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PubMed1991

Tetrahymena thermophila acidic ribosomal protein L37 contains an archaebacterial type of C-terminus.

We have cloned and characterized a Tetrahymena thermophila macronuclear gene (L37) encoding the acidic ribosomal protein (A-protein) L37. The gene contains a single intron located in the 3'-part of the coding region. Two major and three minor transcription start points (tsp) were mapped 39 to 63 nucleotides upstream from the translational start codon. The uppermost tsp mapped to the first T in a putative T. thermophila RNA polymerase II initiator element, TATAA. The coding region of L37 predicts a protein of 109 amino acid (aa) residues. A substantial part of the deduced aa sequence was verified by protein sequencing. The T. thermophila L37 clearly belongs to the P1-type family of eukaryotic A-proteins, but the C-terminal region has the hallmarks of archaebacterial A-proteins.

Amino Acid Sequence↗
PubMed1993

Human L7a ribosomal protein: sequence, structural organization, and expression of a functional gene.

A cDNA coding for the human L7a ribosomal protein (r-protein) was used to isolate the corresponding gene by screening two human genomic libraries constructed in bacteriophage lambda and in a cosmid vector. One of the cosmid clones isolated, cos1.1, contains the whole L7 alpha gene, composed of eight exons and seven introns spanning 3226 bp. As in other mammalian housekeeping genes, the promoter and the first exon of the L7 alpha reside within a CpG-rich island. Furthermore, similar to the other higher eukaryote r-protein-encoding genes characterized so far, the human L7 alpha gene has a C as the major transcriptional start point localized in a pyrimidine-rich region and lacks a canonical TATA sequence. We show that 130 bp of the human L7 alpha gene 5'-flanking region represent the minimal element required to promote its transcription. This element is strikingly conserved between the mouse and human L7 alpha genes. Finally, a comparison of the human L7 alpha gene coding sequence and the predicted amino acid (aa) sequence with the sequences of mouse L7a, rat L7a, and the homologous yeast L4 shows that the aa sequence has been highly conserved during evolution.

Amino Acid Sequence↗
PubMed1996

Expression of alternate growth hormone receptor messenger RNA in ovary and uterus of cattle.

Growth hormone (GH) receptor mRNA is found within the corpus luteum and endometrium of cattle. However, binding sites for placental lactogen (PL) but not GH are found within these tissues. The objectives were to isolate cDNA for the GH receptor within the reproductive tissues of cattle and to examine these cDNA as potential variants of the GH receptor that bind PL. Ten cDNA clones were isolated from a bovine endometrial cDNA library with a 32P-labeled cDNA of the GH receptor extracellular domain. On the basis of restriction enzyme digestion, 2 of the 10 cDNA clones contained exon 1. The DNA sequence of these clones was determined by dideoxy nucleotide sequencing. The exon 1 DNA sequence of each clone (exon 1B) was different from the previously reported exon 1 for the bovine GH receptor cDNA isolated from liver (exon 1A). Analyses of these cDNA sequences showed that exon 1B contained significant homology with placental forms of the GH receptor found in mouse and human. Unlike the human cDNA, the bovine cDNA isolated from endometrium contained an intact third exon. Amplification of GH receptor mRNA by reverse transcriptase polymerase chain reaction, with exon 1A- and 1B-specific forward primers, showed that exon 1B was expressed in liver, corpus luteum, ovary, endometrium, and myometrium. Exon 1A was found almost exclusively in liver, and little was found in reproductive tissues. The predicted initiation of protein coding for the GH receptor was within the second exon and was not changed by the splicing of the alternate first exon. This suggests that the alternate mRNA results in the expression of intact GH receptor protein that is similar to that found within liver. Alternative promoters (1B) may control the expression of the receptor outside the liver. Furthermore, mechanisms other than the differential splicing of GH receptor protein may dictate the specificity of PL binding within the endometrium and corpus luteum.

Animals↗
PubMed1990

A transcribed gene in an intron of the human factor VIII gene.

We have identified a CpG island contained within the largest factor VIII intron. This island is associated with a 1.8-kb transcript and, unlike factor VIII, is produced abundantly in a wide variety of cell types. The nested gene is oriented in a direction opposite to that of factor VIII and contains no intervening sequences. A cDNA of 1739 bases was isolated from a human liver library and found to have a GC-rich, long open reading frame. Two computer-assisted methods (Fickett TESTCODE and Staden-McLachlan codon usage) predict that the gene codes for a protein. Two other copies of this gene are located within 1.1 Mb of the factor VIII gene. Northern blot analysis of RNA isolated from hemophilia patients deleted for factor VIII sequences has shown that both the intron gene and at least one other copy of the gene are transcribed. A homologous, transcribed sequence is also present in mice.

Amino Acid Sequence↗
PubMed2005

Cytochrome P450 2E polymorphism in feline liver.

Only one isoform of cytochrome P450 (CYP) 2E subfamily was known in human and various animals. Three cDNAs corresponding to CYP 2E subfamily members (CYP2E-a, CYP2E-b and CYP2E-c) were obtained from feline liver. These cDNAs each had a 1488-bp nucleotide coding region encoding a predicted amino acid sequence of 495 residues. Eleven amino acid substitutions were observed between CYP2E-a and CYP2E-b, but only one substitution between CYP2E-b and CYP2E-c. The CO difference spectrums about 450 nm wave length and similar values of Vmax and Km of 6-hydoxygenase activity toward chlorzoxazone were observed in all three isoforms expressed in AH22 yeast cells. By PCR-RFLP, mRNA of the CYP2E-a was found to be expressed in liver, mononuclear cells, kidney, lung, stomach, intestine and pancreas, whereas CYP2E-b and CYP2E-c were expressed mainly in the liver and mononuclear cells. Expression of CYP2E-a was observed in the livers of all felines tested, but CYP2E-b and CYP2E-c were not expressed in all cats. The sequences of two different introns between exons I and II and between exons VII and VIII were obtained in genomic DNA from the feline liver. Based on these results, we conclude that cats have two highly similar CYP2E genes.

Amino Acid Sequence↗
PubMed2004

Zebrafish cellular nucleic acid-binding protein: gene structure and developmental behaviour.

Here we analyse the structural organisation and expression of the zebrafish cellular nucleic acid-binding protein (zCNBP) gene and protein. The gene is organised in five exons and four introns. A noteworthy feature of the gene is the absence of a predicted promoter region. The coding region encodes a 163-amino acid polypeptide with the highly conserved general structural organisation of seven CCHC Zn knuckle domains and an RGG box between the first and the second Zn knuckles. Although theoretical alternative splicing is possible, only one form of zCNBP is actually detected. This form is able to bind to single-stranded DNA and RNA probes in vitro. The analysis of zCNBP developmental expression shows a high amount of CNBP-mRNA in ovary and during the first developmental stages. CNBP-mRNA levels decrease while early development progresses until the midblastula transition (MBT) stage and increases again thereafter. The protein is localised in the cytoplasm of blastomeres whereas it is mainly nuclear in developmental stages after the MBT. These findings suggest that CNBP is a strikingly conserved single-stranded nucleic acid-binding protein which might interact with maternal mRNA during its storage in the embryo cell cytoplasm. It becomes nuclear once MBT takes place possibly in order to modulate zygotic transcription and/or to associate with newly synthesised transcripts.

Alternative Splicing↗
PubMed2005

Characterization of LST-R533: uncovering a novel repetitive element in Leishmania.

We have previously isolated and sequenced a novel repetitive element, now named LST-R533, which is present in four different regions of one extremity of Leishmania major chromosome 20. The repeats are polymorphic in size, ranging from 367 to 533 bp and contain an internal 81 bp sequence with highly conserved segments (14-81 bp long) dispersed throughout the parasite's genome. These sequences were not found in coding regions of any predicted gene in L. major Friedlin genome, but are part of untranslated regions of some Leishmania transcripts. Analysis of the 81 bp sequence revealed significant degrees of identity with retrotransposons described in several other organisms. The presence of the sequence in other species from genus Leishmania was determined by Southern hybridisation and DNA sequencing. This analysis indicated the conservation of the 81-nucleotide element in all the Leishmania species evaluated. No sequences corresponding to LST-R533 or the 81 bp element were found on either Trypanosoma brucei or Trypanosoma cruzi databanks.

Animals↗
PubMed2004

Number processing and basal ganglia dysfunction: a single case study.

Numerical processing has never been investigated in a case of Fahr's disease (FD) and only rarely in cases of basal ganglia dysfunction. The study describes the cognitive decline of a pre-morbidly high-functioning patient (medical doctor) affected by FD and his difficulties in number processing. A MRI scan revealed bilateral calcifications in the basal ganglia and a brain PET showed a massive reduction of glucose metabolism in the basal ganglia and both frontal lobes, but no other brain abnormalities. The patient's cognitive deficits included impairments in problem solving, in cognitive set shifting and in mental flexibility, as well as in verbal memory. These deficits are attributed to the disruption of the dorsolateral prefrontal circuit involving the basal ganglia. In number processing, the patient showed a severe deficit in the retrieval of multiplication facts, deficits in all tasks of numerical problem solving and in the execution of complex procedures. Importantly, he also showed a dense deficit in conceptual knowledge, which concerned all test conditions and all operations. The findings confirm the predictions of the triple code model in so far, as a disruption of cortico-subcortical loops involving the basal-ganglia may lead to specific deficits in fact retrieval. However, no verbal deficit, as assumed in the triple code model and reported in similar cases, could be observed. The present findings further add to current knowledge on numerical processing, showing how fronto-executive dysfunction may disrupt conceptual understanding of arithmetic. This study shows that not only parietal lesions may lead to severe deficits in conceptual understanding, but that basal ganglia lesions leading to frontal dysfunction may have a devastating effect.

Basal Ganglia Diseases↗
PubMed2005

Saliency from orthogonal velocity component in texture segregation.

We found that a moving target line, more-vertical than 45 deg-oriented background lines, pops-out (d'=1.2) although it moves at the same speed of background elements and although it is invisible in static presentation (d'=.7). We suggest that the moving more-vertical target is more salient because the motion system responds to the orthogonal-velocity-component (V(perpendicular)=Delta d/Delta t sin theta) that is larger for the more-vertical target than for distracters. However, motion does not produce high d' when the target is more horizontal than background (d'=.6). This result is not expected if saliency resulted from the sum of saliency of orientation and motion independently coded but is instead predicted by visual search asymmetry. A line length effect on the moving target saliency also suggests that V(perpendicular) is extracted on the whole line and this operation is facilitated by line length in the same way for more-vertical and more-horizontal targets. Altogether, these results demonstrate that speed-based segmentation operating on V(perpendicular) not only affects speed and direction of motion discrimination, as previously demonstrated, but accounts for high saliency of image features that would otherwise prove undetectable of the basis of orientation-contrast.

Discrimination, Psychological↗
PubMed1997

Isolation of chicken alpha ENaC splice variants from a cochlear cDNA library.

Three splice variants of the alpha subunit of the amiloride-sensitive epithelial sodium channel (alpha ENaC) have been isolated from a chicken cochlear cDNA library. A PCR product, generated from the cochlear library using degenerate primers to regions of homology between the rat alpha ENaC and the degenerin Mec-4, was used as a probe. The three splice variant cDNAs with sizes of 2321, 3399 and 3845 bp correspond to transcripts of 2.5, 3.5 and 3.9 kb as detected by Northern blot analysis. The 3399 bp clone differs from the 2321 clone solely by the addition of 1079 bases in the 3'-non-coding region. Both these cDNAs code for an identical predicted protein of 637 amino acids which has 68% similarity to the rat alpha ENaC, and is probably the chicken homologue of alpha ENaC. The third cDNA of 3845 bp is similar to the 3399 bp clone but includes two exons within the open reading frame. The first of these exons introduces a premature stop codon resulting in a truncated predicted protein of 434 amino acids. Northern blot analysis shows expression of the 2.5 and 3.5 kb transcripts in cochlea and colon, the 2.5 kb transcript in cartilage, whilst the 3.9 kb transcript is only detected in cochlea. No expression is detected in brain, liver, and heart nor, most notably, in lung or kidney.

Alternative Splicing↗
PubMed2001

Molecular cloning and expression of the cDNA encoding feline granulocyte colony-stimulating factor.

Both genomic DNA and cDNA of the feline granulocyte colony-stimulating factor (G-CSF) gene were cloned from CRFK cells. Southern blot analysis showed that the haploid genome contains a single copy of the G-CSF gene. The RT-PCR analysis of several feline cell lines revealed expression of G-CSF mRNA in response to lipopolysaccharide stimulation. Sequence analysis of genomic and cDNA clones indicated that the intron-exon junction structure is conserved between the human and the feline G-CSF genes. The G-CSF coding region encodes a predicted protein of 195 amino acids including a signal sequence of 21 amino acids. The feline G-CSF amino acid sequence shares a high degree of identity with the canine (90.8%), human (87.4%), ovine (83.9%), bovine (82.8%), porcine (80.5%), murine (70.7%) and rat (66.8%) G-CSF. The feline G-CSF expressed in insect cells using recombinant baculovirus vector was biologically active as measured in a proliferation assay using NFS-60 cells and an induction assay of leukocytes in cats.

Amino Acid Sequence↗
PubMed1997

Distributed Learning, Recognition, and Prediction by ART and ARTMAP Neural Networks.

A class of adaptive resonance theory (ART) models for learning, recognition, and prediction with arbitrarily distributed code representations is introduced. Distributed ART neural networks combine the stable fast learning capabilities of winner-take-all ART systems with the noise tolerance and code compression capabilities of multilayer perceptrons. With a winner-take-all code, the unsupervised model dART reduces to fuzzy ART and the supervised model dARTMAP reduces to fuzzy ARTMAP. With a distributed code, these networks automatically apportion learned changes according to the degree of activation of each coding node, which permits fast as well as slow learning without catastrophic forgetting. Distributed ART models replace the traditional neural network path weight with a dynamic weight equal to the rectified difference between coding node activation and an adaptive threshold. Thresholds increase monotonically during learning according to a principle of atrophy due to disuse. However, monotonic change at the synaptic level manifests itself as bidirectional change at the dynamic level, where the result of adaptation resembles long-term potentiation (LTP) for single-pulse or low frequency test inputs but can resemble long-term depression (LTD) for higher frequency test inputs. This paradoxical behavior is traced to dual computational properties of phasic and tonic coding signal components. A parallel distributed match-reset-search process also helps stabilize memory. Without the match-reset-search system, dART becomes a type of distributed competitive learning network.

Journal Article↗
PubMed2002

Feature-based anticipation of cues that predict reward in monkey caudate nucleus.

A subset of caudate neurons fires before cues that instruct the monkey what he should do. To test the hypothesis that the anticipatory activity of such neurons depends on the context of stimulus-reward mapping, we examined their activity while the monkeys performed a memory-guided saccade task in which either the position or the color of a cue indicated presence or absence of reward. Some neurons showed anticipatory activity only when a particular position was associated with reward, while others fired selectively for color-reward associations. The functional segregation suggests that caudate neurons participate in feature-based anticipation of visual information that predicts reward. This neuronal code influences the general activity level in response to visual features without improving the quality of visual discrimination.

Animals↗
PubMed2002

Tonic and spillover inhibition of granule cells control information flow through cerebellar cortex.

We show that information flow through the adult cerebellar cortex, from the mossy fiber input to the Purkinje cell output, is controlled by furosemide-sensitive, diazepam- and neurosteroid-insensitive GABA(A) receptors on granule cells, which are activated both tonically and by GABA spillover from synaptic release sites. Tonic activation of these receptors contributes a 3-fold larger mean inhibitory conductance than GABA released synaptically by high-frequency stimulation. Tonic and spillover inhibition reduce the fraction of granule cells activated by mossy fiber input, generating an increase of coding sparseness, which is predicted to improve the information storage capacity of the cerebellum.

Action Potentials↗
PubMed2002

Genome sequencing: and then there were six.

The genome of the fission yeast Schizosaccharomyces pombe has been sequenced, bringing the number of sequenced eukaryotic genomes to six. Analysis of the sequence predicts only 4824 protein coding genes, the smallest number yet recorded for a free-living eukaryote.

Animals↗
PubMedDate not supplied

Monte Carlo methods for the in vivo analysis of cisplatin using X-ray fluorescence.

A Monte Carlo method has been used to model the measurement of cisplatin uptake with in vivo X-ray fluorescence. A user-code has been written for the EGS4 Monte Carlo system that incorporates linear polarisation and multiple element fluorescence extensions. The yield of fluorescent photons to the mainly Compton scattered background is computed for our detector arrangement. The detector consists of a mutually orthogonal arrangement of X-ray tube, aluminium polariser and high purity germanium scintillation detector. The influence of tube voltage on the minimum detectable concentration is modelled for 100 through 150 kVp X-radiation. The code is able to predict absorbed dose to the patient which will influence the optimal choice of tube voltage. The influence of alterations to collimator design and scatterer construction can also be examined. A minimum detectable concentration of 50 ppm is determined from measurements with a 115 kVp X-ray source and a 615 ppm cisplatin sample in a water phantom.

Cisplatin↗
PubMed1989

Bleomycin hydrolase: molecular cloning, sequencing, and biochemical studies reveal membership in the cysteine proteinase family.

Bleomycin (BLM) hydrolase catalyzes the inactivation of the antitumor drug BLM and is believed to protect normal and malignant cells from BLM toxicity. The normal physiological function of BLM hydrolase is not known. We now provide evidence for its membership in the cysteine proteinase family. BLM hydrolase was purified to homogeneity from rabbit lungs, and a partial amino acid sequence was determined from a tryptic digest peptide. On the basis of this sequence a 36-mer oligonucleotide was synthesized. The 36-mer oligonucleotide probe hybridized to a single mRNA species of 2.5 kb from several species and was used to isolate an 832-bp cDNA insert from a lambda gt11 rabbit liver cDNA library. This insert encoded the tryptic digest peptide previously identified in rabbit lung BLM hydrolase by amino acid sequencing. Analysis of the predicted amino acid sequence coded by the 832-bp BLM hydrolase cDNA fragment indicated no significant homology with any currently known proteins except for a 15 amino acid portion, which displayed remarkable homology with the active site of cysteine proteinases. Within this active-site region, 10 of the amino acid residues of papain and 9 of aleurain, cathepsin H, and cathepsin L were identical with those of rabbit liver BLM hydrolase. The catalytic cysteine of thiol proteinases was also conserved in BLM hydrolase, and cysteine proteinase specific inhibitors, such as E-64, were found to be potent inhibitors of BLM hydrolase activity. Furthermore, bleomycin hydrolase exhibited cathepsin H like enzymatic activity. Bleomycin hydrolase had, however, no significant cathepsin B or L activities.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗
PubMed2003

In situ trace metal speciation in lake surface waters using DGT, dialysis, and filtration.

In situ measurements of Fe and Mn by dialysis and diffusive gradients in thin-films (DGT) in 5 lakes (pH 4.7-7.5, ionic strength 0.3-5 mmol l(-1)) and Cu and Zn in an acidic and circumneutral lake were compared to results from on site filtration. For the most acidic lake (pH 4.7) all measurements agreed, indicating an absence of colloids and negligible complexation by organic matter. There was little difference in the Mn concentrations measured by the three techniques for any lake, consistent with it being free from complexation. Zn measured by dialysis in circumneutral water was only slightly higher than DGT measurements, appropriate to only partial complexation. Substantial differences between dialysis and DGT for Cu were consistent with complexation by fulvic and humic substances, though not to the extent predicted by the speciation code WHAM. To achieve a good fit it was necessary to adjust the pK for Cu-fulvic binding from 0.8 to 1.3 and to assume that fulvic substances dominated. The presence of low molecular weight strong binding ligands would also be consistent with the data. Differences between the three measurement methods were greatest for Fe, attributable to the presence of large oxyhydroxide colloids, organic complexation and low molecular weight, reactive hydrolysis products. Fe and Mn concentrations measured by DGT on samples returned to the laboratory were much lower than in situ concentrations, illustrating the need for in situ measurements. While use of two in situ techniques provided useful information on the speciation of these natural waters, further refinements are required for unambiguous characterization of the solution. The use of DGT with a more restricted gel that excludes complexes with humic substances should provide complementary information to in situ dialysis.

Dialysis↗