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RNAdb 2.0--an expanded database of mammalian non-coding RNAs.

RNAdb is a comprehensive database of mammalian non-protein-coding RNAs (ncRNAs). There is increasing recognition that ncRNAs play important regulatory roles in multicellular organisms, and there is an expanding rate of discovery of novel ncRNAs as well as an increasing allocation of function. In this update to RNAdb, we provide nucleotide sequences and annotations for tens of thousands of non-housekeeping ncRNAs, including a wide range of mammalian microRNAs, small nucleolar RNAs and larger mRNA-like ncRNAs. Some of these have documented functions and/or expression patterns, but the majority remain of unclear significance, and include PIWI-interacting RNAs, ncRNAs identified from the latest rounds of large-scale cDNA sequencing projects, putative antisense transcripts, as well as ncRNAs predicted on the basis of structural features and alignments. Improvements to the database comprise not only new and updated ncRNA datasets, but also provision of microarray-based expression data and closer interface with more specialized ncRNA resources such as miRBase and snoRNA-LBME-db. To access RNAdb, visit http://research.imb.uq.edu.au/RNAdb.

Animals↗

Quality-of-life benefit from cochlear implantation in the elderly.

OBJECTIVE: To compare the audiologic results of geriatric patients receiving cochlear implants with younger age groups and to evaluate the quality of life after cochlear implantation in the geriatric population by means of validated quality-of-life questionnaires. STUDY DESIGN: Cross-sectional study involving 89 postlingually deafened cochlear implant subjects. SETTING: Tertiary referral center. PATIENTS: A total of 89 postlingually deafened patients were included in the study, among which were 25 patients who were aged 70 years or older. INTERVENTIONS: All patients received a cochlear implant. Subjects were implanted with either the Laura, Nucleus 24, or Med-el Combi 40+ cochlear implant systems implementing the SPEAK, ACE, CIS, or CIS+ coding strategies. MEAN OUTCOME MEASURES: Speech recognition was determined by means of phonetically balanced monosyllabic word lists. The Hearing Handicap Inventory for Adults, the Glasgow Benefit Inventory, and the scale for the prediction of hearing disability in sensorineural hearing loss were used to quantify the quality of life. RESULTS: Mean audiologic performance for the three groups increased significantly after implantation (p < 0.001). Postoperative audiologic performance of the geriatric population led to useful hearing, but these scores were significantly lower than for the younger age groups (p = 0.002). However, the quality-of-life outcomes for the geriatric group were similar to those of the younger age groups (p = 0.411 for the Hearing Handicap Inventory for Adults; p = 0.886 for the Glasgow Benefit Inventory). CONCLUSION: The results of this study prove that cochlear implantation in the elderly provides improvements in quality of life and speech understanding, similar to those for younger adult cochlear implant recipients.

Adolescent↗

Genetic control of cell-mediated immunity in the rat. I. Transfer of DTH to bacterial antigens is restricted by the B region of RT1.

The genetic basis of allogeneic restriction in the transfer of delayed type hypersensitivity (DTH) to soluble Listeria monocytogenes antigens (LMA) in rats was analysed using a set of congenic strains. The DTH parameter assayed was the localization of radiolabelled donor lymphoblasts in subcutaneous antigen stimulation sites in the ear. Sharing of the RT1,B region between the donor and host was essential for the transfer of DTH to LMA, which suggests that the RT12.B region codes for restriction elements controlling antigen recognition by TDTH cells. The recombinant haplotype RT1 rl was exceptional in that transfer of DTH to A-region matched recipients was, at least in part, possible, Measurement of donor lymphoblast localization in DTH sites afforded an opportunity to quantify allogeneic effects influencing localization of sensitized donor cells in DTH sites borne by alien recipients. Both RT1.B and RT1.A region differences could induce allogeneic effects, but they were an order of magnitude lower than those based on restricted recognition by donor T cells.

Animals↗

Design and selection of novel Cys2His2 zinc finger proteins.

Cys2His2 zinc finger proteins offer a stable and versatile framework for the design of proteins that recognize desired target sites on double-stranded DNA. Individual fingers from these proteins have a simple beta beta alpha structure that folds around a central zinc ion, and tandem sets of fingers can contact neighboring subsites of 3-4 base pairs along the major groove of the DNA. Although there is no simple, general code for zinc finger-DNA recognition, selection strategies have been developed that allow these proteins to be targeted to almost any desired site on double-stranded DNA. The affinity and specificity of these new proteins can also be improved by linking more fingers together or by designing proteins that bind as dimers and thus recognize an extended site. These new proteins can then be modified by adding other domains--for activation or repression of transcription, for DNA cleavage, or for other activities. Such designer transcription factors and other new proteins will have important applications in biomedical research and in gene therapy.

Amino Acid Motifs↗

Computational identification of transcription factor binding sites by functional analysis of sets of genes sharing overrepresented upstream motifs.

BACKGROUND: Transcriptional regulation is a key mechanism in the functioning of the cell, and is mostly effected through transcription factors binding to specific recognition motifs located upstream of the coding region of the regulated gene. The computational identification of such motifs is made easier by the fact that they often appear several times in the upstream region of the regulated genes, so that the number of occurrences of relevant motifs is often significantly larger than expected by pure chance. RESULTS: To exploit this fact, we construct sets of genes characterized by the statistical overrepresentation of a certain motif in their upstream regions. Then we study the functional characterization of these sets by analyzing their annotation to Gene Ontology terms. For the sets showing a statistically significant specific functional characterization, we conjecture that the upstream motif characterizing the set is a binding site for a transcription factor involved in the regulation of the genes in the set. CONCLUSIONS: The method we propose is able to identify many known binding sites in S. cerevisiae and new candidate targets of regulation by known transcription factors. Its application to less well studied organisms is likely to be valuable in the exploration of their regulatory interaction network.

5' Flanking Region↗

Relative locations of the centromere and imprinted SNRPN gene within chromosome 15 territories during the cell cycle in HL60 cells.

Investigations of imprinted regions provide clues that increase our understanding of the regulation of gene functions at higher order chromosomal domains. Here, the relative positions of the chromosome 15 centromere and the imprinted SNRPN gene in interphase nuclei of human myeloid leukemia HL60 cells were compared, because the homologous association of this imprinted chromosomal domain was previously observed in lymphocytes and lymphoblasts. Four targets including the chromosome 15 territory, its centromere, the SNRPN gene on this chromosome, and the nucleus, were visualized simultaneously in three-dimensionally preserved nuclei using multicolor fluorescence in situ hybridization, and the spatial distributions of these probes were analyzed with a cooled CCD camera deconvolution system. We found that preferential association of SNRPN interhomologues did not occur during the cell cycle in HL60 cells, although this gene exhibited asynchronous replication and monoallelic expression in this cells. SNRPN was found to localize at the periphery of the chromosome territories, and it preferentially faced the nuclear membrane, unlike the adjacent centromeric repeat. The SNRPN gene and the centromere were located close to each other late in S phase, reflecting that these DNA segments may be compacted into the same intranuclear subcompartments with the progress of S phase and in course of preparation for the following G(2) phase. Our results suggest that, although an imprinted gene has features similar to those observed with intranuclear localization of other gene coding sequences, the characteristic of mutual recognition of imprinted regions is determined by certain cellular regulation, and it is not necessary for the allele-specific features of an imprinted gene.

Autoantigens↗

DNA sequence analysis of the genes for the fowl adenovirus serotype 10 putative 33K and pVIII.

The nucleotide sequence and genomic location of the fowl adenovirus serotype 10 (FAV-10) putative 33K and precursor protein VIII (pVIII) genes have been determined. The total genomic region sequenced was 1814 base pairs (bp) in length with the 33K coding region occupying the sequence from nucleotides 44 to 634 and the pVIII coding region nucleotides 949 to 1689. The location of both the 33K and pVIII genes have the same positional organization as their human adenovirus (HAV) counterparts which is 3 prime (3') to the 100K gene. Along with the 100K the 33K and pVIII form the late transcription unit 4 (L4). The FAV-10 putative 33K coding region could encode a polypeptide of 196 amino acids in length with a relative molecular mass of 21.9 kilodaltons (kDa) while the pVIII produces a polypeptide of 246 amino acids with a calculated relative mass of 26.7 kDa. Two possible splice acceptor sites were identified one 5' to the 33K and one 5' to the pVIII coding regions. A putative poly A recognition sequence of AATAAA was identified 3' to the pVIII, signaling the end of the L4 transcription unit.

Amino Acid Sequence↗

More words in the neighborhood: interference in lexical decision due to deletion neighbors.

This article reports two lexical decision experiments that provide evidence for the automatic activation of deletion neighbors-that is, words that overlap with the presented word save for the deletion of one letter. Experiment 1 showed slower and less accurate no decisions for nonwords with deletion neighbors (e.g., come in scome), relative to control nonwords. Experiment 2 showed slower and less accurate yes decisions for words with higher frequency deletion neighbors, relative to control words. An important methodological implication of these results is that stimuli should be equated using a different definition of orthographic neighborhood from that which is currently the norm. The results also have significant theoretical implications for input coding schemes and the mechanisms underlying recognition of familiar words.

Decision Making↗

[Congruence of monotonous DNA/protein polymers].

The comparative study of the monotonous polynucleotides and polypeptides reveals an univocal congruence between: poly-C and poly-PRO;poly-G and poly-GLY;poly-T and poly-PHE;poly-A and poly-LYS. The hypothesis is presented of a code of congruence ruling the specific recognition of the wide groove of the DNA by proteins.

Chemical Phenomena↗

[Enzymatic synthesis and molecular cloning of the pigeon alpha-globin structural gene].

Double-stranded DNA synthesized from the pigeon globin mRNA by the subsequent actions of avian myeloblastosis virus reverse transcriptase and E. coli DNA polymerase I was split with nuclease S1 and inserted into PstI site in the plasmid pBR322 by poly(dG) times poly(dC) homopolymer extension technique using terminal deoxynucleotidyl transferase. E. coli transformants have been shown to contain pigeon globin sequences by colony hybridization with pigeon globin [32P]cDNA. The inserted DNA fragment deleted from recombinant DNA by PstI treatment hybridizes with globin cDNA. The maximal length of the inserted fragment measured in agarose gel was found to be 550--560 base pairs. Inserted sequences subjected to analysis by hybridization with alpha- and beta-[32P]cDNA have been ascribed to the pigeon alpha globin chain. EcoRI, HindIII, BglII, SalI, BamHI, PstI restriction enzymes did not cleave the inserted DNA fragment. Pigeon DNA coding alpha-globin chain contains recognition sites for AluI, HindII and HaeIII restriction enzymes. Part of the recombinant clones remains resistant to ampicillin and therefore in some of these clones the globin gene could be expressed.

Animals↗

Proficiency testing in clinical cytogenetics. A 6-year experience with photographs, fixed cells, and fresh blood.

The College of American Pathologists and the American Society of Human Genetics offer a proficiency testing program in clinical cytogenetics. Two hundred twenty-five laboratories now provide data for this survey, which was begun in 1986. Challenges have consisted of photographed metaphases, fixed lymphoblastoid cell suspensions, fresh peripheral blood, and disarranged karyotypes. The "correct" response was based on 80% or greater consensus among either the referees or the participants. Referee laboratories performed better than participants. More laboratories were able to report accurate recognition of abnormalities by using a coded list than could write the interpretation in standardized nomenclature. Deletions, unbalanced translocations, and inversions were more difficult challenges than balanced translocations or trisomies. Prenatal and lymphocyte challenges were more likely to result in consensus than were bone marrow challenges. Participants performed best on whole-blood challenges. Fixed cell suspensions were less satisfactory. Excellent quality case material is essential for a successful challenge. A grading system has been devised to separate artifacts of the survey process from proficiency variables.

Chromosome Aberrations↗

Molecular and immunologic characterization of a highly cross-reactive two EF-hand calcium-binding alder pollen allergen, Aln g 4: structural basis for calcium-modulated IgE recognition.

Serum IgE was used to isolate a cDNA coding for a 9.4-kDa two EF-hand calcium-binding allergen, Aln g 4, from a lambda gt11 expression cDNA library constructed from alder (Alnus glutinosa) pollen. rAln g 4 was overexpressed in Escherichia coli and purified to homogeneity. It reacted with serum IgE from 18% of pollen-allergic patients (n = 122); shared IgE epitopes with homologous allergens present in tree, grass, and weed pollens; and thus belongs to a family of highly cross-reactive pollen allergens. Exposure of two E. coli-expressed rAln g 4 fragments comprising amino acids 1-41 and 42-85 to patients' IgE Abs, as well as to a rabbit antiserum raised against purified rAln g 4, indicated that most of the B cell epitopes reside in the N-terminal portion of the protein. IgE recognition of Aln g 4 was strongly modulated by the presence or absence of calcium. Circular dichroism analysis of rAln g 4 revealed that the protein consisted mostly of alpha helical secondary structure and possessed a remarkable thermal stability and refolding capacity, a property that was greatly reduced after calcium depletion. Circular dichroism analysis of the calcium-bound and apo form of rAln g 4 indicated that calcium-induced modulation of IgE binding could be due to changes in the protein conformation. Purified rAln g 4 elicited dose-dependent basophil histamine release and immediate type skin reactions in sensitized patients. It may hence be useful for allergy diagnosis and for specific immunotherapy.

Allergens↗

Tilted letters and tilted words: a possible role for principal axes in visual word recognition.

Subjects made lexical decisions to columnar letter strings in which every letter was either upright or tilted 90 degrees clockwise as if the whole letter string had been rotated from the horizontal. Lexical decisions were faster in the latter case. The advantage for the tilted format was also found when all strings were presented in aLtErNaTiNg CaSe or in uppercase, so this advantage cannot be due to preservation of the tilted words' global shape. The cost for the upright-letter format increased with the number of letters in the columnar strings. These data suggest that words recognition may involve shape description or position coding relative to a reference frame based on the principal axis of the letter string.

Adult↗

Word attributes and lateralization revisited: implications for dual coding and discrete versus continuous processing.

Three attributes of words are their imageability, concreteness, and familiarity. From a literature review and several experiments, I previously concluded (Boles, 1983a) that only familiarity affects the overall near-threshold recognition of words, and that none of the attributes affects right-visual-field superiority for word recognition. Here these conclusions are modified by two experiments demonstrating a critical mediating influence of intentional versus incidental memory instructions. In Experiment 1, subjects were instructed to remember the words they were shown, for subsequent recall. The results showed effects of both imageability and familiarity on overall recognition, as well as an effect of imageability on lateralization. In Experiment 2, word-memory instructions were deleted and the results essentially reinstated the findings of Boles (1983a). It is concluded that right-hemisphere imagery processes can participate in word recognition under intentional memory instructions. Within the dual coding theory (Paivio, 1971), the results argue that both discrete and continuous processing modes are available, that the modes can be used strategically, and that continuous processing can occur prior to response stages.

Adult↗

Cloning and expression in E. coli of a synthetic gene for the bacteriocidal protein caltrin/seminalplasmin.

A synthetic gene coding for the bacteriocidal protein caltrin/seminalplasmin was constructed and expressed in Escherichia coli as a fusion with beta-galactosidase. The gene was designed with a recognition site for the plasma protease, Factor Xa, coded for immediately prior to the N-terminus of caltrin. The beta-galactosidase-caltrin fusion protein was cleaved with Factor Xa to give caltrin, which was identified by its size on SDS-PAGE, its ability to react with an antiserum raised to the N-terminal nonapeptide of caltrin and its N-terminal amino acid sequence. After partial purification, synthetic caltrin was found to be active in an assay involving inhibition of growth of E.coli.

Antineoplastic Agents↗

Analysis of the yeast genome: identification of new non-coding and small ORF-containing RNAs.

The genome sequences from increasing numbers of organisms allow for rapid and organized examination of gene expression. Yet current computational-based paradigms for gene recognition are limited and likely to miss genes expressing non-coding RNAs or mRNAs with small open reading frames (ORFs). We have utilized two strategies to determine if there are additional transcripts in the yeast Saccharomyces cerevisiae that were not identified in previous analyses of the genome. In one approach, we identified strong consensus polymerase III promoters based on sequence, and determined experimentally if these promoters drive the expression of an RNA polymerase III transcript. This approach led to the identification of a new, non-essential 170 nt non-coding RNA. An alternative strategy analyzed RNA expression from large sequence gaps>2 kb between predicted ORFs. Fifteen unique RNA transcripts ranging in size from 161 to 1200 nt were identified from a total of 59 sequence gaps. Several of these RNAs contain unusually small potential ORFs, while one is clearly non-coding and appears to be a small nucleolar RNA. These results suggest that there are likely to be additional previously unidentified non-coding RNAs in yeast, and that new paradigms for gene recognition will be required to identify all expressed genes from an organism.

Base Sequence↗

The emerging functionality of endogenous lectins: A primer to the concept and a case study on galectins including medical implications.

Biochemistry textbooks commonly make it appear that it is a foregone conclusion that the hardware of biological information storage and transfer is confined to nucleotides and amino acids, the letters of the genetic code. However, the remarkable talents of a third class of biomolecules are often overlooked. For example, one of them far surpasses the building blocks of nucleic acids and proteins in terms of theoretical coding capacity by oligomer formation. Although often exclusively assigned to duties in energy metabolism, carbohydrates as part of cellular glycoconjugates (glycoproteins, proteoglycans, glycolipids) have, in fact, other important tasks. Currently, they are increasingly gaining recognition as an operative high-density information coding system. An elaborate enzymatic machinery enables cells to be versatile enough to produce a glycan profile (glycome) that is as characteristic as a fingerprint. Moreover, swift modifications during dynamic processes, such as differentiation or malignant transformation, are readily possible. The translation of the information presented in oligosaccharide determinants to biological responses is carried out by lectins. Recognition of foreign glycosignatures in innate immunity, regulation of cell-cell/matrix interactions, cell migration or growth, and intra- and intercellular glycan routing etc represent physiologically far-reaching lectin-carbohydrate functionality. The classification of endogenous lectins is guided by sequence alignments and conservation of distinct structural traits. For example, a jelly-roll-like folding pattern and maintenance of key residue positioning involved in stacking and C-H/pi-interactions as well as directional hydrogen bonds to the 1-galactoside ligands are common denominators among galectins. Biochemical and biophysical studies are beginning to unravel the intricacies of the selection of a limited set of endogenous ligands, such as certain integrins or ganglioside GM1, and combined with biological cell experiments, its relevance for cell sociology, e.g. in growth regulation and tumor cell invasion or activated T cell apoptosis. Histopathological monitoring accompanies the biological cell investigations, linking expression of certain family members to tumor progression or suppression. Further insights into the functional consequences of the sugar code's translation are thus expected to have notable repercussions for diagnostic and therapeutic procedures.

Amino Acid Sequence↗

Structure and expression of a gene encoding heat-shock protein Hsp70 from the Oomycete fungus Bremia lactucae.

A gene encoding a protein homologous to a 70-kDa heat-shock protein (Hsp70) was isolated from Bremia lactucae and its structure and pattern of expression were determined. This is the first report on the structure of a protein-coding gene from an Oomycete fungus. The cloned gene is a member of a small multigene family. The level of hsp70 mRNA in germlings increases from a low constitutive level in response to heat or cold treatment. A high level of the mRNA is also detected in spores. The hsp70 gene is expressed as a primary transcript of 2241 nucleotides (nt) and contains a continuous open reading frame of 2025 nt. Near the C terminus of the coding sequence is an unusual region that contains repeated enhancer-like sequences. This insert has not been described in other hsp70 genes and is not an intron. Upstream from the 5' terminus of the mRNA are multiple CCAAT motifs, a sequence similar to a consensus heat-shock regulatory element, and an A + T-rich putative 'TATA' box. A canonical polyadenylation recognition sequence is present downstream from the coding sequence. The deduced amino acid sequence is equally similar to yeast and maize Hsp70, providing further evidence of the dissimilarity between Oomycetes and true fungi. The cloning of this gene is part of our strategy to develop a transformation system for B. lactucae.

Amino Acid Sequence↗