[The modification of mumps virus RNA genome and regulation of the genomic expression in BHK21/WI-2 cells persistently infected with mumps virus].
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Animals↗
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Results for “RNA Regulation”
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Mechanism and regulation of yeast RNA polymerase II transcription.
Fractionation of yeast whole-cell extract has led to the identification and purification of four novel proteins required for promoter-dependent transcription by RNA polymerase II in vitro. These initiation factors have been shown to be related in structure and function to RNA polymerase II initiation factors purified from mammalian cells. When combined with the well known TATA-binding protein, the entire set of initiation factors proves necessary and sufficient for accurate initiation from a variety of eukaryotic promoters. Studies with this defined transcription system have revealed the existence of additional proteins required for transcriptional activation or derepression by enhancer-binding regulatory proteins.
Cell-Free System↗
Pursuing gene regulation 'logic' via RNA interference and chromatin immunoprecipitation.
RNA interference and chromatin immunoprecipitation are now firmly established as useful methods for studying mechanisms of gene regulation in vivo. Their combined use can help elucidate gene regulation 'logic' by aiding in target gene identification for transcription factors and chromatin-modifying complexes.
Animals↗
RNA localization and translational regulation during axis specification in the Drosophila oocyte.
The major axes of the oocyte-antero-posterior and dorso-ventral-are established over a one-day period during mid-oogenesis in Drosophila. The same molecule, GURKEN (GRK), functions to initiate signaling between the oocyte and the surrounding, somatically derived follicle cells. This results first in specification of the antero-posterior axis and, later, the dorso-ventral axis of the oocyte and surrounding follicle cells. Central to specification of both axes is a combination of cytoplasmic localization and translational regulation of the grk RNA. Here we discuss the mechanisms by which the grk RNA is localized within the oocyte and the role of translational regulation in spatially restricting the production of GRK protein. We then discuss the generality of these mechanisms during oogenesis by focusing on a second transcript, oskar, whose function is also regulated through a combination of transcript localization and translational control.
Animals↗
Transcriptional regulation of messenger RNA for ferritin heavy chain by thyrotropin.
Thyrotropin increased the steady state levels of ferritin heavy chain messenger RNA in cultured Fisher rat thyroid (FRTL5) cells by about 2.5-fold. Thyrotropin also stimulated the transcription rate of ferritin H gene determined by "nuclear run-on" assay by roughly the same extent as mRNA levels. Thyrotropin showed no effect on the stability of the ferritin heavy chain mRNA, which was suggested using actinomycin D. The results suggest that thyrotropin increases ferritin heavy chain mRNA expression in FRTL5 cells by affecting the step of transcription.
Animals↗
Plant RNA polymerases: structures, regulation, and genes.
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DNA-Directed RNA Polymerases↗
Functional interactions of RNA-capping enzyme with factors that positively and negatively regulate promoter escape by RNA polymerase II.
Capping of the 5' ends of nascent RNA polymerase II transcripts is the first pre-mRNA processing event in all eukaryotic cells. Capping enzyme (CE) is recruited to transcription complexes soon after initiation by the phosphorylation of Ser-5 of the carboxyl-terminal domain of the largest subunit of RNA polymerase II. Here, we analyze the role of CE in promoter clearance and its functional interactions with different factors that are involved in promoter clearance. FCP1-mediated dephosphorylation of the carboxyl-terminal domain results in a drastic decrease in cotranscriptional capping efficiency but is reversed by the presence of DRB sensitivity-inducing factor (DSIF). These results suggest involvement of DSIF in CE recruitment. Importantly, CE relieves transcriptional repression by the negative elongation factor, indicating a critical role of CE in the elongation checkpoint control mechanism during promoter clearance. This functional interaction between CE and the negative elongation factor documents a dynamic role of CE in promoter clearance beyond its catalytic activities.
Nuclear Proteins↗
Interferon induction by viruses. XXII. Vesicular stomatitis virus-Indiana: M-protein and leader RNA do not regulate interferon induction in chicken embryo cells.
Several field isolates, strains, mutants, and revertants of vesicular stomatitis virus (VSV), Indiana (IN) serotype, were studied that differed greatly in their capacity to induce interferon (IFN) in aged chick embryo cells. The predicted M-protein amino acid sequence of a wild-type field isolate that induced > or = 10,000 units/ml IFN in chicken embryo cells was identical to that of a wild-type field isolate that induced < 2 units/ml and of a noninducing wild-type laboratory strain. The 47-base plus-strand leader RNA sequences were the same for five IFN-inducing, and eight noninducing independent isolates of wild-type VSV IN. Our data show that the M-protein and plus-strand leader RNA do not of themselves regulate the induction of IFN in this system. Because the capacity of VSV IN to induce IFN resides in virion-associated elements (Marcus and Sekellick, 1987, J. Interferon Res. 7, 269-284), the differences in IFN yield observed with various isolates must result from changes in other virion components that remain to be determined.
Amino Acid Sequence↗
Binding of RNA to p53 regulates its oligomerization and DNA-binding activity.
The C-terminus of p53 is responsible for maintaining the latent, non-DNA-binding form of p53. However, the mechanism by which the C-terminus regulates DNA binding is not yet fully understood. We show here that p53 interacts with RNA via its C-terminal domain and that disruption of this interaction, by RNase A treatment, truncation or phosphorylation of the C-terminus, restores DNA-binding activity. Furthermore, the oligomerization of p53 is significantly enhanced by disrupting the interaction between p53 and RNA. These findings suggest that binding of RNA to p53 is involved in the mechanism of p53 latency.
Biopolymers↗
The association of TIF-IA and polymerase I mediates promoter recruitment and regulation of ribosomal RNA transcription in Acanthamoeba castellanii.
Large amounts of energy are expended for the construction of the ribosome during both transcription and processing, so it is of utmost importance for the cell to efficiently regulate ribosome production. Understanding how this regulation occurs will provide important insights into cellular growth control and into the coordination of gene expression mediated by all three transcription systems. Ribosomal RNA (rRNA) transcription rates closely parallel the need for protein synthesis; as a cell approaches stationary phase or encounters conditions that negatively affect either growth rate or protein synthesis, rRNA transcription is decreased. In eukaryotes, the interaction of RNA polymerase I (pol I) with the essential transcription initiation factor IA (TIF-IA) has been implicated in this downregulation of transcription. In agreement with the first observation that rRNA transcription is regulated by altering recruitment of pol I to the promoter in Acanthamoeba castellanii, we show here that pol I and an 80-kDa homologue of TIF-IA are found tightly associated in pol I fractions competent for specific transcription. Disruption of the pol I-TIF-IA complex is mediated by a specific dephosphorylation of either pol I or TIF-IA. Phosphatase treatment of TIF-IA-containing A. castellanii pol I fractions results in a downregulation of both transcriptional activity and promoter binding, reminiscent of the inactive pol I fractions purified from encysted cells. The fraction of pol I competent for promoter recruitment is enriched in TIF-IA relative to that not bound by immobilized promoter DNA. This downregulation coincides with an altered electrophoretic mobility of TIF-IA, suggesting at least it is phosphorylated.
Acanthamoeba castellanii↗
Protein synthesis-dependent and -independent mechanisms for the regulation of GnRH RNA transcript levels in GT1 cells.
The cellular mechanism for the suppression of GnRH gene expression by the phorbol ester PMA was investigated in GT1 cells. The protein synthesis inhibitor cycloheximide decreased GnRH primary transcript levels, indicating a protein synthesis requirement for basal GnRH transcription. PMA decreased GnRH primary transcript levels even in the presence of cycloheximide, indicating that the PMA suppression of GnRH gene transcription is protein synthesis-independent. In contrast, the PMA-inhibitory effect on GnRH cytoplasmic mRNA levels was significantly reduced or inhibited in the presence of cycloheximide or RNA synthesis inhibitors given within 4 h of PMA, suggesting a protein/RNA synthesis-dependent mechanism for the regulation of GnRH mRNA levels by PMA. Thus, the mechanism for the PMA inhibition of GnRH primary transcript is mediated through a protein and RNA synthesis-independent mechanism, while the decrease in GnRH mRNA levels occurs through a mechanism that involves the induction of new RNA and protein synthesis that happens within 4 h of PMA administration.
Animals↗
Tomato bushy stunt virus genomic RNA accumulation is regulated by interdependent cis-acting elements within the movement protein open reading frames.
This study on Tomato bushy stunt virus identified and defined three previously unknown regulatory sequences involved in RNA accumulation that are located within the 3'-proximal nested movement protein genes p22 and p19. The first is a 16-nucleotide (nt) element termed III-A that is positioned at the very 3' end of p22 and is essential for RNA accumulation. Approximately 300 nt upstream of III-A resides an approximately 80-nt inhibitory element (IE) that is obstructive to replication only in the absence of a third regulatory element of approximately 30 nt (SUR-III) that is positioned immediately upstream of III-A. Inspection of the nucleotide sequences predicted that III-A and SUR-III can form looped hairpins. A comparison of different tombusviruses showed, in each case, conservation for potential base pairing between the two predicted hairpin-loops. Insertion of a spacer adjacent to the predicted hairpins had no or a minimal effect on RNA accumulation, whereas an insertion in the putative III-A loop abolished genomic RNA multiplication. We conclude that the sequences composing the predicted III-A and SUR-III hairpin-loops are crucial for optimal RNA accumulation and that the inhibitory effect of IE surfaces when the alleged interaction between SUR-III and III-A is disturbed.
Genes, Regulator↗
Mechanistic analysis of RNA polymerase III regulation by the retinoblastoma protein.
The tumour suppressor protein RB restricts cellular growth. This may involve inhibiting the synthesis of tRNA and 5S rRNA by RNA polymerase (pol) III. We have shown previously that RB can repress pol III transcription when overexpressed either in vitro or in vivo. We also demonstrated that pol III activity is elevated substantially in primary fibroblasts from RB-deficient mice. Here we address the molecular mechanism of this regulation. RB is shown to repress all types of pol III promoter. It can do this even if added after transcription complex assembly. Functional assays demonstrate that RB targets specifically the general pol III factor TFIIIB. A physical interaction between TFIIIB and RB is indicated by fractionation, pull-down and immunoprecipitation data. We show that TFIIIB activity is elevated in primary fibroblasts from RB-deficient mice. TFIIIB is a multisubunit complex that includes the TATA-binding protein (TBP) and a TFIIB-related factor called BRF. We show that RB itself contains regions of homology to both TBP and BRF and propose a model in which RB disrupts TFIIIB by mimicking these two components.
Amino Acid Sequence↗
Regulation of ribosomal RNA promoters with a synthetic lac operator.
A synthetic 21-base-pair long DNA fragment containing the central lac operator sequence has been inserted near the initiation point of the cloned Escherichia coli rrnB rRNA promoter P2 in the natural and reverse orientation. RNA synthesis is efficiently repressed in both orientations in lac Iq strains and is induced with isopropyl beta-D-thiogalactoside. When the rrnB promoter P1 is also present, upstream from P2 and the synthetic lac operator, repression of transcription is incomplete. The levels of transcription were measured in vivo, indirectly by the expression of a protein (chloramphenicol acetyltransferase), or directly by the expression of a stable RNA (E. coli 4.5S RNA) in a simple assay involving gel electrophoresis of unlabeled total RNA from E. coli. The rrnB promoter constructions can produce high levels of protein expression as well as high levels of expression of stable RNA.
Acetyltransferases↗
[RNA biosynthesis and regulation of protein biosynthesis in the liver of chicks with experimental coccidiosis].
The aim of this work was to investigate the interrelationship between RNA biosynthesis and that of protein in chick liver during experimental coccidiosis induced by E. tenella. The peculiarity of this model is that in the course of this disease protein synthesis is significantly intensified inspite of the fact that the rate of the biosynthesis is rather high under normal conditions. It has been shown that 4 to 6 days after infection incorporation of labeled amino acids into proteins from chick liver subcellular fractions is greatly increased. The most pronounced changes are in ribosomal and mitochondrial fractions as well as in the postribosomal supernatant. At the same time the specific radioactivity of serum albumin excreted by liver was increased by factor 3. These changes in protein biosynthesis are associated with a significant increase of both the content and intensity of biosynthesis of high molecular weight precursors of rRNA as well as with those of mature 18S rRNA. The amount of 28S rRNA and mRNA per cell is practically without any changes whereas the mRNA turnover is somewhat more extensive. The selective accumulation of 18S rRNA is suggested to be responsible for the intensification of protein biosynthesis.
Animals↗
[Glucocorticoids specifically regulate the expression of dispersed repeating sequences in liver cells. Detection of small ID-like antisense RNA].
Differential regulation of the expression of various families of the rat genome interspersed repetitive sequences by glucocorticoid hormones in liver has been demonstrated. Small ID-like RNA complementarily associated with high-molecular-weight cytoplasmic poly(A)-containing RNAs has been identified. The results obtained indicate that glucocorticoids hormones induce expression of the B2, ID, and L1 families in rat liver. The content of the transcripts of these repetitive elements is increased by the hormones in the molecules of heterogeneous nuclear RNA. In the molecules of high-molecular-mass poly(A)-containing cytoplasmic RNA glucocorticoids increase the content of the ID-homologous sequences. In the population of the small specific RNA the content of the ID-homologous transcripts is also increased. A suggestion is put forward about the involvement of the ID-like element in the hormonal posttranscriptional regulation of gene expression at the mRNA level.
Animals↗
Mutually exclusive exon splicing of type III brain sodium channel alpha subunit RNA generates developmentally regulated isoforms in rat brain.
We have identified two exons of the type III rat brain sodium channel alpha subunit gene that undergo mutually exclusive alternative RNA splicing to produce mRNAs coding either for an isoform predominant in neonatal brain (IIIN) or a different isoform (IIIA) predominant in the adult. These exons are 92 base pairs in length and encode amino acids 203-232, which correspond to part of the S3 and most of the S4 transmembrane segments within domain I and the extracellular loop between them. Despite 21 nucleotide differences between the exons, only a single amino acid at position 209 is altered, specifying either aspartic acid (IIIA) or serine (IIIN). As evidence that these isoforms are generated via alternative splicing, we demonstrate that both exons are encoded within the type III gene. The nucleotide sequences of the neonatal and adult type III exons and the intervening intron as well as the developmental regulation of this splicing are nearly identical in the type II sodium channel gene. The conservation of the exon/intron structure and of the developmentally regulated patterns of expression of the type II and III sodium channel genes suggests that alternative mRNA splicing of this exon may play a substantial role in modulating sodium channel function during brain development by alteration of a single amino acid.
Animals↗
Epidermal growth factor expression in human colon and colon carcinomas: anti-sense epidermal growth factor receptor RNA down-regulates the proliferation of human colon cancer cells.
Human colon cancer cell lines express epidermal growth factor (EGF) mRNA, secrete EGF and may respond to it via the cell-surface EGF receptor (EGFR). Expression of these molecules in human colon and colon tumor, however, is not clear. Reverse transcription-polymerase chain reaction (RT-PCR) analyses of RNA prepared from paired normal human colon and colon tumor samples from 12 individuals followed by Southern blotting analyses of the RT-PCR products revealed a major fragment of 527 bp and a minor fragment of 404 bp that hybridized to a human EGF cDNA probe under stringent conditions. Identical results were obtained from 8 human colon cancer cell lines. Cloning and sequencing of PCR products confirmed that both fragments were from the human EGF gene; the 527-bp fragment corresponded exactly to nucleotides 2,891 to 3,417 of the human EGF mRNA reported by others. A deletion of 123 nucleotides (nucleotides 3,172 to 3,294) was found in the 404-bp fragment. Immunohistochemical studies using cyostat sections of human colon specimens showed that EGF was expressed in the human colon and that expression was restricted to the epithelial colonic crypt cells and epithelium-derived cancer cells. Since EGF and EGF-related molecules are potent mitogens that mediated their effect through the EGFR, we also determined the efficacy of anti-sense EGFR RNA in circumventing the EGFR-related pathway of proliferation. Expression of anti-sense EGFR RNA, by transfection with an inducible anti-sense EGFR expression vector, down-regulated cell-surface EGFR expression and proliferation of these cells and their ability to grow in soft agar. Anti-sense EGFR RNA was found to be an anti-proliferative agent in both relatively non-aggressive and highly aggressive human colon cancer cells.
Adenocarcinoma↗