Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “ENCODE”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 793 records · Page 44Linked to original sources

Message-encoding techniques for augmentative communication systems: the recall performances of adults with severe speech impairments.

This study investigated the cognitive and linguistic processing demands of message-encoding techniques used to retrieve prestored messages from computer-based augmentative communication systems. Twelve physically disabled adults with severe speech impairments participated in six counterbalanced experimental conditions, consisting of three encoding techniques (salient letter, letter category, and iconic codes), each in a personalized condition in which subjects selected their own codes and in a nonpersonalized condition in which codes were preselected by a clinician. In each of the experimental conditions, the subjects participated in five learning and testing sessions. During these sessions, the subjects learned the codes for lists of 80 messages, half of which involved concrete referents and half abstract concepts. Results indicated that the subjects were more accurate in recalling the codes to retrieve preprogrammed messages when using the two letter encoding techniques than when using the iconic technique. No reliable differences were found between recall performances with personalized codes and with nonpersonalized ones for any of the three encoding techniques. Code recall improved consistently across the series of learning sessions; there were no significant differences in the rates of learning across the encoding techniques. Subjects were found to be more accurate at recalling the codes to retrieve concrete messages than those to retrieve abstract messages.

Adult↗

Absence of immunoglobulin E synthesis and airway eosinophilia by vaccination with plasmid DNA encoding ProDer p 1.

BACKGROUND: Various studies have shown that immunization with naked DNA encoding allergens induces T helper 1(Th1)-biased non-allergic responses. OBJECTIVE: To evaluate the polarization of the immune responses induced by vaccinations with plasmid DNA encoding the major mite allergen precursor ProDer p 1. METHODS: A DNA vaccine was constructed on the basis of a synthetic cDNA encoding ProDer p 1 with optimized codon usage. The immunogenicity of ProDer p 1 DNA in CBA/J mice was compared with that of purified natural Der p 1 or recombinant ProDer p 1 adjuvanted with alum. Vaccinated mice were subsequently exposed to aerosolized house dust mite extracts to provoke airway inflammation. The presence of inflammatory cells was examined in bronchoalveolar lavage (BAL) fluids and allergen-specific T cell reactivity was measured. RESULTS: Naive mice immunized with ProDer p 1 DNA developed Th1 immune responses characterized by high titres of specific IgG2a antibodies, low titres of specific IgG1 and, remarkably, the absence of anti-ProDer p 1 IgE. No specific responses were observed in animals vaccinated with the blank DNA vector. By contrast, natural Der p 1 or recombinant ProDer p 1 adsorbed to alum induced pronounced Th2 allergic responses with strong specific IgG1 and IgE titres. Spleen cells from DNA ProDer p 1-vaccinated mice secreted high levels of IFN-gamma and low production of IL-5. Conversely, both adjuvanted allergens stimulated typical Th2-type cytokine profile characterized by high and low levels of IL-5 and IFN-gamma, respectively. Whereas BAL eosinophilia was clearly observed in Der p 1-immunized animals, ProDer p 1 DNA as well as ProDer p 1 vaccinations prevented airway eosinophil infiltrations. CONCLUSIONS: These results suggest that vaccination with DNA encoding ProDer p 1 effectively fails to induce the allergen-induced IgE synthesis and airway cell infiltration. Plasmid DNA encoding ProDer p 1 may provide a novel approach for the treatment of house dust mite allergy.

Animals↗

Human monoclonal antibodies encoded by the V4-34 gene segment show cold agglutinin activity and variable multireactivity which correlates with the predicted charge of the heavy-chain variable region.

We have characterized the reactivities of a panel of V4-34-encoded human IgM monoclonal antibodies (mAb) which bind the erythrocyte Rh D antigen, derived from an immunized individual. These were compared with the specificities of V4-34-encoded autoantibodies with I/i reactivity produced from patients with cold agglutinin disease (CAD), and other V4-34-encoded autoantibodies. The antibodies were evaluated for cold agglutinin activity using haemagglutination tests, immunofluorescence microscopy for reactivity with tissue components, and in solid phase radiobinding assays with purified antigens. We found that (i) cold agglutinin activity was a property of all the V4-34-encoded mAb (ii) the cold agglutinins from CAD patients were generally monospecific for I/i whereas most of the anti-D and the other V4-34-encoded mAb displayed multireactive properties, frequently binding to strongly acidic antigens (iii) computation of the net charge of the heavy-chain V regions showed that the multireactive mAb were generally more positively charged than the monospecific cold agglutinins, which could contribute to their multireactive phenotype. The involvement of charge interactions was further indicated by the effects of pH and ionic strength on the immunofluorescence staining patterns.

Adult↗

Transcriptional regulation of zwf, encoding glucose-6-phosphate dehydrogenase, from the cyanobacterium Nostoc punctiforme strain ATCC 29133.

The gene encoding glucose-6-phosphate dehydrogenase (G6PD), zwf, in Nostoc punctiforme strain ATCC 29133 is part of a four-gene operon that also encodes fructose bisphosphatase (fbp), transaldolase (tal) and a gene product termed OpcA, which is contranscribed with zwf and essential for G6PD activity. The effect of exogenous nitrogen and carbon sources on transcription of these genes was investigated. Growth in the presence of ammonium yielded low levels of transcripts encoding all genes of the operon, while growth under nitrogen-fixing conditions resulted in a large increase of transcripts encoding for fbp and zwf-opcA. When cells are grown in the presence of fructose, levels of transcripts encoding tal and zwf-opcA were increased, relative to levels in ammonium-grown cells. These results indicate that this facultatively heterotrophic cyanobacterium can respond to changes in its environment by altering transcription of genes involved in carbon catabolism. Primer extension identified five 5' ends corresponding to the major regulated transcripts which we conclude arise from independent transcriptional start points.

Bacterial Outer Membrane Proteins↗

The Per regulon of enteropathogenic Escherichia coli : identification of a regulatory cascade and a novel transcriptional activator, the locus of enterocyte effacement (LEE)-encoded regulator (Ler).

Enteropathogenic Escherichia coli (EPEC) is the prototype organism of a group of pathogenic Gram-negative bacteria that cause attaching and effacing (AE) intestinal lesions. All EPEC genes necessary for the AE phenotype are encoded within a 35.6 kb pathogenicity island termed the locus of enterocyte effacement (LEE). The LEE encodes 41 predicted open reading frames (ORFs), including components of a type III secretion apparatus and secreted molecules involved in the disruption of the host cell cytoskeleton. To initiate our studies on regulation of genes within the LEE, we determined the genetic organization of the LEE, defining transcriptional units and mapping transcriptional start points. We found that components of the type III secretion system are transcribed from three polycistronic operons designated LEE1, LEE2 and LEE3. The secreted Esp molecules are part of a fourth polycistronic operon designated LEE4. Using reporter gene fusion assays, we found that the previously described plasmid-encoded regulator (Per) activated operons LEE1, LEE2 and LEE3, and modestly increased the expression of LEE4 in EPEC. Using single-copy lacZ fusions in K-12-derived strains, we determined that Per only directly activated the LEE1:lacZ fusion, and did not directly activate the other operons. Orf1 of the LEE1 operon activated the expression of single-copy LEE2:lacZ and LEE3:lacZ fusions in trans and modestly increased the expression of LEE4:lacZ in K-12 strains. Orf1 was therefore designated Ler, for LEE-encoded regulator. Thus, the four polycistronic operons of the LEE that encode type III secretion components and secreted molecules are now included in the Per regulon, where Ler participates in this novel regulatory cascade in EPEC.

Bacterial Adhesion↗

Homologs of the essential ubiquitin conjugating enzymes UBC1, 4, and 5 in yeast are encoded by a multigene family in Arabidopsis thaliana.

The covalent attachment of the 76 amino acid protein ubiquitin is an important prerequisite for the degradation of many eukaryotic proteins. The specificity of this ligation is accomplished in part by a family of distinct ubiquitin conjugating enzymes (E2s) working in concert with specific ubiquitin-protein ligases (E3s). Three essential E2s in yeast encoded by ScUBC1, -4, and -5 comprise a functionally overlapping E2 subfamily that appears responsible for degrading most abnormal and short-lived proteins. A 15 kDa E2 protein homologous to this family has been identified previously in wheat germ, designated TaE2(15kDa) (Girod and Vierstra (1993) J. Biol. Chem. 268, 955-960). This E2 is responsible for much of the ubiquitin conjugating activity observed in wheat germ extracts and works together with a unique E3 (designated E3 gamma) for substrate recognition. In this paper, the cloning of five genes encoding E2(15kDa) from Arabidiopsis thaliana is described (designated AtUBC8-12). They encode 149 amino acid basic proteins 94-98% similar to each other and 88-92% similar to ScUBC4 at the amino acid sequence level. In contrast, AtUBC8-12 are only 55-65% similar to the Arabidopsis E2s encoded by AtUBC1, -4, and -7. The AtUBC8-12 proteins do not contain N- or C-terminal extensions and have the active site at residue Cys-86, based on their homology with other E2s. Analyses of genomic Southern blots are consistent with the existence of multiple members encoding this E2 subfamily. AtUBC8-12 are transcribed to yield about 800 nucleotide mRNAs that, unlike ScUBC4 and -5, are not strongly induced by heat shock.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗

Molecular cloning of cDNAs encoding the protein backbones of arabinogalactan-proteins from the filtrate of suspension-cultured cells of Pyrus communis and Nicotiana alata.

This paper reports the isolation of cDNAs encoding the protein backbone of two arabinogalactan-proteins (AGPs), one from pear cell suspension cultures (AGPPc2) and the other from suspension cultures of Nicotiana alata (AGPNa2). The proteins encoded by these cDNAs are quite different from the 'classical' AGP backbones described previously for AGPs isolated from pear suspension cultures and extracts of N. alata styles. The cDNA for AGPPc2 encodes a 294 amino acid protein, of which a relatively short stretch (35 amino acids) is Hyp/Pro rich; this stretch is flanked by sequences which are dominated by Asn residues. Asn residues are not a feature of the 'classical' AGP backbones in which Hyp/Pro, Ser, Ala and Thr account for most of the amino acids. The cDNA for AGPNa2 encodes a 437 amino acid protein, which contains two distinct domains: one rich in Hyp/Pro, Ser, Ala, Thr and the other rich in Asn, Tyr and Ser. The composition and sequence of the Pro-rich domain resembles that of the 'classical' AGP backbone. The Asn-rich domains of the two cDNAs described have no sequence similarity; in both cases they are predicted to be processed to give a mature backbone with a composition similar to that of the 'classical' AGPs. The study shows that different AGPs can differ in the amino acid sequence in the protein backbone, as well as the composition and sequence of the arabinogalactan side-chains. It also shows that differential expression of genes encoding AGP protein backbones, as well as differential glycosylation, can contribute to the tissue specificity of AGPs.

Amino Acid Sequence↗

Characterization of the S locus genes, SLG and SRK, of the Brassica S3 haplotype: identification of a membrane-localized protein encoded by the S locus receptor kinase gene.

The S locus, which controls the self-incompatibility response in Brassica, has been shown to contain at least two genes. SLG encodes a secreted S locus glycoprotein whilst SRK encodes a putative S locus receptor kinase. SRK has been shown potentially to encode a functional kinase and genetic evidence indicates that this gene is essential for the self-incompatibility response. Here the characterization of the SRK and SLG genes of a Brassica line homozygous for the S3 haplotype is described. A 120 kDa glycoprotein was identified in stigmas and several lines of evidence indicated that this protein is encoded by the SRK3 gene. First, the 120 kDa glycoprotein was recognized by antibodies raised against peptides based on the SRK3 gene sequence. Secondly, this protein is polymorphic and, in an F2 population segregating for the S3 haplotype, was expressed only in plants possessing the S3 haplotype. Thirdly, the 120 kDa protein was expressed specifically in stigmas. Finally, the 120 kDa protein was only extracted from stigmas in the presence of detergent indicating that it is anchored in the membrane. SRK has been predicted to encode a transmembrane glycoprotein based on the deduced amino acid sequence. Located on the membrane, SRK is in a position to interface between an extracellular recognition event between pollen and pistil and an intracellular signal transduction pathway which initiates the self-incompatibility response.

Amino Acid Sequence↗

Plant disease resistance genes encode members of an ancient and diverse protein family within the nucleotide-binding superfamily.

The nucleotide binding site (NBS) is a characteristic domain of many plant resistance gene products. An increasing number of NBS-encoding sequences are being identified through gene cloning, PCR amplification with degenerate primers, and genome sequencing projects. The NBS domain was analyzed from 14 known plant resistance genes and more than 400 homologs, representing 26 genera of monocotyledonous, dicotyle-donous and one coniferous species. Two distinct groups of diverse sequences were identified, indicating divergence during evolution and an ancient origin for these sequences. One group was comprised of sequences encoding an N-terminal domain with Toll/Interleukin-1 receptor homology (TIR), including the known resistance genes, N, M, L6, RPP1 and RPP5. Surprisingly, this group was entirely absent from monocot species in searches of both random genomic sequences and large collections of ESTs. A second group contained monocot and dicot sequences, including the known resistance genes, RPS2, RPM1, I2, Mi, Dm3, Pi-B, Xa1, RPP8, RPS5 and Prf. Amino acid signatures in the conserved motifs comprising the NBS domain clearly distinguished these two groups. The Arabidopsis genome is estimated to contain approximately 200 genes that encode related NBS motifs; TIR sequences were more abundant and outnumber non-TIR sequences threefold. The Arabidopsis NBS sequences currently in the databases are located in approximately 21 genomic clusters and 14 isolated loci. NBS-encoding sequences may be more prevalent in rice. The wide distribution of these sequences in the plant kingdom and their prevalence in the Arabidopsis and rice genomes indicate that they are ancient, diverse and common in plants. Sequence inferences suggest that these genes encode a novel class of nucleotide-binding proteins.

Amino Acid Motifs↗

Methanobacterium thermoautotrophicum encodes two multisubunit membrane-bound [NiFe] hydrogenases. Transcription of the operons and sequence analysis of the deduced proteins.

Two gene groups, designated energy converting hydrogenase A (eha) and energy converting hydrogenase B (ehb), each encoding a putative multisubunit membrane-bound [NiFe] hydrogenase, were identified in the genome of Methanobacterium thermoautotrophicum. The length of the transcription units was determined using reverse transcription (RT)-PCR. The eha operon (12.5 kb) and the ehb operon (9.6 kb) were found to be composed of 20 and 17 open reading frames, respectively. Competitive RT-PCR was used to compare the amounts of eha and ehb transcripts with the amounts of transcripts of genes encoding the M. thermoautotrophicum catabolic enzymes cyclohydrolase (mch) and a subunit of heterodisulfide reductase (hdrC). In cells grown under conditions in which H2 was nonlimiting, the eha transcripts were 250-fold and 125-fold less abundant and the ehb transcripts were approximately sixfold and threefold less abundant than the hdrC and mch transcripts, respectively. In cells grown under H2 limitation, the amounts of eha and ehb transcripts were about threefold higher than in cells grown with sufficient H2 when compared to the amounts of hdrC and mch transcripts. Sequence analysis of the deduced proteins indicated that the eha and ehb operons each encode a [NiFe] hydrogenase large subunit, a [NiFe] hydrogenase small subunit, and two conserved integral membrane proteins. These proteins show high sequence similarity to subunits of the Ech hydrogenase from Methanosarcina barkeri, Escherichia coli hydrogenases 3 and 4, and CO-induced hydrogenase from Rhodospirillum rubrum, all of which form a distinct group of multisubunit membrane-bound [NiFe] hydrogenases and show high sequence similarity to the energy-conserving NADH:quinone oxidoreductase (complex I) from various organisms. In addition to these four subunits, the eha operon encodes a 6[4Fe-4S] polyferredoxin, a 10[4F-4S] polyferredoxin, four nonconserved hydrophilic subunits, and 10 nonconserved integral membrane proteins; the ehb operon encodes a 2[4Fe-4S] ferredoxin, a 14[4Fe-4S] polyferredoxin, two nonconserved hydrophilic subunits, and nine nonconserved integral membrane proteins. A function of these putative membrane-bound [NiFe] hydrogenases as proton pumps involved in endergonic reactions, such as the synthesis of formylmethanofuran from CO2, H2 and methanofuran, is discussed.

Amino Acid Sequence↗

Regional brain activity during shape recognition impaired by a scopolamine challenge to encoding.

In the present positron emission tomography (PET) study, we examine the effect of a scopolamine-induced challenge to encoding upon the pattern of regional cerebral blood flow during recognition of a list of abstract visual shapes 3 days after encoding of these shapes. This study was conducted to test hypotheses concerning the fusiform and thalamic contributions to object recognition arising from a previous imaging study of impaired recognition. In that study, we demonstrated that activity in the fusiform cortex and the thalamus during shape recognition was modulated by memory challenges. These memory challenges included, on one hand, impaired storage as a consequence of diazepam administration during encoding, and, on the other hand, impaired retrieval caused by a perceptual challenge. Activation in the fusiform cortex decreased during impaired recognition, irrespective of the type of challenge. In contrast, thalamic activation increased only when the recognition deficit resulted from impaired memory storage. Based on these results, we hypothesized that fusiform activation during recognition reflects the matching of an incoming stimulus with a stored one, whereas thalamic activation reflects retrieval attempts. These hypotheses would receive considerable support if scopolamine, which also impairs memory storage, induced similar modulations of fusiform and thalamic activation. In the present study, we observed that a scopolamine challenge to encoding does indeed modulate the activity in the very same regions that were previously modulated by a diazepam challenge. Hence, a similar memory deficit, although primarily effected through different neurochemical pathways, was paralleled by a similar modulation of activity in the same set of nodes in the shape recognition network. In the fusiform cortex, scopolamine decreased recognition-related activity, as did the sensory challenge of retrieval. Furthermore, covariate analysis demonstrated that the level of fusiform activity linearly correlates with behavioural performance. In the thalamus, activation increased following impaired encoding. This is in accordance with the idea that enhanced thalamic activity reflects increased effort expended in retrieval. In addition, in the intraparietal sulcus, differential activation also increased following impaired memory storage, possibly reflecting enhanced visuospatial attention in an effort to compensate for impaired performance.

Administration, Oral↗

A neural correlate of syntactic encoding during speech production.

Spoken language is one of the most compact and structured ways to convey information. The linguistic ability to structure individual words into larger sentence units permits speakers to express a nearly unlimited range of meanings. This ability is rooted in speakers' knowledge of syntax and in the corresponding process of syntactic encoding. Syntactic encoding is highly automatized, operates largely outside of conscious awareness, and overlaps closely in time with several other processes of language production. With the use of positron emission tomography we investigated the cortical activations during spoken language production that are related to the syntactic encoding process. In the paradigm of restrictive scene description, utterances varying in complexity of syntactic encoding were elicited. Results provided evidence that the left Rolandic operculum, caudally adjacent to Broca's area, is involved in both sentence-level and local (phrase-level) syntactic encoding during speaking.

Adult↗

A developmentally regulated mRNA from 3T3 adipocytes encodes a novel serine protease homologue.

We previously have isolated cDNA clones for several mRNAs that increase in abundance during the differentiation of 3T3 adipocytes but whose physiological role is unknown. We show here that a mRNA that is complementary to one of these clones and encodes a protein of 28 kDa is expressed abundantly in mouse fat pads but not in several other mouse tissues. Sequence analysis of the corresponding cDNA clone indicated that the encoded protein shows 30% overall amino acid homology to several serine proteases including trypsin, chymotrypsin, and elastase. Homology is much higher (64%) between the 28-kDa protein and regions that are strongly conserved among the members of the serine protease family. The derived protein also has key features characteristic of active serine proteases, including the histidine, aspartic acid, and serine residues, which comprise the charge relay system, and a potential cleavage site for activation of the zymogen. Primer extension analysis performed to obtain the sequence of the 5' end of mRNA that encodes the 28-kDa protein indicates that two forms of this mRNA exist and probably arise through alternative splicing. The two mRNAs encode signal sequences that differ by the deletion of one amino acid near the predicted cleavage site of the signal peptide. These results demonstrate that adipocyte differentiation is accompanied by the expression of mRNA encoding a serine protease homologue that can be synthesized with two different signal peptides.

Adipose Tissue↗

Pel, the protein that permits lambda DNA penetration of Escherichia coli, is encoded by a gene in ptsM and is required for mannose utilization by the phosphotransferase system.

Mannose uptake and phosphorylation in Escherichia coli is catalyzed by the phosphoenolpyruvate:glycose phosphotransferase system (PTS). The mannose-specific complex of the PTS, designated IIMan, comprises lipid and two membrane proteins, II-AMan and II-BMan. The proteins are encoded by ptsM, located at approximately equal to 40 minutes on the E. coli chromosome. A different genetic marker, pel, maps with ptsM, and is required for lambda DNA penetration of the cytoplasmic membrane. Earlier studies suggested that both pel function and II-BMan are encoded by the same gene, while a different gene (also in ptsM) encodes II-AMan. In the present studies, a ptsM clone, pCS13, was isolated from an E. coli HindIII gene bank in pBR322 and restored both mannose termentation and pel+ function to ptsM mutants defective in II-BMan. Subclones of pCS13 show that two distinct genes, manY and manZ, encode the pel+ function and the II-BMan protein, respectively; each gene may have its own promoter; whereas the protein encoded by manY (Pel) alone seems sufficient for lambda sensitivity, all three gene products are required for mannose fermentation, transport of the mannose analogue 2-deoxyglucose, and phosphorylation of the latter by cytoplasmic membranes. Thus, Pel is required for function of the IIMan complex. The efficiency of the complex may depend on the ratio of Pel to IIMan.

Bacterial Proteins↗

A single gene in mast cells encodes the core peptides of heparin and chondroitin sulfate proteoglycans.

The diversity of the genes encoding mammalian proteoglycan peptide cores was explored using a cDNA clone that encodes the partial sequence of a cell surface/pericellular matrix-localized chondroitin sulfate proteoglycan. Thus we were able to detect the expression of the gene(s) encoding the intracellular chondroitin sulfate proteoglycan produced by a variety of rat and mouse mucosal-like mast cells and the intracellular heparin proteoglycan synthesized by rat serosal mast cells. The cDNA from the proteoglycan cDNA clone pPG-1 was fractionated into two discrete fragments, one of which contained the nucleotides encoding the serine-glycine repeat sequence (pPG-B) and the other of which contained sequences on the 3' side of the repeat (pPG-M). As assessed by Southern blot analysis, pPG-B identified a large gene family, whereas pPG-M identified a single DNA fragment in the rat genome. When the pPG-1 insert and the two subcloned probes pPG-B and pPG-M were used to analyze RNA extracted from the rat and mouse mucosal-like mast cells and the rat serosal mast cells, the same major RNA species was detected at 1.3 kilobases with both probes. These data suggest that the gene responsible for the peptide core of the extracellular chondroitin sulfate proteoglycan synthesized by the rat yolk sac cell line is also the gene that encodes the core peptides of the secretory granule-localized chondroitin sulfate and heparin proteoglycans.

Aggrecans↗

Immunological identification of the genes encoding the four myosin heavy chain isoforms of Caenorhabditis elegans.

The nematode Caenorhabditis elegans produces four distinct myosin heavy chain (MHC) isoforms, A, B, C, and D. The MHC A and MHC B proteins are coordinately expressed in the body wall muscle and are incorporated into different regions of a single kind of thick filament. MHC C and MHC D are exclusively produced in the pharyngeal muscle. Previous studies of mutations that affect MHC B have shown that this isoform is encoded by the unc-54 gene. Three other MHC genes, myo-1, myo-2, and myo-3, were isolated from a C. elegans genomic library by hybridization with fragments of the unc-54 gene. We have now established the MHC isoform encoded by each gene. Restriction fragments from each of these genes were cloned in the plasmid expression vector pUR288, producing fusion proteins between Escherichia coli beta-galactosidase and portions of the MHC rod domains of each gene. The hybrid proteins were screened with a panel of 18 isoform-specific monoclonal antibodies. The results demonstrate that myo-1 encodes MHC D, myo-2 encodes MHC C, and myo-3 encodes MHC A.

Animals↗

Organization of the gene encoding common acute lymphoblastic leukemia antigen (neutral endopeptidase 24.11): multiple miniexons and separate 5' untranslated regions.

The common acute lymphoblastic leukemia antigen (CALLA) is a 749-amino acid type II integral membrane protein that has been identified recently as the neutral endopeptidase 24.11 [NEP (EC 3.4.24.11)]. Herein, we characterize the organization of the human CALLA/NEP gene and show that it spans more than 80 kilobases (kb) and is composed of 24 exons. Exons 1 and 2 encode 5' untranslated sequences; exon 3 [170 base pairs (bp)] encodes the initiation codon and transmembrane and cytoplasmic domain; 20 short exons (exons 4-23), ranging in size from 36 to 162 bp, encode most of the extracellular portion of the enzyme; and exon 24 (approximately 3400 bp) encodes the COOH-terminal 32 amino acids of the protein and contains the entire 3' untranslated region (UTR). Of note, the pentapeptide sequence (His-Glu-Ile-Thr-His) associated with metalloprotease zinc binding and substrate catalysis is encoded within a single exon (exon 19). Three types of CALLA/NEP cDNAs have been identified: these clones contain 5' UTR sequences differing from one another upstream of exon 3. These human 5' sequences are homologous to those found in rat brain and rabbit kidney NEP cDNAs. The three human CALLA cDNA types result from alternative splicing of exons 1, 2a, or 2b to the common exon 3. Moreover, exons 2a and 2b share the same 5' sequence but differ from each other by the use of two distinct donor splice sites 171 bp apart in the gene. The substantial conservation of 5' untranslated sequences among species and the existence of 5' alternative splicing suggest that CALLA gene expression may be differentially controlled in a tissue-specific and/or developmentally regulated fashion.

Amino Acid Sequence↗

Cloning of the cDNA encoding an RNA regulatory protein--the human iron-responsive element-binding protein.

Iron-responsive elements (IREs) are stemloop structures found in the mRNAs encoding ferritin and the transferrin receptor. These elements participate in the iron-induced regulation of the translation of ferritin and the stability of the transferrin receptor mRNA. Regulation in both instances is mediated by binding of a cytosolic protein to the IREs. High-affinity binding is seen when cells are starved of iron and results in repression of ferritin translation and inhibition of transferrin receptor mRNA degradation. The IRE-binding protein (IRE-BP) has been identified as an approximately 90-kDa protein that has been purified by both affinity and conventional chromatography. In this report we use RNA affinity chromatography and two-dimensional gel electrophoresis to isolate the IRE-BP for protein sequencing. A degenerate oligonucleotide probe derived from a single peptide sequence was used to isolate a cDNA clone that encodes a protein containing 13 other sequenced peptides obtained from the IRE-BP. Consistent with previous characterization of the IRE-BP, the cDNA encodes a protein of 87 kDa with a slightly acidic pI, and the corresponding mRNA of approximately 3.6 kilobases is found in a variety of cell types. The encoded protein contains a nucleotide-binding consensus sequence and regions of cysteine and histidine clusters. This mRNA is encoded by a single gene on human chromosome 9, a finding consistent with previous localization by functional mapping. The protein contains no previously defined consensus motifs for either RNA or DNA binding. The simultaneous cloning of a different, but highly homologous, cDNA suggests that the IRE-BP is a member of a distinct gene family.

Amino Acid Sequence↗