Biomedical subjects

Luisa Dandolo

Publications and source records attributed to Luisa Dandolo.

8 recordsLinked to original sources

PubMed2005

The Prader-Willi syndrome murine imprinting center is not involved in the spatio-temporal transcriptional regulation of the Necdin gene.

BACKGROUND: The human Prader-Willi syndrome (PWS) domain and its mouse orthologue include a cluster of paternally expressed genes which imprinted expression is co-ordinately regulated by an imprinting center (IC) closely associated to the Snurf-Snrpn gene. Besides their co-regulated imprinted expression, two observations suggest that the spatio-temporal expression of these genes could also be co-regulated. First, the PWS genes have all been reported to be expressed in the mouse nervous system. Second, Snurf-Snrpn and its associated IC are the most ancient elements of the domain which later acquired additional functional genes by retrotransposition. Although located at least 1.5 megabases from the IC, these retroposons acquired the same imprinted regulation as Snurf-Snrpn. In this study, we ask whether the IC, in addition to its function in imprinting, could also be involved in the spatio-temporal regulation of genes in the PWS domain. RESULTS: We compared the expression pattern of Snurf-Snrpn and C/D-box small nucleolar RNAs (snoRNAs) MBII-85 and MBII-52 to the expression pattern of the two evolutionary related retroposons Ndn and Magel2, in the developing mouse embryo. We show that these genes have highly similar expression patterns in the central nervous system, suggesting that they share a common central nervous system-specific regulatory element. Among these genes, Ndn and Magel2 display the most similar expression patterns. Using transgenic mice containing the Ndn and Magel2 genes, we show that the transgenic Ndn gene whereas not imprinted is correctly expressed. Search for DNase I hypersensitive sites in the Ndn-Magel2 genomic region and comparative genomic analyses were performed in order to identify potential transcriptional cis-regulatory elements. CONCLUSIONS: These results strongly suggest that paternally expressed genes of the PWS domain share a common central nervous system-specific regulatory element. We proposed that this regulatory element could co-localize with the IC. However, we demonstrate that the IC, if required for imprinted regulation, is not involved in the spatio-temporal regulation of distantly located retrotransposed genes such as the Ndn gene in the PWS domain.

Animals↗

PubMed2005

[Epigenetics and development: genomic imprinting].

Genomic imprinting leads to parent-of-origin-specific monoallelic expression of about 60 known genes in the mammalian genome. It was discovered 20 years ago and the aim of this review is to summarize its main characteristics. The nature of the imprint, still unknown, is characterized by differential chromatin structure and DNA methylation. The imprint is reset at each generation during gametogenesis, which can be observed by demethylation in the PGCs, then gamete-specific remethylation. The imprinted genes are usually located in clusters and regulated by cis sequences such as imprinting centres, trans factors such as the insulator protein CTCF and/or large non coding antisense RNAs. Genetic and epigenetic abnormalities of the imprinted clusters can lead to human diseases such as Prader-Willi, Angelman or Beckwith-Wiedemann syndromes.

Animals↗

PubMed2004

An enhancer element at the Igf2/H19 locus drives gene expression in both imprinted and non-imprinted tissues.

The insulin-like growth factor 2 (Igf2) gene encodes a potent growth factor that is expressed in multiple tissues during embryonic development. Expression at this locus is mediated by genomic imprinting. In the developing endodermal tissues, imprinting of Igf2 is mediated by the interaction of a set of enhancers downstream of the linked H19 gene with a differentially methylated domain (DMD) that lies approximately 2-4 kb upstream of H19 that has a boundary or insulator function in the hypomethylated state. In the remainder of tissues that express Igf2 and H19, the cis elements that drive their correct expression and imprinting are not well understood. In addition, enhancers driving expression of Igf2 in the choroid plexus and leptomeninges, tissues where the gene is thought not to be imprinted, have not been isolated. Here we show that biallelic (non-imprinted) expression within the choroid plexus is restricted to the epithelium, and we provide evidence that a conserved intergenic region functions as an enhancer for Igf2 both in tissues where the gene is imprinted, and where Igf2 is biallelically expressed. The presence of an enhancer for imprinted tissues in the intergenic region argues for the existence of imprinting controls distinct from the DMD, which may be provided by differential methylation at sites proximal to Igf2.

Animals↗

PubMed2003

Developmentally regulated functions of the H19 differentially methylated domain.

Igf2 and H19 are physically linked imprinted genes. In embryonic liver, their reciprocal expression (paternal for Igf2 and maternal for H19) is controlled by a paternally methylated region (H19 DMD) located 5' of H19. This region contains a methylation-sensitive insulator that prevents the Igf2 promoters being activated by downstream enhancers on the maternal chromosome. In adult liver, Igf2 is normally not expressed but is reactivated upon tumour formation. By analysing three deletions of the H19 locus, we investigated the mechanism regulating the imprinted expression of the Igf2 gene in the course of liver tumourigenesis. We observed that the role of the H19 DMD in the control of Igf2 expression changes during tumourigenesis. The H19 DMD is required on the paternal chromosome for Igf2 activation in the early stages while its maternal allele is necessary for maintaining Igf2 imprinting only in the late stages. A positive regulatory function of the paternal H19 DMD is also evident in normal neonatal liver, but its relevance for Igf2 expression becomes higher in the second post-natal week. Our results support a model in which both methylated and non-methylated parental copies of the H19 DMD have active roles in the regulation of Igf2 expression in the liver and these activities are under developmental control.

Animals↗

PubMed2003

Disruption of the nonneuronal tph1 gene demonstrates the importance of peripheral serotonin in cardiac function.

Serotonin (5-HT) controls a wide range of biological functions. In the brain, its implication as a neurotransmitter and in the control of behavioral traits has been largely documented. At the periphery, its modulatory role in physiological processes, such as the cardiovascular function, is still poorly understood. The rate-limiting enzyme of 5-HT synthesis, tryptophan hydroxylase (TPH), is encoded by two genes, the well characterized tph1 gene and a recently identified tph2 gene. In this article, based on the study of a mutant mouse in which the tph1 gene has been inactivated by replacement with the beta-galactosidase gene, we establish that the neuronal tph2 is expressed in neurons of the raphe nuclei and of the myenteric plexus, whereas the nonneuronal tph1, as detected by beta-galactosidase expression, is in the pineal gland and the enterochromaffin cells. Anatomic examination of the mutant mice revealed larger heart sizes than in wild-type mice. Histological investigation indicates that the primary structure of the heart muscle is not affected. Hemodynamic analyses demonstrate abnormal cardiac activity, which ultimately leads to heart failure of the mutant animals. This report links loss of tph1 gene expression, and thus of peripheral 5-HT, to a cardiac dysfunction phenotype. The tph1-/- mutant may be valuable for investigating cardiovascular dysfunction observed in heart failure in humans.

Alleles↗

PubMed2003

A real-time polymerase chain reaction assay for quantification of allele ratios and correction of amplification bias.

Allele-specific epigenetic modifications are crucial for several important biological functions, including genomic imprinting and X-inactivation in mammals. Consequently, an ever increasing number of investigations requires accurate quantification of the relative abundance of parental alleles of a specific sequence in a DNA sample. Here, combining the use of polymorphic restriction sites with real-time polymerase chain reaction (PCR) amplification, we describe a simple and quantitative assay to measure allele ratios. The efficiency of the assay was assessed on genomic DNA for several polymorphic restriction sites located in the mouse Igf2/H19 imprinted locus. The assay was also successfully applied to quantify allele ratio in cDNA samples. In addition, we provide an experimental procedure for detection and correction of potential PCR amplification bias which significantly extends the range of application of the assay.

Data Interpretation, Statistical↗

PubMed2002

Turnover of primary transcripts is a major step in the regulation of mouse H19 gene expression.

In the gene expression pathway, RNA biogenesis is a central multi-step process where both message fidelity and steady-state levels of the mature RNA have to be ascertained. An emerging question is whether RNA levels could be regulated at the precursor stage. Until recently, because it was technically very difficult to determine the level of a pre-mRNA, discrimination between changes in transcriptional activity and in pre-mRNA metabolism was extremely difficult. H19 RNA, the untranslated product of an imprinted gene, undergoes post-transcriptional regulation. Here, using a quantitative real-time RT-PCR approach, we accurately quantify its precursor RNA levels and compare these with the transcriptional activity of the gene, assessed by run-on assays. We find that the levels of H19 precursor RNA are regulated during physiological processes and this regulation appears to be related to RNA polymerase II transcription termination. Our results provide direct evidence that turnover of polymerase II primary transcripts can regulate gene expression in mammals.

Animals↗

PubMed2002

Mouse parthenogenetic embryos with monoallelic H19 expression can develop to day 17.5 of gestation.

In mammals, both maternal and paternal genomes are required for a fetus to develop normally to term. This requirement is due to the epigenetic modification of genomes during gametogenesis, which leads to an unequivalent expression of imprinted genes between parental alleles. Parthenogenetic mouse embryos that contain genomes from nongrowing (ng) and fully grown (fg) oocytes can develop into 13.5-day-old fetuses, in which paternally and maternally expressed imprinted genes are expressed and repressed, respectively, from the ng oocyte allele. The H19 gene, however, is biallelically expressed with the silent status Igf2 in such parthenotes. In this study, we examined whether the regulation of H19 monoallelic expression enhances the survival of parthenogenetic embryos. The results clearly show that the ng(H19-KO)/fg(wt) parthenogenetic embryos carrying the ng-oocyte genome that had been deleted by the H19 transcription unit successfully developed as live fetuses for 17.5 gestation days. Control experiments revealed that this unique phenomenon occurs irrespective of the genetic background effect. Quantitative gene expression analysis showed that day 12.5 ng(H19-KO)/fg(wt) parthenogenetic fetuses expressed Igf2 and H19 genes at <2 and 82% of the levels in the controls. Histological analysis demonstrated that the placenta of ng(H19-KO)/fg(wt) parthenotes was afflicted with atrophia with severe necrosis and other anomalies. The present results suggest that the cessation of H19 gene expression from the ng-allele causes extended development of the fetus and that functional defects in the placenta could be fatal for the ontogeny.

Alleles↗