表观调控rDNA转录的研究进展

rDNA是控制细胞核糖体生物合成的串联重复基因,影响着整体蛋白质的翻译水平,与细胞生长代谢息息相关.由于rDNA序列具有多拷贝的重复特征,其转录除了受一般转录机制的调节外,还受多重表观调控机制的调节,精细调控着rDNA的转录状态.一般认为rDNA分为活跃和沉默两种状态,分别与活跃染色质标记和异染色质标记相关.近些年来,发现一种平衡态rDNA的存在,更加丰富了rDNA表观机制的研究.H3.3是一种H3组蛋白变体,是近些年来的研究热点,已有报道H3.3可能在分子伴侣HIRA的介导下整合进入活跃rDNA,然而沉默rDNA的维持是否也与H3.3的作用相关需要更多的探索.CTCF是rDNA重复单元间的绝缘子成分,与H3.3相关但并不清楚是否也调控着rDNA的转录.该综述讨论了几种调控rDNA表观状态的机制,并对可能参与该过程的新机制提出了设想.     

Epigenetic control of cardiomyocyte production in response to a stress during the medaka heart development

The size and morphology of organs are largely determined by a genetic program. However in some cases, an epigenetic mechanism influences the process of organ development. Particularly, epigenetic factors such as hemodynamic stress and blood pressure affect the morphogenesis of cardiac chambers and valves. Here, we report that the epigenetic influences affect the cardiomyocyte production. Taking advantage of longer developmental period of medaka fish, we could examine the later emerging tissue responses to the defect of ventricular beating, which occurred in the hozuki (hoz) mutant that harbors the mutated ventricular myosin heavy chain (vmhc) gene. The mutant showed a remarkable ventricular enlargement, and we showed that this enlargement was due to an excess production of ventricular cardiomyocytes in addition to the lack of concentric chamber growth. By experimental blockade of blood flow, we demonstrated that an elevated cardiac pressure was responsible for the aberrant cardiomyocyte production. From these data, we propose that the epigenetic tissue response to a stressed situation controls the production of cardiomyocytes to attain a fine tuning of heart formation.     

The simultaneous determination of intracellular pH and cell energy status

The relationship between the apparent equilibrium constant of creatine kinase and intracellular pH was evaluated in CHO and murine FSaII tumor cells. The apparent equilibrium constant, K' = [ATP][Cr]/[ADP][PCr], was determined from acid extracts at variable pH. Intracellular pH (pHi) was determined from the intracellular/extracellular distribution of the weak acid 5,5-dimethyl-2,4-oxazolidinedione. Over the intracellular pH range of 7.2 to 6.1, K' increased by a factor of approximately 10. Intracellular pH was related to the apparent equilibrium constant by the equation pHi = -log K' + log K, where the value of the constant log (log[K'/H+]) was 8.09. Over the same pH range, the concentration of phosphocreatine decreased with pH. Essentially identical results were obtained in CHO and FSaII tumor cells. The similar apparent equilibrium constants in CHO and FSaII cells suggest that assessment of the creatine kinase metabolites will be useful not only for determination of cell energy status but also for the determination of intracellular pH. This information may be useful for the design of therapeutic strategies which are influenced by pH or energy status such as hyperthermia, and drugs which are weak acids or bases, including hypoxic cell radiosensitizers.     

Epigenetic heterogeneity at imprinted loci in normal populations

Genomic imprinting is the phenomenon by which the two alleles of certain genes are differentially expressed according to their parental origin. Extensive analysis of allelic expression at multiple imprinted loci in a normal population has not performed so far. In the present study, we examined the allelic expression pattern of three imprinted genes in a panel of 262 Japanese normal individuals. We observed differences in the extent of maintenance of allele-specific expression of the three genes. The allelic expression of small nuclear ribonucleoprotein N (SNRPN) was stringently regulated while that of multimembrane-spanning polyspecific transporter-like gene 1 (IMPT1) showed a large degree of variation. Significant biallelic expression of insulin-like growth factor II (IGF2) was observed in about 10% of normal individuals. Our findings add to the accumulating evidence for variable allelic expression at multiple loci in a normal human population. This epigenetic heterogeneity can be a stable trait and potentially influence individual phenotypes.     

Physical mapping of rDNA loci in Brassica species.

The number of major rDNA loci (the genes coding for 18S-5.8S-26S rRNA) was investigated in the economically important Brassica species and their wild relatives by in situ hybridization of an rDNA probe to metaphase chromosomes and interphase nuclei. The diploid species B. nigra (B genome) has two major pairs of rDNA loci, B. oleracea (C genome) has two major pairs and one minor pair of loci, while B. campestris (A genome) has five pairs of loci. Among the three tetraploid species arising from these three diploid ancestors, B. carinata (BBCC genomes) has four loci, B. juncea (AABB genomes) has five major pairs and one minor pair of loci, and B. napus (AACC genomes) has six pairs of loci, indicating that the number of loci has been reduced during evolution. The complexity of the known rDNA restriction fragment length polymorphism patterns gave little indication of number of rDNA loci. It is probable that chromosome rearrangements have occurred during evolution of the amphidiploid species. The data will be useful for physical mapping of genes relative to rDNA loci, micro- and macro-evolutionary studies and analysis of aneuploids including addition and substitution lines used in Brassica breeding programs.     

细胞自噬发生的表观遗传调节

细胞自噬是一种进化上保守的, 通过吞噬降解自身大分子物质或细胞器来维持细胞生存的活动。自噬与多种生命活动息息相关, 其功能的紊乱往往会导致肿瘤发生、神经退行性疾病、微生物感染等疾病。研究表明, 表观遗传修饰可以调控细胞自噬的发生, 并在细胞自噬的生物学功能调节过程中发挥重要作用, 但具体调控机制尚需进一步探究。文章综述了细胞自噬发生过程中存在的表观遗传效应, 包括组蛋白乙酰化对细胞自噬激活或抑制的负反馈调控, 通过DNA甲基化调节自噬相关基因活性来影响细胞自噬的发生, miRNA通过靶向调节自噬相关基因表达来影响组蛋白修饰, 从而调控细胞自噬的发生及作用过程等, 旨在为人们进一步研究细胞自噬发生过程中的表观遗传修饰及其机制提供信息依据。     

Influence of extracellular pH on intracellular pH and cell energy status: relationship to hyperthermic sensitivity

The effect of variable extracellular pH on intracellular pH, cell energy status, and thermal sensitivity was evaluated in CHO cells over the extracellular pH range of 6.0 to 8.6. Extracellular pH was adjusted with either lactic acid, HCl, or NaOH. Regardless of the method of pH adjustment, the results obtained were similar. The relationship between extracellular and intracellular pH was dependent upon the pH range examined. Intracellular pH was relatively resistant to a change in extracellular pH over the pHe range of 6.8 to 7.8 (i.e., delta pHi congruent to delta pHe X 0.33). Above and below this range, delta pHi congruent to delta pHe X 1.08 or X 0.76, respectively. Cellular survival after a 30-min heat treatment at 44 degrees C remained constant over the extracellular pH range of 7.0 to 8.4, but varied substantially over a similar intracellular pH range. The cellular concentration of the high energy phosphate reservoir, phosphocreatine, decreased with decreasing pH. However, the cellular concentrations of ATP, ADP, and AMP remained constant over the entire pH range examined. It is concluded that increased thermal sensitivity resulting from a change in extracellular pH is not due to cellular energy depletion. Furthermore, intracellular pH is a more accurate indicator of thermal sensitivity than is extracellular pH.     

Distribution of rDNA loci in the genus Glycine Willd.

The objective of this study was to examine the distribution of rDNA loci in the genus Glycine Willd. by fluorescent in situ hybridization (FISH) using the internal transcribed spacer (ITS) region of nuclear ribosomal DNA as a probe. The hybridized rDNA probe produced two distinct yellow signals on reddish chromosomes representing two NORs in 16 diploid (2n=40) species. Aneudiploid (2n=38) and aneutetraploid (2n=78) Glycine tomentella Hayata also exhibited two rDNA sites. However, the probe hybridized with four chromosomes as evidenced by four signals in two diploid species (Glycine curvata Tind. and Glycine cyrtoloba Tind.) and tetraploid (2n=80) G. tabacina (Labill.) Benth. and G. tomentella. Synthesized amphiploids (2n=80) of Glycine canescens F. J. Herm. (2n=40) and the 40-chromosome G. tomentella also showed four signals. This study demonstrates that the distribution of the rDNA gene in the 16 Glycine species studied is highly conserved and that silence of the rDNA locus may be attributed to amphiplasty during diploidization and speciation. Received: 10 October 2000 / Accepted: 6 December 2000     

Epigenetic regulation of the nitrosative stress response and intracellular macrophage survival by extraintestinal pathogenic Escherichia coli

Extraintestinal pathogenic Escherichia coli (ExPEC) reside in the enteric tract as a commensal reservoir, but can transition to a pathogenic state by invading normally sterile niches, establishing infection and disseminating to invasive sites like the bloodstream. Macrophages are required for ExPEC dissemination, suggesting the pathogen has developed mechanisms to persist within professional phagocytes. Here, we report that FimX, an ExPEC-associated DNA invertase that regulates the major virulence factor type 1 pili (T1P), is also an epigenetic regulator of a LuxR-like response regulator HyxR. FimX regulated hyxR expression through bidirectional phase inversion of its promoter region at sites different from the type 1 pili promoter and independent of integration host factor (IHF). In vitro, transition from high to low HyxR expression produced enhanced tolerance of reactive nitrogen intermediates (RNIs), primarily through de-repression of hmpA, encoding a nitric oxide-detoxifying flavohaemoglobin. However, in the macrophage, HyxR produced large effects on intracellular survival in the presence and absence of RNI and independent of Hmp. Collectively, we have shown that the ability of ExPEC to survive in macrophages is contingent upon the proper transition from high to low HyxR expression through epigenetic regulatory control by FimX.     

Epigenetic inheritance of cell differentiation status

Epigenetic modifications influence gene expression pattern and provide a unique signature of a cell differentiation status. Without external stimuli or signalling events, this cell identity remains stable and unlikely to change over many cell divisions. The epigenetic signature of a particular cell fate therefore needs to be replicated faithfully in daughter cells; otherwise a cell lineage cannot be maintained. However, the mechanism of transmission of cellular memory from mother to daughter cells remains unclear. It has been suggested that the inheritance of an active or silent gene state involves different kinds of epigenetic mechanisms, e.g. DNA methylation, histone modifications, replacement of histone variants, Polycomb group (PcG) and Trithorax group (TrxG) proteins. Emerging evidence supports the role of histone variant H3.3 in maintaining an active gene status and in remodelling nucleosomal composition. Here we discuss some recent findings on the propagation of epigenetic memory and propose a model for the inheritance of an active gene state through the interaction of H3.3 with other epigenetic components.     

Extracellular and intracellular acid-base status with regard to the energy metabolism in the oyster Crassostrea gigas during exposure to air

The acid-base status of extra- and intracellular fluids was studied in relation to the anaerobic energy metabolism in the adductor muscle, mantle, gills, and heart of the marine bivalve Crassostrea gigas after exposure to air for periods of 2, 4, 8, 12, 24, and 48 h. Such exposure was found to cause a significant reduction in the pH in the hemolymph (pH(e)) within the first 4 h. The decrease in the pHe was accompanied by elevated Pco2 values, causing [HCO3-] to rise (respiratory acidosis). Thereafter, the pHe fell at a lower rate, and this fall was partially compensated for by a further increase in [HCO3-] in the hemolymph. The increase in the [Ca] levels in the hemolymph indicates a mobilization of Ca2+ from CaCO3 and the involvement of bicarbonates in the buffering of pHe. The main anaerobic end-products that accumulated in the tissues during the first stages of anaerobiosis were alanine and succinate, at a ratio of about 2 : 1. Later on, propionate and acetate were also accumulated at significant rates. In contrast to the adductor muscle, gills, and mantle, opine production in the heart was significant after 12-24 h of exposure to air. Determination of intracellular pH (pHi) revealed that there is a close relationship between the rate of anaerobic end-product accumulation and the extent of intracellular acidosis in the adductor muscle, mantle, and gills. On the contrary, accumulation of anaerobic end-products in the heart did not cause any significant change in its pHi. The intracellular nonbicarbonate, nonphosphate buffering value (beta (NB,NPi)) was determined to be higher in the heart than in the other three tissues and thus probably plays a crucial role in stabilizing heart pHi during exposure to air.     

Epigenetic control of plant immunity

In eukaryotic genomes, gene expression and DNA recombination are affected by structural chromatin traits. Chromatin structure is shaped by the activity of enzymes that either introduce covalent modifications in DNA and histone proteins or use energy from ATP to disrupt histone–DNA interactions. The genomic ‘marks’ that are generated by covalent modifications of histones and DNA, or by the deposition of histone variants, are susceptible to being altered in response to stress. Recent evidence has suggested that proteins generating these epigenetic marks play crucial roles in the defence against pathogens. Histone deacetylases are involved in the activation of jasmonic acid‐ and ethylene‐sensitive defence mechanisms. ATP‐dependent chromatin remodellers mediate the constitutive repression of the salicylic acid‐dependent pathway, whereas histone methylation at the WRKY70 gene promoter affects the activation of this pathway. Interestingly, bacterial‐infected tissues show a net reduction in DNA methylation, which may affect the disease resistance genes responsible for the surveillance against pathogens. As some epigenetic marks can be erased or maintained and transmitted to offspring, epigenetic mechanisms may provide plasticity for the dynamic control of emerging pathogens without the generation of genomic lesions.     

Epigenetic control of reproductive development

Epigenetics is a rapidly expanding research field focused on gene regulation and differentiation. Due to a sessile life, plants have evolved diverse adaptive mechanisms that are controlled by epigenetic processes. Epigenetic molecular mechanisms are in principle shared by plants and animals. There are two basic stages in plant development: The germline is formed after the vegetative period of life. Similarly, a unique stage in plant life is a haploid gametophyte, a short but important period requiring gene expression. These transitions in plant life are controlled by epigenetic processes such as DNA methylation, nucleosomal histone modifications, and regulation via small RNAs. In this review we summarize epigenetic events in the course of transition from vegetative to generative development, formation of spores and gametes, and fertilization and embryogenesis.     

Autophagosome formation in response to intracellular bacterial invasion

Autophagy is an intracellular bulk degradation system in which double‐membrane vesicles, called autophagosomes, engulf cytoplasmic components and later fuse with lysosomes to degrade the autophagosome content. Although autophagy was initially thought a non‐selective process, recent studies have clarified that it can selectively target intracellular bacteria and function as an intracellular innate immune system that suppresses bacterial survival. A key mechanism for the recognition of cytosol‐invading bacteria is ubiquitination, and the recognition of the ubiquitinated target by the autophagy machinery can be accomplished multiple ways. In this review, we discuss recent findings regarding the induction of autophagosome formation in response to intracellular bacterial invasion.     

TSC2 mediates cellular energy response to control cell growth and survival

Mutations in either the TSC1 or TSC2 tumor suppressor gene are responsible for Tuberous Sclerosis Complex. The gene products of TSC1 and TSC2 form a functional complex and inhibit the phosphorylation of S6K and 4EBP1, two key regulators of translation. Here, we describe that TSC2 is regulated by cellular energy levels and plays an essential role in the cellular energy response pathway. Under energy starvation conditions, the AMP-activated protein kinase (AMPK) phosphorylates TSC2 and enhances its activity. Phosphorylation of TSC2 by AMPK is required for translation regulation and cell size control in response to energy deprivation. Furthermore, TSC2 and its phosphorylation by AMPK protect cells from energy deprivation-induced apoptosis. These observations demonstrate a model where TSC2 functions as a key player in regulation of the common mTOR pathway of protein synthesis, cell growth, and viability in response to cellular energy levels.