Biomedical subjects

En Li

Publications and source records attributed to En Li.

62 records · Page 4Linked to original sources

PubMed2002

Neurobehavioral and electroencephalographic abnormalities in Ube3a maternal-deficient mice.

Angelman syndrome (AS), characterized by motor dysfunction, mental retardation, and seizures, is caused by several genetic etiologies involving chromosome 15q11-q13, including mutations of the UBE3A gene. UBE3A encodes UBE3A/E6-AP, a ubiquitin-protein ligase, and shows brain-specific imprinting, with brain expression predominantly from the maternal allele. Lack of a functional maternal allele of UBE3A causes AS. In order to understand the causal relationship between maternal UBE3A mutations and AS, we have constructed a mouse model with targeted inactivation of Ube3a. The inactive allele contains a lacZ reporter gene for analysis of brain-specific imprinting. Maternal, but not paternal, transmission of the targeted allele leads to beta-galactosidase activity in hippocampal and cerebellar neurons. Maternal inheritance of the Ube3a mutant allele also causes impaired performance in tests of motor function and spatial learning, as well as abnormal hippocampal EEG recordings. As predicted from the dependence of UBE3A-mediated ubiquitination of p53 on HPV E6 protein, our maternal-deficient mice show normal brain p53 levels.

Angelman Syndrome↗

PubMed2002

Chromatin modification and epigenetic reprogramming in mammalian development.

The developmental programme of embryogenesis is controlled by both genetic and epigenetic mechanisms. An emerging theme from recent studies is that the regulation of higher-order chromatin structures by DNA methylation and histone modification is crucial for genome reprogramming during early embryogenesis and gametogenesis, and for tissue-specific gene expression and global gene silencing. Disruptions to chromatin modification can lead to the dysregulation of developmental processes, such as X-chromosome inactivation and genomic imprinting, and to various diseases. Understanding the process of epigenetic reprogramming in development is important for studies of cloning and the clinical application of stem-cell therapy.

Animals↗

PubMed2002

Inhibition of Nodal signalling by Lefty mediated through interaction with common receptors and efficient diffusion.

BACKGROUND: Two TGFbeta-related proteins, Nodal and Lefty, are implicated in early embryonic patterning of vertebrates. Genetic data suggest that Nodal is a signalling molecule, while Lefty is an antagonist of Nodal, but their precise function remains unknown. RESULTS: The signalling pathway of Nodal was investigated with the use of a Nodal-responsive assay system based on frog animal caps. Expression of dominant negative mutants of various receptors indicated that ALK4, and either ActRIIA or ActRIIB, function as type I and type II receptors for Nodal, respectively. A soluble form of Cripto lacking the COOH-terminal region interacted with Nodal but failed to mediate Nodal signalling, indicating that the native Cripto protein functions as a membrane-bound co-receptor for Nodal. Processed forms of Lefty proteins, both smaller and larger forms, inhibited Nodal signalling. Such Lefty-induced inhibition was rescued by excess ActRIIA or ActRIIB, suggesting that Lefty antagonizes Nodal signalling through competitive binding to the common receptor ActRIIA or ActRIIB. This idea was supported by the demonstration of a genetic interaction between lefty2 and ActRIIB in mouse. Behaviours of GFP-Nodal and GFP-Lefty2 proteins were also investigated in chick embryos. Both proteins could diffuse over a long distance, but the latter diffused faster than the former. CONCLUSIONS: Efficient inhibition of Nodal signals by Lefty may involve competitive binding of Lefty to the common receptors and faster diffusion of Lefty.

Activin Receptors, Type I↗

PubMed2002

Genetic analyses of DNA methyltransferase genes in mouse model system.

DNA methylation regulates important biological processes and is involved in tumorigenesis and several human diseases, such as Rett and immunodeficiency, centromeric instability and facial anomalies (ICF). The major objective of our research is to investigate the roles of DNA methylation in mammals through genetic analysis of DNA methyltransferase genes in mouse and human. Previously, we found that Dnmt1 knockout embryonic stem (ES) cells are capable of methylating retroviral DNA de novo. In search of enzymes responsible for de novo methylation, we have cloned a novel family of mammalian DNA methyltransferase genes, Dnmt3a and Dnmt3b. Although extensive sequence similarity was found between Dnmt3a and Dnmt3b, little homology was observed between Dnmt1 and Dnmt3a/3b in the catalytic domain as well as in the N-terminal domain. Additionally, biochemical analysis revealed that, unlike Dnmt1, neither Dnmt3a nor Dnmt3b had a strong preference to hemimethylated DNA substrates. Genetic analysis demonstrated that Dnmt3a and Dnmt3b were required for de novo methylation activities in ES cells and during early embryogenesis and were essential for early development. Interestingly, phenotype analyses of single homozygous mice for either Dnmt3a or Dnmt3b suggested that the functions of Dnmt3a and Dnmt3b also were required at the late developmental stage and even at the adult stage.

Animals↗

PubMed2002

Cooperativity between DNA methyltransferases in the maintenance methylation of repetitive elements.

We used mouse embryonic stem (ES) cells with systematic gene knockouts for DNA methyltransferases to delineate the roles of DNA methyltransferase 1 (Dnmt1) and Dnmt3a and -3b in maintaining methylation patterns in the mouse genome. Dnmt1 alone was able to maintain methylation of most CpG-poor regions analyzed. In contrast, both Dnmt1 and Dnmt3a and/or Dnmt3b were required for methylation of a select class of sequences which included abundant murine LINE-1 promoters. We used a novel hemimethylation assay to show that even in wild-type cells these sequences contain high levels of hemimethylated DNA, suggestive of poor maintenance methylation. We showed that Dnmt3a and/or -3b could restore methylation of these sequences to pretreatment levels following transient exposure of cells to 5-aza-CdR, whereas Dnmt1 by itself could not. We conclude that ongoing de novo methylation by Dnmt3a and/or Dnmt3b compensates for inefficient maintenance methylation by Dnmt1 of these endogenous repetitive sequences. Our results reveal a previously unrecognized degree of cooperativity among mammalian DNA methyltransferases in ES cells.

Animals↗

PubMed2002

Dnmt3L cooperates with the Dnmt3 family of de novo DNA methyltransferases to establish maternal imprints in mice.

Genomic imprinting is regulated by differential methylation of the paternal and maternal genome. However, it remains unknown how parental imprinting is established during gametogenesis. In this study, we demonstrate that Dnmt3L, a protein sharing homology with DNA methyltransferases, Dnmt3a and Dnmt3b, but lacking enzymatic activity, is essential for the establishment of maternal methylation imprints and appropriate expression of maternally imprinted genes. We also show that Dnmt3L interacts with Dnmt3a and Dnmt3b and co-localizes with these enzymes in the nuclei of transfected cells, suggesting that Dnmt3L may regulate genomic imprinting via the Dnmt3 family enzymes. Consistent with this model, we show that [Dnmt3a(-/-), Dnmt3b(+/-)] mice also fail to establish maternal methylation imprints. In addition, both Dnmt3a and Dnmt3L are required for spermatogenesis. Together, our findings suggest that Dnmt3L may cooperate with Dnmt3 family methyltransferases to carry out de novo methylation of maternally imprinted genes in oocytes.

3T3 Cells↗

PubMed2002

[Effects of genistein on proliferation and differentiation of cultured osteoblasts in vitro].

OBJECTIVE: To study the effects of genistein on proliferation and differentiation of cultured osteoblasts in vitro. METHODS: MTT and PNPP were used separately to observe the proliferation and the activity of ALP of osteoblasts cultured in vitro with different concentration genistein for different incubation periods. Protein was measured by Lowry method. RESULTS: It was found that genistein had the effects on promoting proliferation of osteoblasts cultured for 24 h and 48 h at concentration ranging from 1 x 10(-9) mol/L to 1 x 0(-5) mol/L. The activity of ALP was stimulated by genistein at concentration ranging from 1 x 10(-7) mol/L to 1 x 10(-5) mol/L after 72 h incubation. CONCLUSION: Genistein has the effects on stimulating the proliferation and differentiation of cultured osteoblasts in vitro.

Alkaline Phosphatase↗

PubMedDate not supplied

5-aza-2'-deoxycytidine induces alterations in murine spermatogenesis and pregnancy outcome.

Because of the ability of cytidine analogues, such as 5-aza-2'-deoxycytidine, to incorporate into DNA and lead to decreases in DNA methylation, there has recently been renewed interest in using these drugs in anticancer therapy. To determine the effects of paternal 5-aza-2'-deoxycytidine treatment on spermatogenesis and progeny outcome in the mouse and whether effects are modulated by decreased levels of the predominant DNA methyltransferase, DNMT1, adult Dnmt1(+/+) and Dnmt1-deficient (Dnmt1(c/+)) male mice were treated with 5-aza-2'-deoxycytidine for 7 weeks, which resulted in dose-dependent decreases in testicular weight, an increase in histological abnormalities, and a decline in sperm counts, with no apparent effect on androgen status. Testes of Dnmt1(c/+) mice, however, were less severely affected by 5-aza-2'-deoxycytidine than were those of wild-type mice. The exposure of Dnmt1(+/+) male mice to even low doses of 5-aza-2'-deoxycytidine followed by mating elicited significantly reduced pregnancy rates and elevated preimplantation loss in females. Dnmt1 deficiency, however, protected against such drug-induced decreases in pregnancy rate but not preimplantation loss. Altered DNA methylation or DNMT1 activity may explain such adverse effects, because treatment resulted in dose-dependent decreases in the global methylation of sperm DNA. Thus, in the mouse, paternal administration of 5-aza-2'-deoxycytidine interferes with normal male germ cell development and results in reduced fertility, whereas lowering DNMT1 levels appears to partially protect the seminiferous epithelium from deleterious drug effects.

Animals↗