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PubMed1982

Development of apparent presynaptic elements formed in response to polylysine coated surfaces.

Formation of apparent presynaptic elements on polylysine-coated surfaces was examined with both scanning and transmission electron microscopy. Neurons from 2- or 8-day-old rat cerebellums were used in dissociated cell cultures as a source of growing neurites. An apparent presynaptic element is a defined swelling on a neurite with 40 nm diameter vesicles accumulated at the membrane which has a slight thickening. Apparent presynaptic elements do not have a normal postsynaptic element. In place of the postsynaptic element was the polylysine-coated surface of a large diameter Sepharose bead. The first apparent presynaptic elements were seen at 2 h of incubation, suggesting that morphologically identifiable synapses may form in this short a time. The number of apparent presynaptic elements on beads increased from 2 h to 24 h incubation and decreased from 5 to 9 days incubation. At all times non-neuronal cells grew up on to the beads and often covered both neurites and apparent presynaptic elements. In the longer incubations degenerating apparent presynaptic elements were seen engulfed by non-neuronal cells, suggesting that non-neuronal cells may have the ability to remove presynaptic elements that are not functioning. The number of synaptic vesicles per apparent presynaptic elements increased continuously between 2 h and 9 day incubation, eventually surpassing the number of synaptic vesicles seen in other presynaptic elements in cultures. This result suggests that an interaction between presynaptic and postsynaptic elements may be necessary to limit the number of synaptic vesicles found in presynaptic elements. Cultures grown for only 5 days in vitro (DIV) and incubated for 1 day with coated beads had the most apparent presynaptic elements, while those at 28 DIV and incubated for 1 day had the least. In the cultures at 5 DIV, neurons formed many apparent presynaptic elements, but these neurons could form only a very few normal synapses between themselves. Thus the beginning of the formation of synaptic contacts may depend on the availability of future postsynaptic elements and not on the lack of future presynaptic elements.

Animals↗

PubMed1989

Characterization of bz1 mutants isolated from mutator stocks with high and low numbers of Mu1 elements.

The high frequency of mutations in Mutator stocks of maize is the result of transposition of Mu elements. Nine different Mu elements that share the 220 bp Mu terminal inverted repeats have been described. Mu1 elements have been found inserted into most of the molecularly characterized mutant alleles isolated from Mutator stocks, and most Mutator stocks contain a high number of Mu1 elements (10-60). However, it is clear that additional Mu elements, which share the Mu1 termini but have unrelated internal sequences, can also transpose in Mutator stocks. We were interested in comparing the mutation frequency and type of elements that inserted into a particular locus when Mutator stocks with differing numbers of Mu1 elements were utilized. Furthermore, previous studies with Mu-induced mutations have demonstrated that the element that inserted most frequently was Mu1. Therefore, to try to obtain Mu elements different from Mu1 we utilized a stock that had a low number (3-6) of Mu1 elements as well as a Mutator stock with a more typical number of Mu1 elements (20-60). Utilizing both stocks, we isolated numerous mutants at one gene, Bronze 1 (Bz1), and compared the type of elements inserted. In this paper we report that both the high and low Mu1 stocks produced bz1 mutants at frequencies characteristic of Mutator stocks, 6.6 and 4.3 x 10(-5), respectively. We describe the isolation of 20 bz1 mutations, and the initial molecular characterization of eight unstable mutations: two from the high Mu1 stock and six from the low Mu1 stock. The six alleles isolated from the low Mu1 stock appear to contain deleted Mu1 elements, and the two alleles isolated from the high Mu1 stock contain elements very similar to Mu1. When the mutants from the low Mu1 stocks were examined, it was found that the Mu1-related elements increased from 3-6 copies to 9-20 copies in one generation. The high number of Mu1-related elements was maintained in subsequent outcrosses. This spontaneous activation and amplification of Mu1-related elements occurred in at least 1% of the low Mu1 plants.

Alleles↗

PubMed2026

In vitro fertilization-conceived offspring exhibit altered Long Interspersed Nuclear Elements-1 retrotransposition dynamics associated with long-term disease risks.

BACKGROUND: In vitro fertilization has transformed reproductive medicine, yet offspring conceived through in vitro fertilization display elevated risks for diverse long-term health conditions, with underlying mechanisms unclear. Long Interspersed Nuclear Elements-1, a mobile genetic element responsive to environmental stress, represents a potential mediator. OBJECTIVE: This study aimed to test the hypothesis that in vitro fertilization procedures may act as an embryonic stressor that alters Long Interspersed Nuclear Elements-1 dynamics, potentially contributing to genomic instability associated with long-term disease susceptibility. STUDY DESIGN: Umbilical cord blood or peripheral blood from 33 in vitro fertilization and 42 naturally conceived neonates were collected for whole-genome sequencing. Total Long Interspersed Nuclear Elements-1 proportion in individual genome was counted with Bowtie2 software. De novo Long Interspersed Nuclear Elements-1 insertion and Long Interspersed Nuclear Elements-1 deletion were detected with Mobile Element Locator Tool. Three parent-matched in vitro fertilization-naturally conceived sibling pairs were included to control for genetic background. Disease association analysis was performed for genes within 500 kb of differential Long Interspersed Nuclear Elements-1 sites in The Database for Annotation, Visualization and Integrated Discovery (DAVID). Statistical analysis was performed using the R language. RESULTS: In vitro fertilization offspring demonstrate elevated global Long Interspersed Nuclear Elements-1 content compared to naturally conceived controls (P=.04). This finding was corroborated in 3 sibling pairs from identical genetic backgrounds, where in vitro fertilization-conceived children consistently exhibited higher Long Interspersed Nuclear Elements-1 levels than their naturally conceived siblings. Eleven genomic loci with differential Long Interspersed Nuclear Elements-1 insertion frequencies and 14 loci with differential Long Interspersed Nuclear Elements-1 deletion frequencies between in vitro fertilization offspring and naturally conceived controls were identified. Notably, these differential Long Interspersed Nuclear Elements-1 sites demonstrated significant enrichment near genes implicated in metabolic, cardiovascular, neuropsychiatric, and neoplastic diseases, conditions associated with in vitro fertilization conception. CONCLUSION: These findings provide preliminary evidence that in vitro fertilization conception is associated with increased Long Interspersed Nuclear Elements-1 content and altered genomic distribution of Long Interspersed Nuclear Elements-1 elements. The proximity of these differential Long Interspersed Nuclear Elements-1 sites to disease-associated genes suggests a plausible genomic mechanism linking in vitro fertilization-associated embryonic stress to elevated disease risk. This work provides valuable molecular insights that may inform the ongoing discussion about assisted reproductive technology safety and suggests that continued attention to genomic integrity in in vitro fertilization-conceived individuals would be beneficial.

Humans↗

PubMed1998

Excision of the tol2 transposable element of the medaka fish, Oryzias latipes, in zebrafish, Danio rerio.

The Tol2 element is a transposable element in Oryzias latipes (the medaka fish) found in the tyrosinase gene locus of the tyrosinase-deficient mutant medaka fish and has been shown to be excised from the genome during medaka embryogenesis (Koga, A., Suzuki, M., Inagaki, H., Bessho, Y., Hori, H., 1996. Transposon element in fish. Nature 383, 30). It is, however, not known whether the Tol2 element is an autonomous element. To determine whether the cloned Tol2 element is an autonomous element and whether excision can occur also in the other fish species, the plasmid DNA harboring the Tol2 element was injected to fertilized eggs of zebrafish, Danio rerio, and the total DNA extracted from the embryos 9-10h after the injection was analyzed by PCR. When a plasmid with the full-length Tol2 element was used for the microinjection, in 39 out of 43 injected embryos, we found generation of short PCR products indicative of the loss of the Tol2 element from the injected plasmid. Ten of these cases were analyzed at the DNA sequence level, and nine of them showed either precise excision of the Tol2 element (three cases) or nearly precise excision of the element with the addition of a few nucleotides of the target duplication (six cases). When a deletion version of the Tol2 element that retained the terminal inverted repeats but lacked about one-fourth of the open reading frame-coding region was used for the microinjection, such short PCR products could not be amplified from any of the injected embryos (0 out of 30). Thus, the Tol2 element is capable of excision in zebrafish embryos, presumably dependent on a putative transposase encoded by the Tol2 element itself. This transient embryonic excision assay using zebrafish should be useful to analyze the structure and the function of the transposase and cis-elements necessary for excision. Also, this study implies the potential use of the Tol2 element in transgenesis and insertional mutagenesis in both zebrafish and the medaka fish.

Animals↗

PubMed1992

A temperature-regulated, retrotransposon-like element from Candida albicans.

A repetitive element was isolated from the genome of Candida albicans. This repetitive element, which we designated alpha, was localized to a 500-bp fragment of genomic DNA. The alpha element was dispersed in the genome and varied in copy number and genomic location in the strains examined. Analyses of various loci containing the alpha element identified a locus containing a composite element. This composite element consisted of two direct repeats of the alpha element separated by approximately 5.5 kb of DNA, a structural arrangement similar to that of retrovirus-like transposable elements. The flanking alpha elements of the composite structure were 388 bp in length and were identical in sequence. They were bounded by the nucleotides 5'-TG. ... CA-3', which were part of a delimiting inverted repeat, a feature conserved in the long terminal repeats of retroviruses and retrovirus-like elements. As in retrovirus-like elements, the entire composite element, including the alpha elements, was transcribed into an approximately unit-length mRNA. The expression of this transcript was greatly increased when cells were grown at 25 versus 37 degrees C. As has been found in many retrotransposons, the composite element was flanked by a 5-bp duplication and varied in both copy number and genomic location in various strains. We conclude that the composite element is a retrotransposon-like element, and we have designated this element Tca1. We suggest that Tca1 may be relevant to the genomic evolution of C. albicans and the pathogenic potential of the organism.

Base Sequence↗

PubMedDate not supplied

Geometric modeling of the human torso using cubic hermite elements.

We discuss the advantages and problems associated with fitting geometric data of the human torso obtained from magnetic resonance imaging, with high-order (bicubic Hermite) surface elements. These elements preserve derivative (C1) continuity across element boundaries and permit smooth anatomically accurate surfaces to be obtained with relatively few elements. These elements are fitted to the data with a new nonlinear fitting procedure that minimizes the error in the fit while maintaining C1 continuity with nonlinear constraints. Nonlinear Sobelov smoothing is also incorporated into this fitting scheme. The structures fitted along with their corresponding root mean-squared error, number of elements used, and number of degrees of freedom (df) per variable are: epicardium (0.91 mm, 40 elements, 142 df), left lung (1.66 mm, 80 elements, 309 df), right lung (1.69 mm, 80 elements. 309 df), skeletal muscle surface (1.67 mm, 264 elements, 1,010 df), fat layer (1.79 mm, 264 elements, 1.010 df), and the skin layer (1.43 mm, 264 elements, 1,010 df). The fitted surfaces are assembled into a combined finite element/boundary element model of the torso in which the exterior surfaces of the heart and lungs are modeled with two-dimensional boundary elements and the layers of the skeletal muscle, fat, and skin are modeled with finite elements. The skeletal muscle and fat layers are modeled with bicubic Hermite linear elements and are obtained by joining the adjacent surface elements for each layer. Applications for the torso model include the forward and inverse problems of electrocardiography, defibrillation studies, radiation dosage studies, and heat transfer studies.

Abdomen↗

PubMed1990

TATA-dependent and TATA-independent function of the basal and heat shock elements of a human hsp70 promoter.

We have characterized the interactions between the TATA element and other sequence elements of a human heat shock protein 70 (hsp70) promoter by a mutational approach. Expression of a distal element of this promoter requires an intact TATA element in human cell lines. The hsp70 TATA element can be functionally replaced for this interaction by TATA elements from the simian virus 40 early and adenovirus EIIa promoters. The TATA element in this promoter therefore both determines the appropriate start site and determines strength by allowing function of the distal element. In contrast, three proximal upstream elements necessary for basal and heat-regulated transcription have no requirement either for a TATA element or for any other proximal element. The behavior of promoters multiply mutant in these proximal elements implies that these elements function independently. We examined the interaction between the heat shock element (HSE) and the TATA element as the distance between the two factor-binding sites was increased. It was necessary to create a mutant HSE with an extended consensus sequence in order for the HSE to function at a distance. Moving this extended HSE 500 bases upstream did not increase its dependence on the TATA element, suggesting that the TATA independence of this element is intrinsic to its function and is not determined by distance from the promoter.

Animals↗

PubMed1999

Site-specific integration of the conjugal Vibrio cholerae SXT element into prfC.

Vibrio cholerae O139, the first non-O1 serogroup of V. cholerae to give rise to epidemic cholera, is characteristically resistant to the antibiotics sulphamethoxazole, trimethoprim, chloramphenicol and streptomycin. Resistances to these antibiotics are encoded by a 62 kb self-transmissible, conjugative, chromosomally integrating element designated the 'SXT element'. We found that the SXT element integrates site specifically into both V. cholerae and Escherichia coli K-12 into the 5' end of prfC, the gene encoding peptide chain release factor 3. Integration of the SXT element interrupts the chromosomal prfC gene, but the element encodes a new 5' end of prfC that restores the reading frame of this gene. The recombinant of prfC allele created upon element integration is functional. The integration and excision mechanism of the SXT element shares many features with site-specific recombination found in lambdoid phages. First, like lambda, the SXT element forms a circular extrachromosomal intermediate through specific recombination of the left and right ends of the integrated element. Second, chromosomal integration of the element occurs via site-specific recombination in a 17 bp sequence found in the circular form of the SXT element and a similar 17 bp sequence in prfC. Third, both chromosomal integration and excision of the SXT element were found to require an element-encoded int gene with strong similarities to the lambda integrase family. Based on the properties of the SXT element, we propose to classify this element as a CONSTIN, an acronym for a conjugative, self-transmissible, integrating element.

Amino Acid Sequence↗

PubMed1994

Heterogeneous functional Ty1 elements are abundant in the Saccharomyces cerevisiae genome.

Despite the abundance of Ty1 RNA in Saccharomyces cerevisiae, Ty1 retrotransposition is a rare event. To determine whether transpositional dormancy is the result of defective Ty1 elements, functional and defective alleles of the retrotransposon in the yeast genome were quantitated. Genomic Ty1 elements were isolated by gap repair-mediated recombination of pGTy1-H3(delta 475-3944) HIS3, a multicopy plasmid containing a GAL1/Ty1-H3 fusion element lacking most of the gag domain (TYA) and the protease (PR) and integrase (IN) domains. Of 39 independent gap repaired pGTyHIS3 elements isolated, 29 (74%) transposed at high levels following galactose induction. The presence of restriction site polymorphisms within the gap repaired region of the 29 functional pGTyHIS3 elements indicated that they were derived from at least eight different genomic Ty1 elements and one Ty2 element. Of the 10 defective pGTyHIS3 elements, one was a partial gap repair event while the other nine were derived from at least six different genomic Ty1 elements. These results suggest that most genomic Ty1 elements encode functional TYA, PR and IN proteins. To understand how functional Ty1 elements are regulated, we tested the hypothesis that a TYB protein associates preferentially in cis with the RNA template that encodes it, thereby promoting transposition of its own element. A genomic Ty1 mhis3AI element containing either an in-frame insertion in PR or a deletion in TYB transposed at the same rate as a wild-type Ty1mhis3AI allele, indicating that TYB proteins act efficiently in trans. This result suggests in principle that defective genomic Ty1 elements could encode trans-acting repressors of transposition; however, expression of only one of the nine defective pGTy1 isolates had a negative effect on genomic Ty1 mhis3AI element transposition in trans, and this effect was modest. Therefore, the few defective Ty1 elements in the genome are not responsible for transpositional dormancy.

Amino Acid Sequence↗

PubMed2005

Saccadic search performance: the effect of element spacing.

In a saccadic search task, we investigated whether spacing between elements affects search performance. Since it has been suggested in the literature that element spacing can affect the eye movement strategy in several ways, its effects on search time per element are hard to predict. In the first experiment, we varied the element spacing (3.4 degrees -7.1 degrees distance between elements) and target-distracter similarity. As expected, search time per element increased with target-distracter similarity. Decreasing element spacing decreased the search time per element. However, this effect was surprisingly small in comparison to the effect of varying target-distracter similarity. In a second experiment, we elaborated on this finding and decreased element spacing even further (between 0.8 degrees and 3.2 degrees). Here, we did not find an effect on search time per element for element spacings from 3.2 degrees to spacings as small as 1.5 degrees . It was only at distances smaller than 1.5 degrees that search time per element increased with decreasing element spacing. In order to explain the remarkable finding that search time per element was not affected for such a wide range of element spacings, we propose that irrespective of the spacing crowding kept the number of elements processed per fixation more or less constant.

Adult↗

PubMed2006

Introduction of the factor of partitioning in the lithogenic enrichment factors of trace element bioaccumulation in plant tissues.

Bioindicators are widely used in the study of trace elements inputs into the environment and great efforts have been conducted to separate atmospheric from soil borne inputs on biomass accumulation. Many monitoring studies of trace element pollution take into account the dust particles located in the plant surface plus the contents of the plant tissues. However, it is usually only the trace element content in the plant tissues that is relevant on plant health. Enrichment factor equations take into account the trace element enrichment of biomasses with respect soil or bedrocks by comparing the ratios of the trace element in question to a lithogenic element, usually Al. However, the enrichment equations currently in use are inadequate because they do not take into account the fact that Al (or whichever reference element) and the element in question may have different solubility-absorption-retention levels depending on the rock and soil types involved. This constrain will become critical when results from different sites are compared and so in this article we propose that the solubility factors of each element are taken into account in order to overcome this constrain. We analysed Sb, Co, Ni, Cr, Pb, Cd, Mn, V, Zn, Cu, As, Hg, and Al concentration in different zones of Catalonia (NE Spain) using the evergreen oak Quercus ilex and the moss Hypnum cupressiforme as target species. We compared the results obtained in rural and non industrial areas with those from the Barcelona Metropolitan Area. We observed differences in Al concentrations of soils and bedrocks at each different site, together with the differences in solubility between Al and the element in question, and a weak correlation between total soil content and water extract content through different sites for most trace elements. All these findings show the unsuitability of the current enrichment factors for calculating lithospheric and atmospheric contributions to trace element concentrations in biomass tissues. The trace element enrichment factors were calculated by subtracting the part predicted by substrate composition (deduced from water extracts from soils and bedrock) from total concentrations. Results showed that for most of the trace elements analysed, trace elements enrichment factors were higher inside the Barcelona Metropolitan Area than outside, a finding that indicates that greater atmospheric inputs occur in urban areas. The results show that the most useful and correct way of establishing a reference for lithospheric and atmospheric inputs into the plant tissues is, first, to analyse samples of the same plant species collected from a number of sites possessing similar environmental conditions (climate, vegetation type, soil type) and, second, to use this new enrichment factor obtained by subtracting from the total concentration in plant tissue the predicted contribution of soil or bedrock extracts instead of that of total soil or bedrock concentrations.

Biomass↗

PubMed1985

Assessment of trace element status.

Biochemical and clinical investigations involving trace elements are made for the diagnosis of inherited or acquired deficiencies of essential trace elements and their treatment, to monitor the efficacy of the therapeutic administration of non-essential trace elements in order to achieve maximum clinical response with minimum toxicity, and for the early detection of excessive ingestion of non-essential toxic trace elements. The wide range of tests used to assess trace element status in these three areas of clinical importance is discussed with examples of essential and of toxic trace elements since therapeutic use of trace elements is discussed elsewhere in this issue. Particular attention is given to zinc, copper, selenium, lead and cadmium because the various tests used to assess the status of these elements encompass the principles of all currently available tests. Although trace element analysis of body fluids and tissues is the most useful and most commonly used method of assessment of trace element status, this is of limited value and no single test may be considered as ideal for any element. The provision of more detailed information from elemental analysis of cellular and subcellular fractions and of protein fractions from plasma leads inexorably to measurements of element-dependent enzymes, metalloproteins and of low molecular weight element-binding ligands. Even at this level of discrimination the choice of body tissue or tissue fluid for investigation is determined by the trace element and its principal metabolic targets.

Cadmium↗

PubMed2002

Mobile elements in archaeal genomes.

The recent availability of several archaeal genome sequences has provided a basis for detailed analyses of the frequency, location and phylogeny of archaeal mobile elements. All the known elements fall into two main types, autonomous insertion sequence (IS) elements and the non-autonomous miniature inverted repeat element (MITE)-like elements. Both classes are considered to be mobilized via transposases that are encoded by the IS elements, although mobility has only been demonstrated experimentally for a few elements. The number, and diversity, of the elements differs greatly between the genomes. At one extreme Sulfolobus solfataricus P2 and Halobacterium NRC-1 are very rich in elements while Methanobacterium thermoautotrophicum contains none. The former also show examples of complex clusters of interwoven elements. An analysis of the genomic distribution in S. solfataricus suggests that the putative oriC and terC regions act as barriers for the mobility of both IS and MITE-like elements. Moreover, the very high level of truncated IS elements in the genomes of S. solfataricus, Sulfolobus tokodaii and Thermoplasma volcanium suggests that there may be a cellular mechanism for selectively inactivating IS elements at a point when they become too numerous and disadvantageous for the cell. Phylogenetically, archaeal IS elements are confined to 11 of the 17 known families of bacterial and eukaryal IS elements where some generate distinct subgroups. Finally, DNA viruses, plasmids and DNA fragments can also be inserted into, and excised from, archaeal genomes by means of an integrase-mediated mechanism that has special archaeal characteristics.

Crenarchaeota↗

PubMedDate not supplied

Trace element contamination of total parenteral nutrition. 1. Contribution of component solutions.

BACKGROUND: Trace elements have been shown to contaminate total parenteral nutrition (TPN) solutions. METHODS: This study used the multi-elemental technology of inductively coupled plasma-mass spectrometry to demonstrate the extent to which trace elements were present in amounts above (ie, as contaminants) or below expected levels in eight TPN component solutions. RESULTS: Of the 66 trace elements scanned, there were 12 trace element contaminants in amounts >1 microg/L (zinc, copper, manganese, chromium, selenium, boron, aluminum, titanium, barium, vanadium, arsenic, and strontium) in the eight component solutions studied. Trace element contaminants were present in all solutions, and different trace elements contaminated the solutions at various concentrations. Component solutions of amino acid, potassium chloride, calcium gluconate, and sodium chloride contained the greatest numbers of trace element contaminants, whereas the lowest numbers were present in sterile water and magnesium sulfate. Interlot and intermanufacturer variations were apparent. Measured concentrations of trace elements in the multi-trace element additive solution also were higher than the labeled values. A comparison of the amounts of contaminated trace elements delivered by a typical TPN mixture relative to the amounts typically absorbed by the gastrointestinal tract indicates that the inadvertent delivery of trace elements from contaminated TPN solutions may be substantial. CONCLUSIONS: All eight components tested were contaminated with trace elements not intended to be present in the product, and similarly, the multi-trace element component contained trace elements either above or below that which the label claimed.

Drug Contamination↗

PubMed1998

Characterization of the initiator and downstream promoter elements of herpes simplex virus 1 late genes.

Previously identified cis-acting regulatory elements of herpes simplex virus (HSV) 1 late promoters include a TATA element upstream from the start of transcription, an initiator-like element at the start of transcription, and sequences downstream from the start of transcription. To determine whether these elements are functionally equivalent to similar elements from other eukaryotic genes, model late promoters were constructed using well-characterized regulatory elements from non-HSV genes. These modular promoters were then inserted into the viral genome upstream from a lacZ marker gene. Results showed that a eukaryotic initiator element, along with a TATA element, can function as a late HSV promoter. Several initiator sequences from both viral and nonviral genes were functionally similar to the initiator-like element in HSV-1 late promoters; however, a random sequence of the same size and a similarly located sequence from the HSV-1 early thymidine kinase promoter could not substitute for the initiator element. These results indicate that eukaryotic initiator elements are functionally equivalent to HSV-1 late promoter initiator elements. In addition, the downstream element of the late glycoprotein C promoter was further analyzed by construction of a series of small deletions and insertions. The presence of the downstream glycoprotein C region in a promoter consisting of a strong TATA and initiator element increased mRNA expression by a modest amount; this effect appeared to be sequence specific and dependent on its exact alignment with the upstream elements of the promoter.

Animals↗

PubMed2002

P element-mediated duplications of genomic regions in Drosophila melanogaster.

Previously we have described highly unstable yellow mutations induced by chimeric elements that consist of genomic sequences originating from different regions of the X chromosome flanked by identical copies of an internally deleted 1.2 kb P element. To study further the origin and the mechanism of formation of chimeric mobile elements, we analyzed complex y-sc mutations, induced by inversions between P elements located in the neighboring yellow and scute loci. The breakpoints of the inversions are flanked by two P elements in head-to-head orientation on one side and by one P element on the other side. Such an arrangement of P elements leads to frequent duplication into the site between the two P element copies located in head-to-head orientation of the yellow sequences adjacent to the single P element. The duplicated yellow sequences either partly replace the sequence of one of the P elements or are inserted between the conserved head-to-head oriented P elements. In some cases two copies of the yellow sequence are duplicated between the P elements in inverted tail-to-tail orientation. The structure of the P elements at the place of duplication and of the P element- yellow junction suggests that the described duplications, which form chimeric mobile elements, are generated through the previously proposed synthesis-dependent strand annealing mechanism.

Animals↗

PubMed1996

Human proteinase-3 expression is regulated by PU.1 in conjunction with a cytidine-rich element.

Human proteinase-3 is one of three serine proteinases present in the azurophil granules of polymorphonuclear leukocytes along with elastase and cathepsin G. Proteinase-3 gene expression is confined to the promyelocytic stage of polymorphonuclear leukocyte maturation. The present investigation identifies elements responsible for this highly controlled tissue- and developmental-specific expression of proteinase-3. Within the first 200 base pairs of the proteinase-3 promoter, two elements were identified as important for expression, these elements at -101 and -190 confer the majority of the activity. The element at -101 has a PU.1 consensus. It binds a myeloid nuclear protein of approximately 45 kDa that "supershifts" with PU.1 antibody and is competed by the CD11b PU.1 element. The element at -190 has a core sequence of CCCCGCCC (CG element). The cytidines but not the guanidine are essential for promoter activity. The CG element binds a second nuclear protein with a molecular mass of approximately 40 kDa that is found in cells of myeloid lineage as well as non-myeloid HeLa cells. However, the proteinase-3 promoter is not active in HeLa cells which suggests that the CG element alone is not sufficient for proteinase-3 gene expression. Maturation of promyelocytic cells results in an inhibition of proteinase-3 gene expression and a reduction in nuclear protein binding to the PU.1 and CG elements. Similar elements occur in the elastase and cathepsin G promoters. Using the elastase and cathepsin G PU.1 and CG-like elements as probes results in identical band-shift patterns to that obtained with proteinase-3 PU.1 and CG elements. These data suggest that there is cooperative interaction between a PU.1 and a CG element with a consensus of CCCCXCCC and that they are important control elements for tissue- and developmental-specific expression of azurophil serine proteinases of polymorphonuclear leukocytes.

Autoantigens↗

PubMed1992

Definition of the upstream efficiency element of the simian virus 40 late polyadenylation signal by using in vitro analyses.

The polyadenylation signal for the late mRNAs of simian virus 40 is known to have sequence elements located both upstream and downstream of the AAUAAA which affect efficiency of utilization of the signal. The upstream efficiency element has been previously characterized by using deletion mutations and transfection analyses. Those studies suggested that the upstream element lies between 13 and 48 nucleotides upstream of the AAUAAA. We have utilized in vitro cleavage and polyadenylation reactions to further define the upstream element. 32P-labeled substrate RNAs were prepared by in vitro transcription from wild-type templates as well as from mutant templates having deletions and linker substitutions in the upstream region. Analysis of these substrates defined the upstream region as sequences between 13 and 51 nucleotides upstream of the AAUAAA, in good agreement with the in vivo results. Within this region, three core elements with the consensus sequence AUUUGURA were identified and were specifically mutated by linker substitution. These core elements were found to contain the active components of the upstream efficiency element. Using substrates with both single and double linker substitution mutations of core elements, we observed that the core elements function in a distance-dependent manner. In mutants containing only one core element, the effect on efficiency increases as the distance between the element and the AAUAAA decreases. In addition, when core elements are present in multiple copies, the effect is additive. The core element consensus sequence, which bears homology to the Sm protein complex-binding site in human U1 RNA, is also found within the upstream elements of the ground squirrel hepatitis B and cauliflower mosaic virus polyadenylation signals (R. Russnak, Nucleic Acids Res. 19:6449-6456, 1991; H. Sanfacon, P. Brodmann, and T. Hohn, Genes Dev. 5:141-149, 1991), suggesting functional conservation of this element between mammals and plants.

Base Sequence↗