scholarly journals RNA transcription in myocardial-cell nuclei during postnatal development. A study establishing an assay system for transcription in vitro

1988 ◽  
Vol 256 (2) ◽  
pp. 441-445 ◽  
Author(s):  
J D McCully ◽  
C C Liew
Keyword(s):  
Gene Expression ◽  
Rna Synthesis ◽  
Myocardial Cell ◽  
Assay System ◽  
Specific Gene ◽  
Cell Nuclei ◽  
Rna Transcription ◽  
Nuclear Rna ◽  
Transcription In Vitro

A system for RNA transcription in vitro was established in order to determine the relative rate of RNA synthesis in neonatal and adult rat myocardial cells. This assay system optimizes the incorporation of [3H]UMP into RNA by using 3.5 x 10(7) myocardial-cell nuclei, and minimizes RNA degradation for at least 1 h in transcription in vitro, by the addition of human placental RNAase inhibitor. A 100% increase in the incorporation of [3H]UMP into myocardial-cell RNA was found on addition of this inhibitor. Myocardial-cell nuclei derived from 5-, 10-, 15-, 20-, and greater than 100-day-old rat hearts indicated that there is a progressive decrease in RNA synthesis with age. A 3-fold increase in RNA synthesis in 5-day-old myocardial cell nuclei as compared with 20-day-old rat heart was found. RNA synthesis in the adult myocardial cell nuclei decreased more than 10-fold in comparison with the 5-day-old newborn. The incorporation of [3H]UMP into rat liver nuclear RNA was 3-fold greater than in the myocardial-cell nuclear RNA, even when compared with the highly active transcription of 12-day-old heart nuclei. In order to determine the relationship between total RNA synthesis and the extent of specific gene expression in myocardial-cell nuclei during development, two distinct cDNA probes were used for Northern-blot analysis. Our results indicate that myosin-heavy-chain gene expression is remarkably decreased with age, whereas the ‘housekeeping’ gene is continually expressed independently of age.

RNA transcription and translation in the hearts of normal and cardiomyopathic Syrian hamsters

1991 ◽  
Vol 69 (1) ◽  
pp. 88-92 ◽  
Author(s):  
J. D. McCully ◽  
J. D. Mably ◽  
M. J. Sole ◽  
C. C. Liew
Keyword(s):  
Syrian Hamster ◽  
Myocardial Cell ◽  
In Vitro Translation ◽  
Rna Transcription ◽  
Syrian Hamsters ◽  
Rna Translation ◽  
Nuclear Rna ◽  
Reticulocyte Lysate ◽  
Recessive Defect

The cardiomyopathic Syrian hamster has an autosomal recessive defect that results in the development of an early onset cardiac myopathy leading to cardiac dysfunction and, eventually, complete heart failure. To assess the regulatory mechanisms modulating gene expression in the normal and myopathic myocardium, we investigated both RNA transcription and translation. Our results indicated that the incorporation of [3H]UMP into myocardial cell nuclear RNA decreased 10-fold from 7 to 210 days of age in the normal Syrian hamster. The incorporation of [3H]UMP was approximately 50% lower in the cardiomyopathic as compared with the normal Syrian hamster. RNA translation, as assessed by rabbit reticulocyte lysate in vitro translation, indicated that a coordinated 50% decrease in RNA translation occurred in the normal Syrian hamster from 7 to 210 days of age. A further reduction of 20% in translation was found in cardiomyopathic Syrian hamster ventricular RNA translation as compared with matched random bred control groups. Two-dimensional polyacrylamide gel analysis of cell-free translated protein products demonstrated two myocardial peptides that were found to be consistently altered when the normal and cardiomyopathic Syrian hamsters were compared. These results indicate that transcription and translation decrease with age and that these processes are further downregulated, in an additive manner, with the genesis of the disease process.Key words: cardiomyopathy, ribonucleic acid, transcription, translation.

Female-specific gene expression in Schistosoma mansoni is regulated by pairing

Parasitology ◽  
1997 ◽  
Vol 115 (6) ◽  
pp. 635-640 ◽  
Author(s):  
C. G. GREVELDING ◽  
G. SOMMER ◽  
W. KUNZ
Keyword(s):  
Gene Expression ◽  
Schistosoma Mansoni ◽  
Rna Synthesis ◽  
De Novo ◽  
Transcript Level ◽  
Specific Gene ◽  
Expression Studies ◽  
Gene Expression Studies ◽  
Female Specific

Gene expression studies in adult females of Schistosoma mansoni cultured in vitro revealed that the transcription of female-specifically expressed genes is influenced by pairing. In contrast, the activity of genes that are expressed in both genders was not affected by contact with the male. The transcription of genes was monitored in paired, separated and remated females. The transcript level of female-specifically expressed genes decreases within a few days following separation from males. Remating of uncoupled females with males leads to the reinitiation of transcription. These results provide strong evidence for the influence of the male on gene transcription in the female and contribute a molecular basis for the classical histological observation that the maturation of females is male dependent. The data also show that the culture system is suitable to monitor gene expression and, furthermore, they indicate de novo RNA synthesis in vitro.

Serial analysis of gene expression (SAGE) during porcine embryo development

2004 ◽  
Vol 16 (2) ◽  
pp. 87 ◽  
Author(s):  
Le Ann Blomberg ◽  
Kurt A. Zuelke
Keyword(s):  
Gene Expression ◽  
Functional Genomics ◽  
Embryo Development ◽  
Molecular Mechanisms ◽  
Physiological Role ◽  
Transgenic Animals ◽  
Specific Gene ◽  
Research Strategies

Functional genomics provides a powerful means for delving into the molecular mechanisms involved in pre-implantation development of porcine embryos. High rates of embryonic mortality (30%), following either natural mating or artificial insemination, emphasise the need to improve the efficiency of reproduction in the pig. The poor success rate of live offspring from in vitro-manipulated pig embryos also hampers efforts to generate transgenic animals for biotechnology applications. Previous analysis of differential gene expression has demonstrated stage-specific gene expression for in vivo-derived embryos and altered gene expression for in vitro-derived embryos. However, the methods used to date examine relatively few genes simultaneously and, thus, provide an incomplete glimpse of the physiological role of these genes during embryogenesis. The present review will focus on two aspects of applying functional genomics research strategies for analysing the expression of genes during elongation of pig embryos between gestational day (D) 11 and D12. First, we compare and contrast current methodologies that are being used for gene discovery and expression analysis during pig embryo development. Second, we establish a paradigm for applying serial analysis of gene expression as a functional genomics tool to obtain preliminary information essential for discovering the physiological mechanisms by which distinct embryonic phenotypes are derived.

scholarly journals INHIBITING EFFECT OF THE NEW CYTOTOXIC ANTIBIOTIC DAUNOMYCIN ON NUCLEIC ACIDS AND MITOTIC ACTIVITY OF HELA CELLS

1965 ◽  
Vol 27 (3) ◽  
pp. 545-550 ◽  
Author(s):  
A. Di Marco ◽  
R. Silvestrini ◽  
S. Di Marco ◽  
T. Dasdia
Keyword(s):  
Mitotic Activity ◽  
Rna Synthesis ◽  
Thymidine Incorporation ◽  
Acid Synthesis ◽  
Acetyl Derivative ◽  
Total Inhibition ◽  
Nuclear Rna ◽  
Nucleolar Rna ◽  
Higher Doses

The effect has been studied of Actinomycin D, Daunomycin (Da.), and Da. N acetyl derivative on mitotic activity and on the nucleic acid synthesis of in vitro HeLa cell cultures. The experiments were carried out by means of the radioautographic technique using stripping films. The relative uptake of thymidine-H3 and uridine-H3 was determined by means of the reduced silver grain count present in the nuclei of controls and treated cells. The mitotic activity and thymidine incorporation were noticeably reduced by Daunomycin and Actinomycin, whereas both processes appeared less affected by Da. N acetyl derivative. As regards nuclear RNA synthesis, all three antibiotics at low doses chiefly inhibit nucleolar RNA synthesis. On the other hand, whilst Actinomycin at higher doses causes an almost total inhibition of the synthesis of the whole nuclear RNA, in Daunomycin- and Da. N acetyl derivative-treated cells extranucleolar RNA synthesis is less susceptible to inhibition.

scholarly journals A Dual Role for Oncostatin M Signaling in the Differentiation and Death of Mammary Epithelial Cells in Vivo

2008 ◽  
Vol 22 (12) ◽  
pp. 2677-2688 ◽  
Author(s):  
Paul G. Tiffen ◽  
Nader Omidvar ◽  
Nuria Marquez-Almuina ◽  
Dawn Croston ◽  
Christine J. Watson ◽  
...  
Keyword(s):  
Gene Expression ◽  
Epithelial Cell ◽  
Epithelial Cells ◽  
Mammary Epithelial Cells ◽  
Mammary Epithelial ◽  
Oncostatin M ◽  
Specific Gene ◽  
Mammary Involution

Abstract Recent studies in breast cancer cell lines have shown that oncostatin M (OSM) not only inhibits proliferation but also promotes cell detachment and enhances cell motility. In this study, we have looked at the role of OSM signaling in nontransformed mouse mammary epithelial cells in vitro using the KIM-2 mammary epithelial cell line and in vivo using OSM receptor (OSMR)-deficient mice. OSM and its receptor were up-regulated approximately 2 d after the onset of postlactational mammary regression, in response to leukemia inhibitory factor (LIF)-induced signal transducer and activator of transcription-3 (STAT3). This resulted in sustained STAT3 activity, increased epithelial apoptosis, and enhanced clearance of epithelial structures during the remodeling phase of mammary involution. Concurrently, OSM signaling precipitated the dephosphorylation of STAT5 and repressed expression of the milk protein genes β-casein and whey acidic protein (WAP). Similarly, during pregnancy, OSM signaling suppressed β-casein and WAP gene expression. In vitro, OSM but not LIF persistently down-regulated phosphorylated (p)-STAT5, even in the continued presence of prolactin. OSM also promoted the expression of metalloproteinases MMP3, MMP12, and MMP14, which, in vitro, were responsible for OSM-specific apoptosis. Thus, the sequential activation of IL-6-related cytokines during mammary involution culminates in an OSM-dependent repression of epithelial-specific gene expression and the potentiation of epithelial cell extinction mediated, at least in part, by the reciprocal regulation of p-STAT5 and p-STAT3.

scholarly journals Sex differentiation in grayling (Salmonidae) goes through an all-male stage and is delayed in genetic males who instead grow faster

2017 ◽  
Author(s):  
Diane Maitre ◽  
Oliver M. Selmoni ◽  
Anshu Uppal ◽  
Lucas Marques da Cunha ◽  
Laetitia G. E. Wilkins ◽  
...  
Keyword(s):  
Gene Expression ◽  
Sex Determination ◽  
Sex Differentiation ◽  
Sex Ratios ◽  
Molecular Genetic ◽  
Specific Gene ◽  
Genetic Sexing ◽  
Experimental Conditions ◽  
Two Samples

AbstractFish can be threatened by distorted sex ratios that arise during sex differentiation. It is therefore important to understand sex determination and differentiation, especially in river-dwelling fish that are often exposed to environmental factors that may interfere with sex differentiation. However, sex differentiation is not sufficiently understood in keystone taxa such as the Thymallinae, one of the three salmonid subfamilies. Here we study a wild grayling (Thymallus thymallus) population that suffers from distorted sex ratios. We found sex determination in the wild and in captivity to be genetic and linked to the sdY locus. We therefore studied sex-specific gene expression in embryos and early larvae that were bred and raised under different experimental conditions, and we studied gonadal morphology in five monthly samples taken after hatching. Significant sex-specific changes in gene expression (affecting about 25,000 genes) started around hatching. Gonads were still undifferentiated three weeks after hatching, but about half of the fish showed immature testes around seven weeks after hatching. Over the next few months, this phenotype was mostly replaced by the “testis-to-ovary” or “ovaries” phenotypes. The gonads of the remaining fish, i.e. approximately half of the fish in each sampling period, remained undifferentiated until six months after fertilization. Genetic sexing of the last two samples revealed that fish with undifferentiated gonads were all males, who, by that time, were on average larger than the genetic females (verified in 8-months old juveniles raised in another experiment). Only 12% of the genetic males showed testicular tissue six months after fertilization. We conclude that sex differentiation starts around hatching, goes through an all-male stage for both sexes (which represents a rare case of “undifferentiated” gonochoristic species that usually go through an all-female stage), and is delayed in males who, instead of developing their gonads, grow faster than females during these juvenile stages.Author contributionMRR and CW initiated the project. DM, OS, AU, LMC, LW, and CW sampled the adult fish, did the experimental in vitro fertilizations, and prepared the embryos for experimental rearing in the laboratory. All further manipulations on the embryos and the larvae were done by DM, OS, AU, LMC, and LW. The RNA-seq data were analyzed by OS, JR, and MRR, the histological analyses were done by DM, supervised by SK, and the molecular genetic sexing was performed by DM, OS, AU, and KBM. DM, OS, and CW performed the remaining statistical analyses and wrote the first version of the manuscript that was then critically revised by all other authors.

Changes in liver-specific compared to common gene transcription during primary culture of mouse hepatocytes

1983 ◽  
Vol 3 (9) ◽  
pp. 1552-1561
Author(s):  
D F Clayton ◽  
J E Darnell
Keyword(s):  
Gene Transcription ◽  
Primary Cultures ◽  
Specific Gene ◽  
Cell Nuclei ◽  
Gradual Loss ◽  
Nuclear Rna ◽  
Transcriptional Effect ◽  
Specific Mrnas ◽  
Mouse Hepatocytes ◽  
Specific Mrna

Liver-specific mRNA sequences were examined in primary cultures of mouse hepatocytes. After cell disaggregation by collagenase treatment and for at least 24 h in culture, little change in liver-specific mRNA concentrations was noted. Gradually over a period of 140 h, liver-specific mRNAs declined. In contrast, transcriptional assays in which liver cell nuclei were used to produce 32P-labeled nuclear RNA showed that liver-specific gene transcription was greatly diminished within 24 h, while polymerase II transcription of "common" genes and transcription of tRNA and rRNA did not decline. Thus, a prompt differential transcriptional effect seems to underlie the gradual loss of tissue specificity of the primary cultures.

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