Different Imprinting Status of IGF-2 in Epithelial Ov arian Tumors

To explore whether the imprinting status of IGF 2 in the malignant epithelial ovarian tumors is different from that in benign tumors, the target sequences (DNA and RNA) which contain a polymorphism site for ApaI restriction endonuclease digestion were amplified with PCR and RT PCR methods. Then the PCR/RT PCR products were digested by ApaI. The IGF 2 transcriptional pattern came out from the results of endonucleases digestion. Among the 36 cases of benign epithelial ovarian tumors, 20 were heterozygous for ApaI locus and all showed genomic imprinting. While in the malignant group, 22 were heterozygous for ApaI locus but six were found to lose imprinting. Significant differences existed between the two groups ( P <0.05). Loss of imprinting of IGF 2 may serve as a marker for differentiating the malignant ovarian cancers from the benign ones. In a new field of molecular genetics, our research provides an experimental basis for genetic diagnosis and treatment of the ovarian cancers.

IDENTIFICATION OF HUMAN MURR1, THE HOMOLOGUE OF MOUSE IMPRINTED Murr1 GENE

Objective To identify the mRNA sequence, genetic construction, imprinting status, and expression profile of human MURR1 gene, the homologue of mouse imprinted Murr1 gene. Methods The MURR1 mRNA sequence was identified by colony hybridization screening of human cDNA library and the 5'-RACE analyses; Absence of U2AF1-RS1 gene within MURR1 was confirmed by Southern Blotting; Expression profile of MURR1 was examined by Northern Blotting; The imprinting status of MURR1 were revealed by SNP investigation and RT-PCR followed by sequencings and RFLP analyses. Results The full-length mRNA sequence of MURR1 spans 711 bp, transcribed from 3 exons, encodes predicted MURR1 protein of 190 amino acids. The gene was expressed in all the 12 kinds of human adult tissues and 6 kinds of fetal tissues. It showed biallelic expression in all 32 investigated samples including 6 kinds of human fetal tissues and 8 adult brains. Unlike mouse imprinted U2af1-rs1 gene existing in the intron of Murr1, the human U2AF1-RS1 gene was not located in the MURR1 locus. Conclusion Human MURR1 gene is not imprinted and the non-imprinting is possible due to the absence of human homologue of mouse U2af1-rs1 within MURR1 locus.

The maternally expressed polycomb group gene OsEMF2a is essential for endosperm cellularization and imprinting in rice

Cellularization is a key event in endosperm development.Polycomb group(PcG)genes,such as Fertilization-Independent Seed 2(FIS2),are vital for the syncytium-to-cellularization transition in Arabidopsis plants.In this study,we found that OsEMF2a,a rice homolog of the Arabidopsis PcG gene Embryonic Flower2(EMF2),plays a role similar to that of FIS2 in regard to seed development,although there is limited sequence similarity between the genes.Delayed cellularization was observed in osemf2a,associated with an unusual activation of type I MADS-box genes.The cell cycle was persistently activated in osemf2a caryopses,which was likely caused by cytokinin overproduction.However,the overaccumulation of auxin was not found to be associated with the delayed cellularization.As OsEMF2a is a maternally expressed gene in the endosperm,a paternally inherited functional allele was unable to recover the maternal defects of OsEMF2a.Many imprinted rice genes were deregulated in the defective hybrid seeds of osemf2a(♀)/9311(♂)(m9).The paternal expression bias of some paternally expressed genes was disrupted in m9 due to either the activation of maternal alleles or the repression of paternal alleles.These findings suggest that OsEMF2a-PRC2-mediated H3K27me3 is necessary for endosperm cellularization and genomic imprinting in rice.

Haploid embryonic stem cells:an ideal tool for mammalian genetic analyses

Identification of the function of all genes in the mammalian genome is critical in understanding basic mechanisms of biology.However,the diploidy of mammalian somatic cells has greatly hindered efforts to elucidate the gene function in numerous biological processes by mutagenesis-based genetic approaches.Recently,mouse haploid embryonic stem(haES)cells have been successfully isolated from parthenogenetic and androgenetic embryos,providing an ideal tool for genetic analyses.In these studies,mouse haES cells have already shown that they could be used in cell-based forward or reverse genetic screenings and in generating gene-targeting via homologous recombination.In particular,haES cells from androgenetic embryos can be employed as novel,renewable form of fertilization agent for yielding live-born mice via injection into oocytes,thus showing the possibility that genetic analysis can be extended from cellular level to organism level.

Epigenetic Programming： The Challenge to Species Hybridization

In many organisms, the genomes of individual species are isolated by a range of reproductive barriers that act before or after fertilization. Successful mating between species results in the presence of different genomes within a cell （hybridization）, which can lead to incompatibility in cellular events due to adverse genetic interactions. In addition to such genetic interactions, recent studies have shown that the epigenetic control of the genome, silencing of transposons, control of non-additive gene expression and genomic imprinting might also contribute to reproductive barriers in plant and animal species. These genetic and epigenetic mechanisms play a significant role in the prevention of gene flow between species. In this review, we focus on aspects of epigenetic control related to hybrid incompatibility during species hybrid- ization, and also consider key mechanism（s） in the interaction between different genomes.

Dynam,c and Antagonistic Allele-Specific Epigenetic Modifications Controlling the Expression of Imprinted Genes in Maize Endosperm

Genomic imprinting is often associated with aUele-specific epigenetic modifications. Although many reports suggested potential roles of DNA methylation and H3K27me3 in regulating genomic imprinting, the contribu- tions of allele-specific active histone modifications to imprinting remain still unclear in plants. Here, we report the identification of 337 high-stringency allele-specific H3K4me3 and H3K36me3 peaks in maize endosperm. Paternally preferred H3K4me3 and H3K36me3 peaks mostly co-localized with paternally expressed genes （PEGs）, while endosperm-specific maternally expressed genes （endo-MEGs） were associated with mater- nally preferred H3K4me3 and H3K36me3 peaks. A unique signature for PEGs was observed, where the active H3K4me4 and H3K36me3 as well as repressive H3K27me3 appeared together. At the gene body of con-PEGs （constitutively expressed PEG）, H3K27me3 and H3K36me3 were specifically deposited on hypomethylated maternal alleles and hypermethylated paternal alleles, respectively. Around the transcription start sites of endo-MEGs, DNA methylation and H3K4me3 specifically marked paternal and maternal alleles, respectively. In addition, 35 maternally expressed non-coding RNAs exhibited the same allele-specific epigenetic features as endo-MEGs, indicating similar mechanisms for the regulation of imprinted genes and non-coding RNAs. Taken together, our results uncover the complex patterns of mutually exclusive epigenetic modifications deposited at different alleles of imprinted genes that are required for genomic imprinting in maize endosperm.

Dissection of gene function at clonal level using mosaic analysis with double markers

MADM （Mosaic Analysis with Double Markers） technology offers a genetic approach in mice to visualize and concomitantly manipulate genetically defined cells at clonal level and single cell resolution. MADM employs Cre recombinase/loxP-dependent interchromosomal mitotic recombination to reconstitute two split marker genes--green GFP and red tdTomato -- and can label sparse clones of homozygous mutant cells in one color and wild-type cells in the other color in an otherwise unlabeled background. At present, major MADM applications include lineage tracing, single cell labeling, conditional knockouts in small populations of cells and induction of uniparental chromosome disomy to assess effects of genomic imprinting. MADM can be applied universally in the mouse with the sole limitation being the specificity of the promoter controlling Cre recombinase expression. Here I review recent developments and extensions of the MADM technique and give an overview of the major discoveries and progresses enabled by the implementation of the novel genetic MADM tools.