mRNA expression and DNA methylation in three key genes involved in caste differentiation in female honeybees(Apis mellifera)

In honeybee(Apis mellifera)colonies,queens and workers are alternative forms of the adult female honeybee that develop from genetically identical zygotes but that depend on differential nourishment.Queens and workers display distinct morphologies,anatomies and behavior,better known as caste differentiation.Despite some basic insights,the exact mechanism responsible for this phenomenon,especially at the molecular level,remains unclear although some progress has been achieved.In this study,we examined mRNA levels of the TOR(target of rapamycin)and Dnmt3(DNA methyltransferase 3)genes,closely related to caste differentiation in honeybees.We also investigated mRNA expression of the S6K(similar to RPS6-p70-protein kinase)gene linked closely to organismal growth and development in queen and worker larvae(1-day and 3-day old).Last,we investigated the methylation status of these three genes in corresponding castes.We found no difference in mRNA expression for the three genes between 1st instar queen and worker larvae;however,3rd instar queen larvae had a higher level of TOR mRNA than worker larvae.Methylation levels of all three genes were lower in queen larvae than worker larvae but the differences were not statistically significant.These findings provide basic data for broadening our understanding of caste differentiation in female honeybees.     

熊蜂级型分化研究进展

熊蜂是众多野生植物及农作物的有效授粉昆虫,具有重要的经济和生态价值。熊蜂复杂的生长发育过程及社会性使其成为社会生物学研究的最佳模式生物之一。社会性昆虫的生殖劳动分工具有重要的进化意义,而级型分化是形成生殖劳动分工的基础。蜜蜂级型分化的研究已取得诸多重要成果,其机理也得到了较为深入的阐释,而熊蜂的社会性研究尚未形成系统,与蜜蜂研究相差甚远。近来的研究表明,饲喂频率或者饲喂总量的差异能够引起熊蜂级型分化的发生。保幼激素和蜕皮激素与熊蜂幼虫的发育紧密联系,在熊蜂级型分化的过程中发挥重要作用。一些参与蜜蜂级型分化的基因,在熊蜂级型间也存在差异表达。此外,群体间的相互作用以及蜂王和工蜂间的竞争也是促进熊蜂级型分化发生的重要因素。本文从营养、激素调控、群体发展及相互作用等方面综述熊蜂级型分化机制,并对未来的研究提出可能方向。     

Pre-mRNA选择性剪接的调控及选择性剪接数据库

Pre-mRNA选择性剪接是真核生物转录组和蛋白质组多样性的主要来源,也是细胞分化、发育等过程中重要的基因表达调控方式。约95%的人类多外显子基因存在RNA选择性剪接|很多人类基因疾病的发生与RNA剪接错误相关。随着共转录现象的发现,RNA选择性剪接调控机制研究也取得了很大进展。本文分别从序列层面和核小体定位、组蛋白修饰、DNA甲基化及非编码RNA等表观遗传层面,系统地阐述了RNA选择性剪接的调控机制。为便于搜索,本文介绍了近10年来RNA选择性剪接相关的数据库。     

Patterns of DNA methylation in development, division of labor and hybridization in an ant with genetic caste determination

Background

DNA methylation is a common regulator of gene expression, including acting as a regulator of developmental events and behavioral changes in adults. Using the unique system of genetic caste determination in Pogonomyrmex barbatus, we were able to document changes in DNA methylation during development, and also across both ancient and contemporary hybridization events.

Methodology/Principal Findings

Sodium bisulfite sequencing demonstrated in vivo methylation of symmetric CG dinucleotides in P. barbatus. We also found methylation of non-CpG sequences. This validated two bioinformatics methods for predicting gene methylation, the bias in observed to expected ratio of CpG dinucleotides and the density of CpG/TpG single nucleotide polymorphisms (SNP). Frequencies of genomic DNA methylation were determined for different developmental stages and castes using ms-AFLP assays. The genetic caste determination system (GCD) is probably the product of an ancestral hybridization event between P. barbatus and P. rugosus. Two lineages obligately co-occur within a GCD population, and queens are derived from intra-lineage matings whereas workers are produced from inter-lineage matings. Relative DNA methylation levels of queens and workers from GCD lineages (contemporary hybrids) were not significantly different until adulthood. Virgin queens had significantly higher relative levels of DNA methylation compared to workers. Worker DNA methylation did not vary among developmental stages within each lineage, but was significantly different between the currently hybridizing lineages. Finally, workers of the two genetic caste determination lineages had half as many methylated cytosines as workers from the putative parental species, which have environmental caste determination.

Conclusions/Significance

These results suggest that DNA methylation may be a conserved regulatory mechanism moderating division of labor in both bees and ants. Current and historic hybridization appear to have altered genomic methylation levels suggesting a possible link between changes in overall DNA methylation and the origin and regulation of genetic caste determination in P. barbatus.     

Molecular Profiling of DNA Methylation and Alternative Splicing of Genes in Skeletal Muscle of Obese Rabbits

DNA methylation and the alternative splicing of precursor messenger RNAs (pre-mRNAs) are two important genetic modification mechanisms. However, both are currently uncharacterized in the muscle metabolism of rabbits. Thus, we constructed the Tianfu black rabbit obesity model (obese rabbits fed with a 10% high-fat diet and control rabbits from 35 days to 70 days) and collected the skeletal muscle samples from the two groups for Genome methylation sequencing and RNA sequencing. DNA methylation data showed that the promoter regions of 599 genes and gene body region of 2522 genes had significantly differential methylation rates between the two groups, of which 288 genes had differential methylation rates in promoter and gene body regions. Analysis of alternative splicing showed 555 genes involved in exon skipping (ES) patterns, and 15 genes existed in differential methylation regions. Network analysis showed that 20 hub genes were associated with ubiquitinated protein degradation, muscle development pathways, and skeletal muscle energy metabolism. Our findings suggest that the two types of genetic modification have potential regulatory effects on skeletal muscle development and provide a basis for further mechanistic studies in the rabbit.     

长链非编码RNA的免疫调节机制研究进展

真核生物的基因表达受多个层面调控,包括染色体水平、DNA水平、转录水平和转录后水平的调控等.长链非编码RNA(lnc RNA)是一类转录本超过200 nt的非编码RNA,其对基因表达的调控涉及上述各个层面,如组蛋白修饰、DNA甲基化的调控、转录的促进和抑制、m RNA的剪辑及对转录因子的调控等.其作用方式复杂多样,可与DNA、mRNA和蛋白质等相互作用而发挥调节作用.LncRNA保守性较差,但其表达却有较高的细胞、组织和分化阶段特异性.免疫系统的发育和分化受到精密的调控,且具有较高的阶段性和特异性.因此研究lnc RNA的功能及作用机制,免疫系统是较好的选择,这能促进我们对免疫调控的理解,为免疫性疾病的治疗提供新的思路和方法.本文主要介绍lnc RNA的分类和lnc RNA作用的一般分子机制,及其对T细胞、B细胞、固有免疫细胞和炎症因子的分子调控机制及其进展.     

Transcriptomic analysis of epigenetic modification genes in the termite Reticulitermes speratus

Eusocial insects display a caste system in which different castes are morpho-logically and physiologically specialized for different tasks.Recent studies have revealed that epigenetic modifications,including DNA methylation and histone modification,me-diate caste determination and differentiation,longevity,and polyethism in eusocial insects.Although there has been a growing interest in the relationship between epigenetic mech-anisms and phenotypic plasticity in termites,there is ltte information about differential expression levels among castes and expression sites for these genes in termites.Here we show royaltissuc-specific expression of epigenetic modification genes in the termite Reticulitermes speratus.Using RNA-seq,we identified 74 genes,including three DNA methyltransferases,seven sirtuins,48 Trithorax group proteins,and 16 Polycomb group proteins.Among these genes,15 showed king-specific expression,and 52 showed age-dependent differential expression in kings and queens.Quantitative real-time PCR revealed that DNA methyltransferase 3 is expressed specifically in the king's testis and fat body,whereas some histone modification genes are remarkably expressed in the king's testis and queen's ovary.These findings imply that epigenetic modification plays important roles in the gamete production process in termite kings and queens.     

Integrated epigenome profiling of repressive histone modifications, DNA methylation and gene expression in normal and malignant urothelial cells

Epigenetic regulation of gene expression is commonly altered in human cancer. We have observed alterations of DNA methylation and microRNA expression that reflect the biology of bladder cancer. This common disease arises by distinct pathways with low and high-grade differentiation. We hypothesized that epigenetic gene regulation reflects an interaction between histone and DNA modifications, and differences between normal and malignant urothelial cells represent carcinogenic events within bladder cancer. To test this we profiled two repressive histone modifications (H3K9m3 and H3K27m3) using ChIP-Seq, cytosine methylation using MeDIP and mRNA expression in normal and malignant urothelial cell lines. In genes with low expression we identified H3K27m3 and DNA methylation each in 20-30% of genes and both marks in 5% of genes. H3K9m3 was detected in 5-10% of genes but was not associated with overall expression. DNA methylation was more closely related to gene expression in malignant than normal cells. H3K27m3 was the epigenetic mark most specifically correlated to gene silencing. Our data suggest that urothelial carcinogenesis is accompanied by a loss of control of both DNA methylation and H3k27 methylation. From our observations we identified a panel of genes with cancer specific-epigenetic mediated aberrant expression including those with reported carcinogenic functions and members potentially mediating a positive epigenetic feedback loop. Pathway enrichment analysis revealed genes marked by H3K9m3 were involved with cell homeostasis, those marked by H3K27m3 mediated pro-carcinogenic processes and those marked with cytosine methylation were mixed in function. In 150 normal and malignant urothelial samples, our gene panel correctly estimated expression in 65% of its members. Hierarchical clustering revealed that this gene panel stratified samples according to the presence and phenotype of bladder cancer.     

The effects of culture on genomic imprinting profiles in human embryonic and fetal mesenchymal stem cells

Human embryonic stem (hES) cells and fetal mesenchymal stem cells (fMSC) offer great potential for regenerative therapy strategies. It is therefore important to characterize the properties of these cells in vitro. One major way the environment impacts on cellular physiology is through changes to epigenetic mechanisms. Genes subject to epigenetic regulation via genomic imprinting have been characterized extensively. The integrity of imprinted gene expression therefore provides a measurable index for epigenetic stability. Allelic expression of 26 imprinted genes and DNA methylation at associated differentially methylated regions (DMRs) was measured in fMSC and hES cell lines. Both cell types exhibited monoallelic expression of 13 imprinted genes, biallelic expression of six imprinted genes, and there were seven genes that differed in allelic expression between cell lines. fMSC s exhibited the differential DNA methylation patterns associated with imprinted expression. This was unexpected given that gene expression of several imprinted genes was biallelic. However, in hES cells, differential methylation was perturbed. These atypical methylation patterns did not correlate with allelic expression. Our results suggest that regardless of stem cell origin, in vitro culture affects the integrity of imprinted gene expression in human cells. We identify biallelic and variably expressed genes that may inform on overall epigenetic stability. As differential methylation did not correlate with imprinted expression changes we propose that other epigenetic effectors are adversely influenced by the in vitro environment. Since DMR integrity was maintained in fMSC but not hES cells, we postulate that specific hES cell derivation and culturing practices result in changes in methylation at DMRs.Key words: genomic imprinting, embryonic stem cells, mesenchymal stem cells, differentiation, methylation, epigenetic stability     

Evolution of gene expression in fire ants: the effects of developmental stage, caste, and species

Ants provide remarkable examples of equivalent genotypes developing into divergent and discrete phenotypes. Diploid eggs can develop either into queens, which specialize in reproduction, or workers, which participate in cooperative tasks such as building the nest, collecting food, and rearing the young. In contrast, the differentiation between males and females generally depends upon whether eggs are fertilized, with fertilized (diploid) eggs giving rise to females and unfertilized (haploid) eggs giving rise to males. To obtain a comprehensive picture of the relative contributions of gender (sex), caste, developmental stage, and species divergence to gene expression evolution, we investigated gene expression patterns in pupal and adult queens, workers, and males of two species of fire ants, Solenopsis invicta and S. richteri. Microarray hybridizations revealed that variation in gene expression profiles is influenced more by developmental stage than by caste membership, sex, or species identity. The second major contributor to variation in gene expression was the combination of sex and caste. Although workers and queens share equivalent diploid nuclear genomes, they have highly distinctive patterns of gene expression in both the pupal and the adult stages, as might be expected given their extraordinary level of phenotypic differentiation. Overall, the difference in the proportion of differentially expressed genes was greater between workers and males than between workers and queens or queens and males, consistent with the fact that workers and males share neither gender nor reproductive capability. Moreover, between-species comparisons revealed that the greatest difference in gene expression patterns occurred in adult workers, a finding consistent with the fact that adult workers most directly experience the distinct external environments characterizing the different habitats occupied by the two species. Thus, much of the evolution of gene expression in ants may occur in the worker caste, despite the fact that these individuals are largely or completely sterile. Analyses of gene expression evolution revealed a combination of positive selection and relaxation of stabilizing selection as important factors driving the evolution of such genes.