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Acting Factors (acting + factor)
Selected AbstractsA trans -acting factor, isolated by the three-hybrid system, that influences alternative splicing of the amyloid precursor protein minigeneFEBS JOURNAL, Issue 13 2000Andrej Poleev Two clones were isolated in a three-hybrid screen of a rat fetal brain P5 cDNA library with an intronic splicing enhancer of the amyloid precursor protein (APP) gene as RNA bait. These clones represent the rat homologues of the previously described genes CUG-binding protein (CUG-BP) and Siah-binding protein (Siah-BP). Both interact in a sequence-specific manner with the RNA bait used for library screening as well as with the CUG repeat. In contrast, no interactions were observed in the three-hybrid assay with other baits tested. In two-hybrid assays, Siah-BP interacts with U2AF65 as well as with itself. EWS, an RGG-type RNA-binding protein associated with Ewing sarcoma, was identified as an interacting partner for the CUG-BP homologue in a two-hybrid assay for protein,protein interactions performed with various factors involved in RNA metabolism. Splicing assays performed by RT-PCR from cells cotransfected with certain cDNAs and an APP minigene, used as a reporter, indicate exclusion of exon 8 if the CUG-BP homologue is present. We conclude that clone AF169013 and its counterpart in human CUG-BP could be the trans -acting factors that interact with the splicing enhancer downstream of exon 8, and in this way influence alternative splicing of the APP minigene. [source] Roles of CmpR, a LysR family transcriptional regulator, in acclimation of the cyanobacterium Synechococcus sp. strain PCC 7942 to low-CO2 and high-light conditionsMOLECULAR MICROBIOLOGY, Issue 3 2004Yukari Takahashi Summary The cmp operon of Synechococcus sp. strain PCC 7942, encoding a high-affinity bicarbonate transporter, is induced under low CO2 conditions by a LysR family protein CmpR. CmpR was found to be required also for induction of the operon by transfer of the cells from low-light to high-light conditions, indicating involvement of a common mechanism in the high-light- and low-CO2 -responsive regulation. Expression of the high-light inducible genes psbAII and psbAIII, on the other hand, was found to be induced also by low-CO2 conditions. A single promoter was responsible for the high-light and low-CO2 induction of each of psbAII and psbAIII, suggesting involvement of a common regulatory mechanism in the light and CO2 responses of the psbA genes. CmpR was, however, not required for the induction of psbAII and psbAIII, indicating the presence of multiple mechanisms for induction of genes under high-light and low-CO2 conditions. The CmpR-deficient mutant nevertheless showed lower levels of the psbAII and psbAIII transcripts than the wild-type strain under all the light and CO2 conditions examined. Gel shift assays showed that CmpR binds to the enhancer elements of psbAII and psbAIII, through specific interaction with a sequence signature conforming to the binding motif of similar LysR family proteins. These findings showed that CmpR acts as a trans -acting factor that enhances transcription of the photosystem II genes involved in acclimation to high light, revealing a complex network of gene regulation in the cyanobacterium. [source] Mechanisms of imprint dysregulation,AMERICAN JOURNAL OF MEDICAL GENETICS, Issue 3 2010Bernhard Horsthemke Abstract Genomic imprinting is an epigenetic process by which the male and the female germ line confer specific marks (imprints) onto certain gene regions, so that one allele of an imprinted gene is active and the other allele is silent. Genomic imprints are erased in primordial germ cells, newly established during later stages of germ cell development, and stably inherited through somatic cell divisions during postzygotic development. Defects in imprint erasure, establishment, or maintenance result in a paternal chromosome carrying a maternal imprint or in a maternal chromosome carrying a paternal imprint. A wrong imprint leads to activation of an allele that should be silent or silencing of an allele that should be active. Since the dosage of imprinted genes is very important for development and growth, imprinting defects lead to specific diseases. Imprinting defects can occur spontaneously without any DNA sequence change (primary imprinting defect) or as the result of a mutation in a cis -regulatory element or a trans -acting factor (secondary imprinting defect). The distinction between primary and secondary imprinting defects is important for assessing the recurrence risk in affected families. © 2010 Wiley-Liss, Inc. [source] Functional analysis of the osteoarthritis susceptibility,associated GDF5 regulatory polymorphismARTHRITIS & RHEUMATISM, Issue 7 2009Rainer J. Egli Objective Single-nucleotide polymorphism (SNP) rs143383 (T to C) in the 5,-untranslated region (5,-UTR) of GDF5 has recently been reported to be associated with osteoarthritis (OA) susceptibility, with lower expression of the risk-associated T allele observed in vitro and in vivo. The in vivo studies were performed on cartilage tissue from OA patients. The present study was undertaken to expand the analysis of the effect of this SNP on GDF5 allelic expression to more joint tissue types, to investigate for cis and trans factors that interact with the SNP, and to examine novel cis -acting GDF5 regulatory polymorphisms. Methods Tissue samples were collected from OA patients undergoing joint replacement of the hip or knee. Nucleic acid was extracted, and, using rs143383 and an assay that discriminates and quantifies allelic expression, the relative amount of GDF5 expression from the T and C alleles was measured. Additional common variants in the GDF5 transcript sequence were interrogated as potential regulatory elements using allelic expression and luciferase reporter assays, and electrophoretic mobility shift assays were used to search for trans factors binding to rs143383. Results We observed a consistent allelic expression imbalance of GDF5 in all tissues tested, implying that the functional effect mediated by rs143383 on GDF5 expression is joint-wide. We identified a second polymorphism, located in the 3,-UTR of GDF5, that influenced allelic expression of the gene independent of rs143383. Finally, we observed differential binding of deformed epidermal autoregulatory factor 1 (DEAF-1) to the 2 alleles of rs143383. Conclusion These findings show that the OA susceptibility mediated by polymorphism in GDF5 is not restricted to cartilage, emphasizing the need to consider the disease as involving the whole joint. The existence of an additional cis -acting regulatory polymorphism highlights the complexity of the regulation of expression of this important OA susceptibility locus. DEAF-1 is a trans -acting factor that merits further investigation as a potential tool for modulating GDF5 expression. [source] Crystallization and preliminary crystallographic analysis of the second RRM of Pub1 from Saccharomyces cerevisiaeACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 2 2009Yingji Cui mRNA stability is elaborately regulated by elements in the mRNA transcripts and their cognate RNA-binding proteins, which play important roles in regulating gene expression at the post-transcriptional level in eukaryotes. Poly(U)-binding protein 1 (Pub1), which is a major nuclear and cytoplasmic polyadenylated RNA-binding protein in Saccharomyces cerevisiae, is involved in the regulation of mRNA turnover as a trans -acting factor. It binds to transcripts containing the AU-rich element in order to protect them from degradation. Pub1 contains three RNA-recognition motifs (RRMs) which play significant roles in mRNA binding at AU-rich elements and stabilizer elements. In this study, the second RRM of Pub1 was crystallized by the hanging-drop vapour-diffusion method using polyethylene glycol 4000 as a precipitant at 283,K. An X-ray diffraction data set was collected using a single flash-cooled crystal that belonged to space group H3. [source] Utp25p, a nucleolar Saccharomyces cerevisiae protein, interacts with U3 snoRNP subunits and affects processing of the 35S pre-rRNAFEBS JOURNAL, Issue 13 2010Mauricio B. Goldfeder In eukaryotes, pre-rRNA processing depends on a large number of nonribosomal trans -acting factors that form intriguingly organized complexes. Two intermediate complexes, pre-40S and pre-60S, are formed at the early stages of 35S pre-rRNA processing and give rise to the mature ribosome subunits. Each of these complexes contains specific pre-rRNAs, some ribosomal proteins and processing factors. The novel yeast protein Utp25p has previously been identified in the nucleolus, an indication that this protein could be involved in ribosome biogenesis. Here we show that Utp25p interacts with the SSU processome proteins Sas10p and Mpp10p, and affects 18S rRNA maturation. Depletion of Utp25p leads to accumulation of the pre-rRNA 35S and the aberrant rRNA 23S, and to a severe reduction in 40S ribosomal subunit levels. Our results indicate that Utp25p is a novel SSU processome subunit involved in pre-40S maturation. Structured digital abstract ,,MINT-7889901: SAS10 (uniprotkb:Q12136) physically interacts (MI:0915) with Utp25p (uniprotkb:P40498) by pull down (MI:0096) ,,MINT-7889915: NIP7 (uniprotkb:Q08962) physically interacts (MI:0915) with RRP43 (uniprotkb:P25359) by two hybrid (MI:0018) ,,MINT-7889852: Utp25p (uniprotkb:P40498) physically interacts (MI:0915) with MPP10 (uniprotkb:P47083) by two hybrid (MI:0018) ,,MINT-7890065: NOP1 (uniprotkb:P15646) and Utp25p (uniprotkb:P40498) colocalize (MI:0403) by fluorescence microscopy (MI:0416) ,,MINT-7889865: Utp25p (uniprotkb:P40498) physically interacts (MI:0915) with SAS10 (uniprotkb:Q12136) by two hybrid (MI:0018) [source] A trans -acting factor, isolated by the three-hybrid system, that influences alternative splicing of the amyloid precursor protein minigeneFEBS JOURNAL, Issue 13 2000Andrej Poleev Two clones were isolated in a three-hybrid screen of a rat fetal brain P5 cDNA library with an intronic splicing enhancer of the amyloid precursor protein (APP) gene as RNA bait. These clones represent the rat homologues of the previously described genes CUG-binding protein (CUG-BP) and Siah-binding protein (Siah-BP). Both interact in a sequence-specific manner with the RNA bait used for library screening as well as with the CUG repeat. In contrast, no interactions were observed in the three-hybrid assay with other baits tested. In two-hybrid assays, Siah-BP interacts with U2AF65 as well as with itself. EWS, an RGG-type RNA-binding protein associated with Ewing sarcoma, was identified as an interacting partner for the CUG-BP homologue in a two-hybrid assay for protein,protein interactions performed with various factors involved in RNA metabolism. Splicing assays performed by RT-PCR from cells cotransfected with certain cDNAs and an APP minigene, used as a reporter, indicate exclusion of exon 8 if the CUG-BP homologue is present. We conclude that clone AF169013 and its counterpart in human CUG-BP could be the trans -acting factors that interact with the splicing enhancer downstream of exon 8, and in this way influence alternative splicing of the APP minigene. [source] Closely linked cis -acting modifier of expansion of the CGG repeat in high risk FMR1 haplotypes,HUMAN MUTATION, Issue 12 2007S. Ennis Abstract In its expanded form, the fragile X triplet repeat at Xq27.3 gives rise to the most common form of inherited mental retardation, fragile X syndrome. This high population frequency persists despite strong selective pressure against mutation-bearing chromosomes. Males carrying the full mutation rarely reproduce and females heterozygous for the premutation allele are at risk of premature ovarian failure. Our diagnostic facility and previous research have provided a large databank of X chromosomes that have been tested for the FRAXA allele. Using this resource, we have conducted a detailed genetic association study of the FRAXA region to determine any cis -acting factors that predispose to expansion of the CGG triplet repeat. We have genotyped SNP variants across a 650-kb tract centered on FRAXA in a sample of 877 expanded and normal X chromosomes. These chromosomes were selected to be representative of the haplotypic diversity encountered in our population. We found expansion status to be strongly associated with a ,50-kb region proximal to the fragile site. Subsequent detailed analyses of this region revealed no specific genetic determinants for the whole population. However, stratification of chromosomes by risk subgroups enabled us to identify a common SNP variant which cosegregates with the subset of D group haplotypes at highest risk of expansion (,=17.84, p=0.00002). We have verified that this SNP acts as a marker of repeat expansion in three independent samples. Hum Mutat 28(12), 1216,1224, 2007. © 2007 Wiley-Liss, Inc. [source] Developmental regulation of the glyoxylate cycle in the human pathogen Penicillium marneffeiMOLECULAR MICROBIOLOGY, Issue 6 2006David Cánovas Summary Penicillium marneffei is a thermally dimorphic opportunistic human pathogen with a saprophytic filamentous hyphal form at 25°C and a pathogenic unicellular yeast form at 37°C. During infection. P. marneffei yeast cells exist intracellularly in macrophages. To cope with nutrient deprivation during the infection process, a number of pathogens employ the glyoxylate cycle to utilize fatty acids as carbon sources. The genes which constitute this pathway have been implicated in pathogenesis. To investigate acetate and fatty acid utilization, the acuD gene encoding a key glyoxylate cycle enzyme (isocitrate lyase) was cloned. The acuD gene is regulated by both carbon source and temperature in P. marneffei, being strongly induced at 37°C even in the presence of a repressing carbon source such as glucose. When introduced into the non-pathogenic monomorphic fungus Aspergillus nidulans, the P. marneffei acuD promoter only responds to carbon source. Similarly, when the A. nidulans acuD promoter is introduced into P. marneffei it only responds to carbon source suggesting that P. marneffei possesses both cis elements and trans -acting factors to control acuD by temperature. The Zn(II)2Cys6 DNA binding motif transcriptional activator FacB was cloned and is responsible for carbon source-, but not temperature-, dependent induction of acuD. The expression of acuD at 37°C is induced by AbaA, a key regulator of morphogenesis in P. marneffei, but deletion of abaA does not completely eliminate temperature-dependent induction, suggesting that acuD and the glyoxylate cycle are regulated by a complex network of factors in P. marneffei which may contribute to its pathogenicity. [source] Regulation of fungal gene expression via short open reading frames in the mRNA 5,untranslated regionMOLECULAR MICROBIOLOGY, Issue 4 2003Cristina Vilela Summary We review how the expression of fungal mRNAs can be controlled by ribosome interactions with short upstream open reading frames (uORFs) within the 5,untranslated region. The efficiency of uAUG recognition modulates the impact of a uORF but steps during and after translation of the uORF also influence uORF function. The post-termination behaviour of ribosomes, therefore, plays a major role in determining the expression level of these main ORFs. Translation of a uORF can produce a cis -acting peptide that causes effector molecule-dependent stalling of the ribosomes at the end of the uORF. In other cases it is the length or position, or other features of the uORF, rather than the peptide it encodes, that determine the efficiency with which ribosomes reinitiate translation downstream of it. Whether the form of the ribosome that resumes scanning after termination is the 40S subunit alone or the entire 80S ribosome is not known. Translation of the uORF can also control gene expression by affecting the stability of the mRNA. Finally, trans -acting factors may participate in the regulatory mechanisms. Future work will need not only to provide more information on the mechanisms underlying the known cases of uORF-mediated control but also to define the full complement of uORF-containing mRNAs in at least one fungal organism. [source] An embryonic story: Analysis of the gene regulative network controlling Xist expression in mouse embryonic stem cellsBIOESSAYS, Issue 7 2010Pablo Navarro Abstract In mice, dosage compensation of X-linked gene expression is achieved through the inactivation of one of the two X-chromosomes in XX female cells. The complex epigenetic process leading to X-inactivation is largely controlled by Xist and Tsix, two non-coding genes of opposing function. Xist RNA triggers X-inactivation by coating the inactive X, while Tsix is critical for the designation of the active X-chromosome through cis -repression of Xist RNA accumulation. Recently, a plethora of trans -acting factors and cis -regulating elements have been suggested to act as key regulators of either Xist, Tsix or both; these include ubiquitous factors such as Yy1 and Ctcf, developmental proteins such as Nanog, Oct4 and Sox2, and X-linked regulators such as Rnf12. In this paper we summarise recent advances in our knowledge of the regulation of Xist and Tsix in embryonic stem (ES) and differentiating ES cells. [source] Genomic mutation rates: what high-throughput methods can tell usBIOESSAYS, Issue 9 2009Koodali T. Nishant Abstract High-throughput DNA analyses are increasingly being used to detect rare mutations in moderately sized genomes. These methods have yielded genome mutation rates that are markedly higher than those obtained using pre-genomic strategies. Recent work in a variety of organisms has shown that mutation rate is strongly affected by sequence context and genome position. These observations suggest that high-throughput DNA analyses will ultimately allow researchers to identify trans -acting factors and cis sequences that underlie mutation rate variation. Such work should provide insights on how mutation rate variability can impact genome organization and disease progression. [source] Wingless can't fly so it hitches a ride with dyneinBIOESSAYS, Issue 10 2001Steven H. Myster Asymmetric RNA localization is required for many developmental processes in a wide range of organisms. For example, wingless and pair-rule transcripts are localized to the apical membrane of polarized cells. It has been unclear, however, if this localization is important for biological activity and, in addition, how the transcripts are transported. Two recent studies(1,2) have identified cis -elements and trans -acting factors that are required for the asymmetric localization of mRNAs. Correct localization is shown to be required for biological activity, and a mechanism of RNA transport involving the microtubule motor dynein has been revealed. BioEssays 23:869,872, 2001. © 2001 John Wiley & Sons, Inc. [source] | |||||||||||||