Regulation of IMP dehydrogenase gene expression by its end products, guanine nucleotides

To study the regulation of IMP dehydrogenase (IMPDH), the rate-limiting enzyme of guanine nucleotide biosynthesis, we examined the effects of nucleosides, nucleotides, nucleotide analogs, or the IMPDH inhibitor mycophenolic acid (MPA) on the steady-state levels of IMPDH mRNA. The results indicated that IMPDH gene expression is regulated inversely by the intracellular level of guanine ribonucleotides. We have shown that treatment with guanosine increased the level of cellular guanine ribonucleotides and subsequently reduced IMPDH steady-state mRNA levels in a time- and dose-dependent manner. Conversely, MPA treatment diminished the level of guanine ribonucleotides and increased IMPDH mRNA levels. Both of these effects on the steady-state level of IMPDH mRNA could be negated by cotreatment with guanosine and MPA. The down regulation of IMPDH gene expression by guanosine or its up regulation by MPA was not due to major changes in transcriptional initiation and elongation or mRNA stability in the cytoplasm but rather was due to alterations in the levels of the IMPDH mRNA in the nucleus. These results suggest that IMPDH gene expression is regulated by a posttranscriptional, nuclear event in response to fluctuations in the intracellular level of guanine ribonucleotides.

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Complex transcriptional regulation of myc family gene expression in the developing mouse brain and liver

Molecular and Cellular Biology ◽

10.1128/mcb.11.12.6007-6015.1991 ◽

1991 ◽

Vol 11 (12) ◽

pp. 6007-6015

Author(s):

L Xu ◽

S D Morgenbesser ◽

R A DePinho

Keyword(s):

Gene Expression ◽

Transcriptional Regulation ◽

Steady State ◽

High Rate ◽

Developmental Expression ◽

Tissue Type ◽

Mrna Levels ◽

Postnatal Brain ◽

Transcriptional Initiation ◽

Family Gene

myc family genes (c-, N-, and L-myc) have been shown to be differentially expressed with respect to tissue type and developmental stage. To define and compare the regulatory mechanisms governing their differential developmental expression, we examined the transcriptional regulation of each myc family member during murine postnatal brain and liver development. Nuclear run-on transcription assays demonstrated that both the rate of transcriptional initiation and the degree of transcriptional blocking contribute in a complex manner to the regulation of all three genes. During postnatal brain development, the relative contribution of each transcriptional control mechanism to the regulation of myc family gene expression was found to be different for each gene. For instance, while modulation of transcriptional attenuation did not appear to contribute to the down-regulation of L-myc expression, attenuation was found to be the dominant mechanism by which steady-state N-myc mRNA levels were down-regulated. Different transcriptional strategies were found to be employed in newborn versus adult developing liver for repression of N- and L-myc expression. Undetectable steady-state N- and L-myc mRNA levels in newborn liver were associated with a very low rate of transcriptional initiation, whereas the lack of N- and L-myc expression at the adult stage was accompanied by a high rate of initiation and a striking degree of transcriptional attenuation. Transcriptional attenuation in the N-myc gene was found to map to a region encoding a potential stem-loop structure followed by a thymine tract within the first exon and was not dependent on the use of a specific transcriptional start site.

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Complex transcriptional regulation of myc family gene expression in the developing mouse brain and liver.

Molecular and Cellular Biology ◽

10.1128/mcb.11.12.6007 ◽

1991 ◽

Vol 11 (12) ◽

pp. 6007-6015 ◽

Cited By ~ 13

Author(s):

L Xu ◽

S D Morgenbesser ◽

R A DePinho

Keyword(s):

Gene Expression ◽

Transcriptional Regulation ◽

Steady State ◽

High Rate ◽

Developmental Expression ◽

Tissue Type ◽

Mrna Levels ◽

Postnatal Brain ◽

Transcriptional Initiation ◽

Family Gene

myc family genes (c-, N-, and L-myc) have been shown to be differentially expressed with respect to tissue type and developmental stage. To define and compare the regulatory mechanisms governing their differential developmental expression, we examined the transcriptional regulation of each myc family member during murine postnatal brain and liver development. Nuclear run-on transcription assays demonstrated that both the rate of transcriptional initiation and the degree of transcriptional blocking contribute in a complex manner to the regulation of all three genes. During postnatal brain development, the relative contribution of each transcriptional control mechanism to the regulation of myc family gene expression was found to be different for each gene. For instance, while modulation of transcriptional attenuation did not appear to contribute to the down-regulation of L-myc expression, attenuation was found to be the dominant mechanism by which steady-state N-myc mRNA levels were down-regulated. Different transcriptional strategies were found to be employed in newborn versus adult developing liver for repression of N- and L-myc expression. Undetectable steady-state N- and L-myc mRNA levels in newborn liver were associated with a very low rate of transcriptional initiation, whereas the lack of N- and L-myc expression at the adult stage was accompanied by a high rate of initiation and a striking degree of transcriptional attenuation. Transcriptional attenuation in the N-myc gene was found to map to a region encoding a potential stem-loop structure followed by a thymine tract within the first exon and was not dependent on the use of a specific transcriptional start site.

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Corticotropin-Releasing Factor Inhibits Luteinizing Hormone-Stimulated P450c17 Gene Expression and Androgen Production by Isolated Thecal Cells of Human Ovarian Follicles1

The Journal of Clinical Endocrinology & Metabolism ◽

10.1210/jcem.83.2.4546 ◽

1998 ◽

Vol 83 (2) ◽

pp. 448-452

Author(s):

H. F. Erden ◽

I. H. Zwain ◽

H. Asakura ◽

S. S. C. Yen

Keyword(s):

Gene Expression ◽

Granulosa Cells ◽

Inhibitory Effect ◽

Corticotropin Releasing Factor ◽

Mrna Levels ◽

Dependent Manner ◽

Messenger Ribonucleic Acid ◽

Estrogen Production ◽

Thecal Cells ◽

Crf System

Recently, we reported that the thecal compartment of the human ovary contains a CRF system replete with gene expression and protein for corticotropin-releasing factor (CRF), CRF-Receptor 1 (CRF-R1), and the blood-derived high affinity CRF-binding protein (CRF-BP). Granulosa cells are devoid of the CRF system. The parallel increases in intensity of CRF, CRF-R1, and 17α-hydroxylase messenger ribonucleic acid (mRNA) and proteins in thecal cells with follicular maturation suggest that the intraovarian CRF system may play an autocrine role regulating androgen biosynthesis, with a downstream effect on estrogen production by granulosa cells. The functionality of the ovarian CRF system may be conditioned by the relative presence of plasma-derived CRF-BP by virtue of its localization of protein, but not transcript in thecal cells and its ability to compete with CRF for the CRF receptor. To further these findings, in the present study we have examined the effect of CRF on LH-stimulated 17α-hydroxylase (P450c17) gene expression and androgen production by isolated thecal cells from human ovarian follicles (11–13 mm). During the 48-h culture, addition of LH (10 ng/mL) to the medium increased by 5- and 6-fold dehydroepiandrosterone and androstenedione production by thecal cells. Remarkably, the LH-stimulated, but not basal, androgen production was inhibited by CRF in a time- and dose-dependent manner. The half-maximal (ID50) effect dose of CRF occurred at 5 × 10−8 mol/L, and at a maximal concentration of 10−6 mol/L, CRF completely inhibited LH-stimulated androgen production. This inhibitory effect of CRF became evident at 12 h (45%), and by 24 h the effect was more pronounced, with a 70% reduction from baseline. As determined by Northern analyses, CRF dose dependently decreased LH-stimulated P450c17 mRNA levels, with a maximal inhibition of 85% P450c17 gene expression at a CRF concentration of 10−6 mol/L. With the addition of 10−6 mol/L of the antagonist α-helical CRF-(9–41), the inhibitory effect of CRF was partially reversed for both P450c17 mRNA (75%) and androgen production (50%), indicating the CRF-R1-mediated event. In conclusion, the present study demonstrated a potent inhibitory effect of CRF on LH-stimulated dehydroepiandrosterone and androstenedione production that appears to be mediated through the reduction of P450c17 gene expression. Thus, the ovarian CRF system may function as autocrine regulators for androgen biosynthesis in the thecal cell compartment to maintain optimal substrate for estrogen biosynthesis by granulosa cells. Further studies to define the role of CRF-BP in the endocrine modulation of the intraovarian CRF system are needed.

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Codon replacement in the PGK1 gene of Saccharomyces cerevisiae: experimental approach to study the role of biased codon usage in gene expression

Molecular and Cellular Biology ◽

10.1128/mcb.7.8.2914-2924.1987 ◽

1987 ◽

Vol 7 (8) ◽

pp. 2914-2924

Author(s):

A Hoekema ◽

R A Kastelein ◽

M Vasser ◽

H A de Boer

Keyword(s):

Gene Expression ◽

Saccharomyces Cerevisiae ◽

Steady State ◽

Codon Usage ◽

Experimental Approach ◽

Mrna Translation ◽

Yeast Cells ◽

Expression Level ◽

Start Codon ◽

Mrna Levels

The coding sequences of genes in the yeast Saccharomyces cerevisiae show a preference for 25 of the 61 possible coding triplets. The degree of this biased codon usage in each gene is positively correlated to its expression level. Highly expressed genes use these 25 major codons almost exclusively. As an experimental approach to studying biased codon usage and its possible role in modulating gene expression, systematic codon replacements were carried out in the highly expressed PGK1 gene. The expression of phosphoglycerate kinase (PGK) was studied both on a high-copy-number plasmid and as a single copy gene integrated into the chromosome. Replacing an increasing number (up to 39% of all codons) of major codons with synonymous minor ones at the 5' end of the coding sequence caused a dramatic decline of the expression level. The PGK protein levels dropped 10-fold. The steady-state mRNA levels also declined, but to a lesser extent (threefold). Our data indicate that this reduction in mRNA levels was due to destabilization caused by impaired translation elongation at the minor codons. By preventing translation of the PGK mRNAs by the introduction of a stop codon 3' and adjacent to the start codon, the steady-state mRNA levels decreased dramatically. We conclude that efficient mRNA translation is required for maintaining mRNA stability in S. cerevisiae. These findings have important implications for the study of the expression of heterologous genes in yeast cells.

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Abstract 2423: B-type Natriuretic Peptide Upregulates Erythropoietin Receptor Gene Expression via Protein Kinase G in Heart Failure

Circulation ◽

10.1161/circ.118.suppl_18.s_724 ◽

2008 ◽

Vol 118 (suppl_18) ◽

Author(s):

Yoshiaki Ohyama ◽

Toru Tanaka ◽

Takehisa Shimizu ◽

Hiroshi Doi ◽

Norimichi Koitabashi ◽

...

Keyword(s):

Gene Expression ◽

Heart Failure ◽

Protein Kinase ◽

Cardiac Myocytes ◽

Natriuretic Peptide ◽

Neonatal Rat ◽

Mrna Levels ◽

Dependent Manner ◽

Failing Heart ◽

Epor Expression

Backgroud: Recent studies demonstrated non-hematopoietical effects of Erythropoietin (Epo) and its receptor (EpoR) in a variety of tissues including cardiovascular system. Epo treatment improves cardiac function in patients with heart failure and reduces infarct size after ischemia/reperfusion injury in the heart. However, little attention has been paid for the endogenous regulatory mechanisms regulating EpoR expression. In this study, we hypothesize that B-type natriuretic peptide upregulates EpoR gene expression in failing heart. Methods and Results: Wister rats underwent transverse aortic constriction surgery to induce hypertrophy. RT-PCR analyses of those rats showed that EpoR mRNA levels were increased in the left ventricle and positively correlated with the levels of BNP mRNA (n=10, r=0.67, p<0.05). Next we examined the expression of EpoR in human failing heart by using autopsy specimens and found that EpoR mRNA levels were significantly elevated in patients with dilated cardiomyopathy compared with those in normal heart. Immunohistochemistry of endomyocardial biopsy specimens of failing heart (n=54) showed that EpoR mRNA levels were correlated with severity of cardiac dysfunction estimated by diameter of cardiac chambers, pathomorphology, serum BNP concentration and functional class of New York Heart Association. Interestingly, stimulation of cultured neonatal rat cardiac myocytes with BNP, but not with hypertrophic reagents including endothelin I, angiotensin II and norepinephrine, significantly increased the EpoR mRNA levels in a time-dependent manner. Overexpression of cGMP-dependent protein kinase (PKG) increased EpoR transcript in cultured cardiac myocytes. BNP-induced EpoR expression was abrogated in the presence of KT5823, a specific inhibitor for PKG. Conclusion: These results suggest a role for BNP in mediating an induction of EpoR expression in failing myocardium and indicate that the cardiac EpoR gene is a target of cGMP/PKG signaling.

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Interferon-gamma downregulates CFTR gene expression in epithelial cells

AJP Cell Physiology ◽

10.1152/ajpcell.1994.267.5.c1398 ◽

1994 ◽

Vol 267 (5) ◽

pp. C1398-C1404 ◽

Cited By ~ 75

Author(s):

F. Besancon ◽

G. Przewlocki ◽

I. Baro ◽

A. S. Hongre ◽

D. Escande ◽

...

Keyword(s):

Gene Expression ◽

Interferon Gamma ◽

Posttranscriptional Regulation ◽

Bacterial Infections ◽

Cftr Gene ◽

Mrna Levels ◽

Dependent Manner ◽

Necrosis Factor Alpha ◽

Ifn Gamma ◽

Cl Transport

Cystic fibrosis (CF) is caused by mutations in the CF transmembrane conductance regulator (CFTR) gene, resulting in defective transepithelial Cl- transport. The regulation of CF gene expression is not fully understood. We report that interferon-gamma (IFN-gamma), but not IFN-alpha or -beta, downregulates CFTR mRNA levels in two colon-derived epithelial cell lines, HT-29 and T84, in a time- and concentration (from 0.1 IU/ml)-dependent manner. IFN-gamma has no effect on the transcription rate of the CFTR gene but reduces CFTR mRNA half-life, indicating that it exerts a posttranscriptional regulation of CFTR expression, at least partly, through destabilization of the transcripts. Cells treated with IFN-gamma contain subnormal amounts of 165-kDa CFTR protein. Assays of adenosine 3',5'-cyclic monophosphate-stimulated 36Cl- efflux and whole cell currents show that CFTR function is diminished in IFN-gamma-treated cells. IFN-gamma and tumor necrosis factor-alpha synergistically reduce CFTR gene expression. Our results suggest that production of these cytokines in response to bacterial infections and inflammatory disorders may alter transmembrane Cl- transport.

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Genotype-dependent and non-gradient patterns of RSV gene expression

10.1101/641878 ◽

2019 ◽

Author(s):

Felipe-Andrés Piedra ◽

Xueting Qiu ◽

Michael N. Teng ◽

Vasanthi Avadhanula ◽

Annette A. Machado ◽

...

Keyword(s):

Gene Expression ◽

Transcriptional Regulation ◽

Steady State ◽

Transcription Initiation ◽

Mrna Levels ◽

Viral Pathogen ◽

Negative Strand ◽

Gene Start ◽

Syncytial Virus ◽

Conserved Gene

AbstractRespiratory syncytial virus (RSV) is a nonsegmented negative-strand (NNS) RNA virus and a leading cause of severe lower respiratory tract illness in infants and the elderly. Transcription of the ten RSV genes proceeds sequentially from the 3’ promoter and requires conserved gene start (GS) and gene end (GE) signals. Previous studies using the prototypical GA1 genotype Long and A2 strains have indicated a gradient of gene transcription. However, recent reports show data that appear inconsistent with a gradient. To better understand RSV transcriptional regulation, mRNA abundances from five RSV genes were measured by quantitative real-time PCR (qPCR) in three cell lines and cotton rats infected with virus isolates belonging to four different genotypes (GA1, ON, GB1, BA). Relative mRNA levels reached steady-state between four and 24 hours post-infection. Steady-state patterns were genotype-specific and non-gradient, where mRNA levels from the G (attachment) gene exceeded those from the more promoter-proximal N (nucleocapsid) gene across isolates. Transcript stabilities could not account for the non-gradient patterns observed, indicating that relative mRNA levels more strongly reflect transcription than decay. While the GS signal sequences were highly conserved, their alignment with N protein in the helical ribonucleocapsid, i.e., N-phase, was variable, suggesting polymerase recognition of GS signal conformation affects transcription initiation. The effect of GS N-phase on transcription efficiency was tested using dicistronic minigenomes. Ratios of minigenome gene expression showed a switch-like dependence on N-phase with a period of seven nucleotides. Our results indicate that RSV gene expression is in part sculpted by polymerases that initiate transcription with a probability dependent on GS signal N-phase.Author SummaryRSV is a major viral pathogen that causes significant morbidity and mortality, especially in young children. Shortly after RSV enters a host cell, transcription from its nonsegmented negative-strand (NNS) RNA genome starts at the 3’ promoter and proceeds sequentially. Transcriptional attenuation is thought to occur at each gene junction, resulting in a gradient of gene expression. However, recent studies showing non-gradient levels of RSV mRNA suggest that transcriptional regulation may have additional mechanisms. We show using RSV isolates belonging to four different genotypes that gene expression is genotype-dependent and one gene (the G or attachment gene) is consistently more highly expressed than an upstream neighbor. We hypothesize that variable alignment of highly conserved gene start (GS) signals with nucleoprotein (i.e., variable GS N-phase) can affect transcription and give rise to non-gradient patterns of gene expression. We show using dicistronic RSV minigenomes wherein the reporter genes differ only in the N-phase of one GS signal that GS N-phase affects gene expression. Our results suggest the existence of a novel mechanism of transcriptional regulation that might play a role in other NNS RNA viruses.

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Gene expression of human DNA polymerase alpha during cell proliferation and the cell cycle

Molecular and Cellular Biology ◽

10.1128/mcb.8.11.5016-5025.1988 ◽

1988 ◽

Vol 8 (11) ◽

pp. 5016-5025

Author(s):

A F Wahl ◽

A M Geis ◽

B H Spain ◽

S W Wong ◽

D Korn ◽

...

Keyword(s):

Gene Expression ◽

Cell Cycle ◽

Cell Proliferation ◽

Steady State ◽

Dna Polymerase ◽

Transformed Cells ◽

Mrna Levels ◽

Dna Polymerase Alpha ◽

Human Dna ◽

Alpha Gene

We studied the expression of the human DNA polymerase alpha gene during cell proliferation, during cell progression through the cell cycle, and in transformed cells compared with normal cells. During the activation of quiescent cells (G0 phase) to proliferate (G1/S phases), the steady-state mRNA levels, rate of synthesis of nascent polymerase protein, and enzymatic activity in vitro exhibited a substantial and concordant increase prior to the peak of in vivo DNA synthesis. In transformed cells, the respective values were amplified greater than 10-fold. In actively growing cells separated into discrete stages of the cell cycle by counterflow elutriation or by mitotic shakeoff, levels of steady-state transcripts, translation rates, and enzymatic activities of polymerase alpha were constitutively and concordantly expressed at all stages of the cell cycle, with only a moderate elevation prior to the S phase and a slight decline in the G2 phase. These findings support the conclusion that the regulation of human DNA polymerase alpha gene expression is at the transcriptional level and strongly suggest that the regulatory mechanisms that are operative during the entrance of a cell into the mitotic cycle are fundamentally different from those that modulate polymerase alpha expression in continuously cycling cells.

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Changes in Gene Expression of the MCU Complex Are Induced by Electrical Stimulation in Adult Skeletal Muscle

Frontiers in Physiology ◽

10.3389/fphys.2020.601313 ◽

2021 ◽

Vol 11 ◽

Author(s):

Esteban R. Quezada ◽

Alexis Díaz-Vegas ◽

Enrique Jaimovich ◽

Mariana Casas

Keyword(s):

Gene Expression ◽

Skeletal Muscle ◽

Electrical Stimulation ◽

Muscle Fibers ◽

Low Frequency ◽

Extracellular Atp ◽

Mrna Levels ◽

Dependent Manner ◽

Adult Skeletal Muscle ◽

Muscle Plasticity

The slow calcium transient triggered by low-frequency electrical stimulation (ES) in adult muscle fibers and regulated by the extracellular ATP/IP3/IP3R pathway has been related to muscle plasticity. A regulation of muscular tropism associated with the MCU has also been described. However, the role of transient cytosolic calcium signals and signaling pathways related to muscle plasticity over the regulation of gene expression of the MCU complex (MCU, MICU1, MICU2, and EMRE) in adult skeletal muscle is completely unknown. In the present work, we show that 270 0.3-ms-long pulses at 20-Hz ES (and not at 90 Hz) transiently decreased the mRNA levels of the MCU complex in mice flexor digitorum brevis isolated muscle fibers. Importantly, when ATP released after 20-Hz ES is hydrolyzed by the enzyme apyrase, the repressor effect of 20 Hz on mRNA levels of the MCU complex is lost. Accordingly, the exposure of muscle fibers to 30 μM exogenous ATP produces the same effect as 20-Hz ES. Moreover, the use of apyrase in resting conditions (without ES) increased mRNA levels of MCU, pointing out the importance of extracellular ATP concentration over MCU mRNA levels. The use of xestospongin B (inhibitor of IP3 receptors) also prevented the decrease of mRNA levels of MCU, MICU1, MICU2, and EMRE mediated by a low-frequency ES. Our results show that the MCU complex can be regulated by electrical stimuli in a frequency-dependent manner. The changes observed in mRNA levels may be related to changes in the mitochondria, associated with the phenotypic transition from a fast- to a slow-type muscle, according to the described effect of this stimulation frequency on muscle phenotype. The decrease in mRNA levels of the MCU complex by exogenous ATP and the increase in MCU levels when basal ATP is reduced with the enzyme apyrase indicate that extracellular ATP may be a regulator of the MCU complex. Moreover, our results suggest that this regulation is part of the axes linking low-frequency stimulation with ATP/IP3/IP3R.

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