In vivo dissection of cis-acting determinants for plastid RNA editing.

1996 ◽  
Vol 15 (18) ◽  
pp. 5052-5059 ◽  
Author(s):  
R. Bock ◽  
M. Hermann ◽  
H. Kössel
Keyword(s):  
Rna Editing ◽  
Cis Acting

scholarly journals Recognition of RNA Editing Sites Is Directed by Unique Proteins in Chloroplasts: Biochemical Identification of cis-Acting Elements and trans-Acting Factors Involved in RNA Editing in Tobacco and Pea Chloroplasts

2002 ◽  
Vol 22 (19) ◽  
pp. 6726-6734 ◽  
Author(s):  
Tetsuya Miyamoto ◽  
Junichi Obokata ◽  
Masahiro Sugiura
Keyword(s):  
Rna Editing ◽  
Editing Site ◽  
Mrna Editing ◽  
Higher Plant ◽  
In Vitro System ◽  
Cis Acting ◽  
Biochemical Identification ◽  
Editing Activity

ABSTRACT RNA editing in higher-plant chloroplasts involves C-to-U conversions at specific sites. Although in vivo analyses have been performed, little is known about the biochemical aspects of chloroplast editing reactions. Here we improved our original in vitro system and devised a procedure for preparing active chloroplast extracts not only from tobacco plants but also from pea plants. Using our tobacco in vitro system, cis-acting elements were defined for psbE and petB mRNAs. Distinct proteins were found to bind specifically to each cis-element, a 56-kDa protein to the psbE site and a 70-kDa species to the petB site. Pea chloroplasts lack the corresponding editing site in psbE since T is already present in the DNA. Parallel in vitro analyses with tobacco and pea extracts revealed that the pea plant has no editing activity for psbE mRNAs and lacks the 56-kDa protein, whereas petB mRNAs are edited and the 70-kDa protein is also present. Therefore, coevolution of an editing site and its cognate trans-factor was demonstrated biochemically in psbE mRNA editing between tobacco and pea plants.

scholarly journals APOBEC1 mediated C-to-U RNA editing: target sequence and trans-acting factor contribution to 177 RNA editing events in 119 murine transcripts in-vivo

2021 ◽  
Author(s):  
Saeed Soleymanjahi ◽  
Valerie Blanc ◽  
Nicholas Davidson
Keyword(s):  
Rna Editing ◽  
Nearest Neighbor ◽  
Cytidine Deaminase ◽  
Target Sequence ◽  
Flanking Sequence ◽  
Cis Acting ◽  
Rna Targets ◽  
Genome Wide ◽  
Nucleotide Modification

Mammalian C-to-U RNA editing was described more than 30 years ago as a single nucleotide modification in APOB RNA in small intestine, later shown to be mediated by the RNA-specific cytidine deaminase APOBEC1. Reports of other examples of C-to-U RNA editing, coupled with the advent of genome-wide transcriptome sequencing, identified an expanded range of APOBEC1 targets. Here we analyze the cis-acting regulatory components of verified murine C-to-U RNA editing targets, including nearest neighbor as well as flanking sequence requirements and folding predictions. We summarize findings demonstrating the relative importance of trans-acting factors (A1CF, RBM47) acting in concert with APOBEC1. Using this information, we developed a multivariable linear regression model to predict APOBEC1 dependent C-to-U RNA editing efficiency, incorporating factors independently associated with editing frequencies based on 103 Sanger-confirmed editing sites, which accounted for 84% of the observed variance. Co-factor dominance was associated with editing frequency, with RNAs targeted by both RBM47 and A1CF observed to be edited at a lower frequency than RBM47 dominant targets. The model also predicted a composite score for available human C-to-U RNA targets, which again correlated with editing frequency.

Transgenic models for study of pulmonary development and disease

1994 ◽  
Vol 267 (5) ◽  
pp. L489-L497 ◽  
Author(s):  
S. W. Glasser ◽  
T. R. Korfhagen ◽  
S. E. Wert ◽  
J. A. Whitsett
Keyword(s):  
Epithelial Cell ◽  
Animal Models ◽  
Gene Targeting ◽  
Transgenic Mouse ◽  
Embryonic Stem ◽  
Lung Epithelial Cell ◽  
Major Advance ◽  
Cis Acting ◽  
Lung Epithelial

This review summarizes progress in the application of transgenic mouse technology to the study of lung development and disease. Since advances in molecular genetics have greatly facilitated the isolation of cDNA and genes, our ability to readily assess roles of both normal and mutated genes in transgenic mouse in vivo represents a major advance, bridging molecular biology and whole animal physiology. Strategies have been developed in which lung epithelial cell promoter elements are used to drive normal or mutated genes into specific subsets of respiratory epithelial cells in the lungs of developing and mature transgenic mice. These mice have been used to elucidate the cis-acting elements controlling lung epithelial cell gene expression, to discern the role of specific polypeptides in lung morphogenesis and tumorigenesis, and to create animal models of pulmonary disease. The ability to mutate genes at their precise chromosomal locations through gene targeting in embryonic stem cells has lead to the production of animal models of lung diseases such as cystic fibrosis. Both gene insertion and gene targeting create permanent mouse lines that pass the modified gene to their progeny, providing animals for the study of the pathogenesis and treatment of pulmonary disorders.

Functional characterization of the human hypoxanthine phosphoribosyltransferase gene promoter: evidence for a negative regulatory element

1991 ◽  
Vol 11 (8) ◽  
pp. 4157-4164
Author(s):  
D E Rincón-Limas ◽  
D A Krueger ◽  
P I Patel
Keyword(s):  
Regulatory Element ◽  
Genomic Structure ◽  
Functional Characterization ◽  
Translation Initiation Site ◽  
Hypoxanthine Phosphoribosyltransferase ◽  
Hprt Gene ◽  
Negative Element ◽  
Cis Acting ◽  
Human Hprt Gene

The enzyme hypoxanthine phosphoribosyltransferase (HPRT) catalyzes the metabolic salvage of the purine bases hypoxanthine and guanine. We previously characterized the genomic structure of the human HPRT gene and described its promoter sequence. In this report, we identify cis-acting transcriptional control regions of the human HPRT gene by linking various 5'-flanking sequences to the bacterial chloramphenicol acetyltransferase gene. The sequence from positions -219 to -122 relative to the translation initiation site is required for maximal expression of this gene, and it functions equally in both normal and reverse orientations. In addition, a cis-acting negative element is present in the region spanning from positions -570 to -388. This negative element can also repress promoters of heterologous genes, such as those of adenosine deaminase and dihydrofolate reductase, which are structurally and functionally similar to the human HPRT promoter. Furthermore, this repressor element functions independently of its orientation but appears to be distance dependent. In vivo competition assays demonstrated that the trans-acting factor(s) that binds to this negative element specifically inhibits human HPRT promoter activity. Taken together, these data localize cis-acting sequences important in the regulation of human HPRT gene expression and should allow the study of protein-DNA interactions which modulate the transcription of this gene.

Escherichia coli phenylalanyl-tRNA synthetase operon: transcription studies of wild-type and mutated operons on multicopy plasmids

1982 ◽  
Vol 152 (2) ◽  
pp. 661-668
Author(s):  
J A Plumbridge ◽  
M Springer
Keyword(s):  
Structural Gene ◽  
Trna Synthetase ◽  
Mrna Levels ◽  
Structural Genes ◽  
Cis Acting ◽  
Trna Synthetases ◽  
Hybridization Probes ◽  
Steady State Levels ◽  
Derivatives Of

The construction of three lambda bacteriophages containing parts of the structural gene for threonyl-tRNA synthetase, thrS, and those for the two subunits of phenylalanyl-tRNA synthetases, pheS and pheT, is described. These phages were used as hybridization probes to measure the in vivo levels of mRNA specific to these three genes. Plasmid pB1 carries the three genes thrS, pheS, and pheT, and strains carrying the plasmid show enhanced levels of mRNA corresponding to these genes. Although the steady-state levels of threonyl-tRNA synthetase and phenylalanyl-tRNA synthetase produced by the presence of the plasmid differed by a factor of 10, their pulse-labeled mRNA levels were about the same. Mutant derivatives of pB1 were also analyzed. Firstly, a cis-acting insertion located before the structural genes for phenylalanyl-tRNA synthetase caused a major decrease in both pheS and pheT mRNA. Secondly, mutations affecting either structural gene pheS or pheT caused a reduction in the mRNA levels for both pheS and pheT. This observation suggests that autoregulation plays a role in the expression of phenylalanyl-tRNA synthetase.

scholarly journals How affinity of the ELT-2 GATA factor binding to cis-acting regulatory sites controls Caenorhabditis elegans intestinal gene transcription

Development ◽  
2020 ◽  
Vol 147 (14) ◽  
pp. dev190330
Author(s):  
Brett R. Lancaster ◽  
James D. McGhee
Keyword(s):  
Transcription Factor ◽  
Caenorhabditis Elegans ◽  
Binding Affinity ◽  
Experimental System ◽  
Quantitative Relationship ◽  
Gata Factor ◽  
Unknown Parameters ◽  
Cis Acting ◽  
Complicated Model

ABSTRACTWe define a quantitative relationship between the affinity with which the intestine-specific GATA factor ELT-2 binds to cis-acting regulatory motifs and the resulting transcription of asp-1, a target gene representative of genes involved in Caenorhabditis elegans intestine differentiation. By establishing an experimental system that allows unknown parameters (e.g. the influence of chromatin) to effectively cancel out, we show that levels of asp-1 transcripts increase monotonically with increasing binding affinity of ELT-2 to variant promoter TGATAA sites. The shape of the response curve reveals that the product of the unbound ELT-2 concentration in vivo [i.e. (ELT-2free) or ELT-2 ‘activity’] and the largest ELT-XXTGATAAXX association constant (Kmax) lies between five and ten. We suggest that this (unitless) product [Kmax×(ELT-2free) or the equivalent product for any other transcription factor] provides an important quantitative descriptor of transcription-factor/regulatory-motif interaction in development, evolution and genetic disease. A more complicated model than simple binding affinity is necessary to explain the fact that ELT-2 appears to discriminate in vivo against equal-affinity binding sites that contain AGATAA instead of TGATAA.

scholarly journals Chi-Dependent Intramolecular Recombination in Escherichia coli

Genetics ◽  
1998 ◽  
Vol 148 (2) ◽  
pp. 545-557
Author(s):  
Rachel Friedman-Ohana ◽  
Iris Karunker ◽  
Amikam Cohen
Keyword(s):  
Escherichia Coli ◽  
Homologous Recombination ◽  
Experimental System ◽  
Cis Acting ◽  
Enzymes Activity ◽  
Intramolecular Recombination ◽  
Insight Into

Abstract Homologous recombination in Escherichia coli is enhanced by a cis-acting octamer sequence named Chi (5′-GCTGGTGG-3′) that interacts with RecBCD. To gain insight into the mechanism of Chi-enhanced recombination, we recruited an experimental system that permits physical monitoring of intramolecular recombination by linear substrates released by in vivo restriction from infecting chimera phage. Recombination of the released substrates depended on recA, recBCD and cis-acting Chi octamers. Recombination proficiency was lowered by a xonA mutation and by mutations that inactivated the RuvABC and RecG resolution enzymes. Activity of Chi sites was influenced by their locations and by the number of Chi octamers at each site. A single Chi site stimulated recombination, but a combination of Chi sites on the two homologs was synergistic. These data suggest a role for Chi at both ends of the linear substrate. Chi was lost in all recombinational exchanges stimulated by a single Chi site. Exchanges in substrates with Chi sites on both homologs occurred in the interval between the sites as well as in the flanking interval. These observations suggest that the generation of circular products by intramolecular recombination involves Chi-dependent processing of one end by RecBCD and pairing of the processed end with its duplex homolog.

scholarly journals Wilms' Tumor Suppressor GeneWT1

2000 ◽  
Vol 11 (suppl 2) ◽  
pp. S106-S115 ◽  
Author(s):  
CHRISTIAN MROWKA ◽  
ANDREAS SCHEDL
Keyword(s):  
Tumor Suppressor ◽  
Rna Editing ◽  
Kidney Development ◽  
Wilms Tumor ◽  
Suppressor Gene ◽  
Structural Features ◽  
Genomic Locus ◽  
Developmental Abnormalities ◽  
Complex Process

Abstract.Normal development of the kidney is a highly complex process that requires precise orchestration of proliferation, differentiation, and apoptosis. In the past few years, a number of genes that regulate these processes, and hence play pivotal roles in kidney development, have been identified. The Wilms' tumor suppressor geneWT1has been shown to be one of these essential regulators of kidney development, and mutations in this gene result in the formation of tumors and developmental abnormalities such as the Denys-Drash and Frasier syndromes. A fascinating aspect of theWT1gene is the multitude of isoforms produced from its genomic locus. In this review, our current understanding of the structural features ofWT1, how they modulate the transcriptional and post-transcriptional activities of the protein, and how mutations affecting individual isoforms can lead to diseased kidneys is summarized. In addition, results from transgenic experiments, which have yielded important findings regarding the function of WT1in vivo, are discussed. Finally, data on the unusual feature of RNA editing ofWT1transcripts are presented, and the relevance of RNA editing for the normal functioning of the WT1 protein in the kidney is discussed.

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