomes are comprized of microsatellite sequences. However, many differences prevail in the abundance and frequency of the individual microsatellite classes. Many of the frequently occurring microsatellites in Fugu are known to code in other species for regions in proteins such as transcription factors, whilst others are associated with known functions, such as transcription factor binding sites and form part of promoter regions in DNA sequences of genes. Therefore, it is likely that such repeats in genomes have a role in the evolution of genes, regulation of gene expression and consequently the evolution of species.

Animals↗
PubMed1996

Identification, molecular cloning, and transcription analysis of the Choristoneura fumiferana nuclear polyhedrosis virus spindle-like protein gene.

The Choristoneura fumiferana nuclear polyhedrosis virus spindle-like protein (slp) gene has been identified and localized immediately downstream and in the same orientation as the CfMNPV DNA polymerase gene. The slp gene is 1101 bp long, predicted to code for a 366 amino acid (42.1 kDa) polypeptide. Transcriptional analysis revealed that the CfMNPV slp gene is expressed at late times postinfection, beginning at 24 hr postinfection and is most abundantly expressed after 36 hr. Transcription initiates within a single baculovirus consensus late start site sequence (GTAAG) at position -18 relative to the translation start codon. Based on amino acid comparisons, the CfMNPV gene is closely related to other similar baculovirus genes and distantly but recognizably related to the fusolin proteins of two entomopoxviruses. The conservation of amino acid sequence, glycosylation signals and specific domains throughout the protein suggest that this gene product may play an important role in insect DNA virus replication.

Amino Acid Sequence↗
PubMed1997

Recent advances in molluscum contagiosum virus research.

Molluscum contagiosum virus (MCV) and variola virus (VAR) are the only two poxviruses that are specific for man. MCV causes skin tumors in humans and primarily in children and immunocompromised individuals. MCV is unable to replicate in tissue culture cells or animals. Recently, the DNA sequence of the 190 kbp MCV genome was reported by Senkevich et al. MCV was predicted to encode 163 proteins of which 103 were clearly related to those of smallpox virus. In contrast, it was found that MCV lacks 83 genes of VAR, including those involved in the suppression of the host response to infection, nucleotide biosynthesis, and cell proliferation. However, MCV possesses 59 genes predicted to code for novel proteins including MHC-class I, chemokine and glutathione peroxidase homologs not found in other poxviruses. The MCV genomic data allow the investigation of novel host defense mechanisms and provide new possibilities for the development of therapeutics for treatment and prevention of the MCV infection.

Animals↗
PubMed1991

The POL1 gene from the fission yeast, Schizosaccharomyces pombe, shows conserved amino acid blocks specific for eukaryotic DNA polymerases alpha.

The POL1 gene of the fission yeast, Schizosaccharomyces pombe, was isolated using a POL1 gene probe from the budding yeast Saccharomyces cerevisiae, cloned and sequenced. This gene is unique and located on chromosome II. It includes a single 91 bp intron and is transcribed into a mRNA of about 4500 nucleotides. The predicted protein coded for by the S. pombe POL1 gene is 1405 amino acid long and its calculated molecular weight is about 160,000 daltons. This peptide contains seven amino acid blocks conserved among several DNA polymerases from different organisms and shares overall 37% and 34% identity with DNA polymerases alpha from S. cerevisiae and human cells, respectively. These results indicate that this gene codes for the S. pombe catalytic subunit of DNA polymerase alpha. The comparisons with human DNA polymerase alpha and with the budding yeast DNA polymerases alpha, delta and epsilon reveal conserved blocks of amino acids which are structurally and/or functionally specific only for eukaryotic alpha-type DNA polymerases.

Amino Acid Sequence↗
PubMed1993

Molecular structure and genetic regulation of SFA, a gene responsible for resistance to formaldehyde in Saccharomyces cerevisiae, and characterization of its protein product.

A 3.7 kb DNA fragment of yeast chromosome IV has been sequenced that contains the SFA gene which, when present on a multi-copy plasmid in Saccharomyces cerevisiae, confers hyper-resistance to formaldehyde. The open reading frame of SFA is 1158 bp in size and encodes a polypeptide of 386 amino acids. The predicted protein shows strong homologies to several mammalian alcohol dehydrogenases and contains a sequence characteristic of binding sites for NAD. Overexpression of the SFA gene leads to enhanced consumption of formaldehyde, which is most probably the reason for the observed hyper-resistance phenotype. In sfa::LEU2 disruption mutants, sensitivity to formaldehyde is correlated with reduced degradation of the chemical. The SFA gene shares an 868 bp divergent promoter with UGX2 a gene of yet unknown function. Promoter deletion studies with a SFA promoter-lacZ gene fusion construct revealed negative interference on expression of SFA by upstream sequences. The upstream region between positions -145 and -172 is totally or partially responsible for control of inducibility of SFA by chemicals such as formaldehyde (FA), ethanol and methyl methanesulphonate. The 41 kDa SFA-encoded protein was purified from a hyper-resistant transformant; it oxidizes long-chain alcohols and, in the presence of glutathione, is able to oxidize FA. SFA is predicted to code for a long-chain alcohol dehydrogenase (glutathione-dependent formaldehyde dehydrogenase) of the yeast S. cerevisiae.

Alcohol Dehydrogenase↗
PubMed1996

Genomic organization of a mouse MHC class II region including the H2-M and Lmp2 loci.

The region encompassing the Ma, Mb1, Mb2, and Lmp2 genes of the mouse class II major histocompatibility complex (MHC) was sequenced. Since this region contains clusters of genes required for efficient class I and class II antigen presentation, it was interesting to search for putative additional genes in the 21 kilobase gap between the Mb1 and Lmp2 genes. Computer predictions of coding regions and CpG islands, exon trapping experiments, and cross-species comparison with the corresponding human sequence indicate that no additional functional gene is present in that stretch. However, computer analysis revealed the possible existence of an alternative 3' exon for Mb1. Except for the fact that the mouse MHC contains two Mb genes, the genomic organization of the H2-M loci was found to be almost identical to the organization of the human HLA-DM genes. The promoter regions of the Ma and Mb genes also resemble classical class II promoters, containing typical S, X, and Y boxes. Like the human genes, the three H2-M genes displayed very limited polymorphism when we compared the cDNA sequences from six haplotypes. Finally, comparison of DMB with Mb1 and Mb2, both at the genomic level and in their coding regions, suggests that the Mb gene was recently duplicated, probably only in certain rodents.

Animals↗
PubMedDate not supplied

Psychophysiological components of imagery.

McGuigan's neuromuscular model of information processing (1978a, 1978b, and 1989) was investigated by electrically recording eye movements (electro-oculograms), covert lip and preferred arm responses (electromyograms), and electroencephalograms. This model predicts that codes are generated as the lips are uniquely activated when processing words beginning with bilabial sounds like "p" or "b," as is the right arm to words like "pencil" that refer to its use. Twelve adult female participants selected for their high imagery ratings were asked to form images to three orally presented linguistic stimuli: the letter "p," the words "pencil" and "pasture," and to a control stimulus, the words "go blank." The following findings were significant beyond the 0.05 level: an increased covert lip response only to the letter "p," increased vertical eye activity to "p" and to the word "pencil," right arm response only to the word "pencil," and a decreased percentage of alpha waves from the right 02 lead only to the word "pasture." Since these covert responses uniquely occurred during specific imagery processes, it is inferred that they are components of neuromuscular circuits that function in accord with the model of information processing tested.

Adult↗
PubMed2000

The bovine alpha-glucosidase gene: coding region, genomic structure, and mutations that cause bovine generalized glycogenosis.

We report here cDNA and genomic sequence of the bovine acidic alpha-glucosidase gene, from the initiation codon to the most 3' polyadenylation signal. The 2814-bp coding sequence predicts a 937-amino acid protein, which is highly conserved compared with the human alpha-glucosidase gene (86% and 83% identity respectively). The intron/exon boundaries are also conserved between the two species. Two mutations have been identified in Brahmans, and one in Shorthorns, that lead to generalized glycogenosis. All three mutations result in premature termination of translation. Evidence is also presented for a missense mutation segregating with the Brahman population, which is responsible for a 70-80% reduction in alpha-glucosidase activity.

Amino Acid Substitution↗