Absence of nucleolar dominance in mouse-human heterokaryons

Autoradiographic analysis of [³H]uridine incorporation 48 h after polyethylene glycol-mediated cell fusion indicates that nucleolar RNA synthesis persists in both human and mouse nuclei in interspecific heterokaryons. The absence of nucleolar dominance in heterokaryons has been confirmed by zinc-dithizone nucleolus-specific staining, and is true even when there are considerably more nuclei of one species than of the other in the heterokaryon. Studies of actinomycin D-induced nucleolar segregation indicate that the zinc-binding proteins responsible for zinc-dithizone staining are located in a different nucleolar component than the protein responsible for silver staining.     

Locus-specific ribosomal RNA gene silencing in nucleolar dominance

The silencing of one parental set of rRNA genes in a genetic hybrid is an epigenetic phenomenon known as nucleolar dominance. We showed previously that silencing is restricted to the nucleolus organizer regions (NORs), the loci where rRNA genes are tandemly arrayed, and does not spread to or from neighboring protein-coding genes. One hypothesis is that nucleolar dominance is the net result of hundreds of silencing events acting one rRNA gene at a time. A prediction of this hypothesis is that rRNA gene silencing should occur independent of chromosomal location. An alternative hypothesis is that the regulatory unit in nucleolar dominance is the NOR, rather than each individual rRNA gene, in which case NOR localization may be essential for rRNA gene silencing. To test these alternative hypotheses, we examined the fates of rRNA transgenes integrated at ectopic locations. The transgenes were accurately transcribed in all independent transgenic Arabidopsis thaliana lines tested, indicating that NOR localization is not required for rRNA gene expression. Upon crossing the transgenic A. thaliana lines as ovule parents with A. lyrata to form F1 hybrids, a new system for the study of nucleolar dominance, the endogenous rRNA genes located within the A. thaliana NORs are silenced. However, rRNA transgenes escaped silencing in multiple independent hybrids. Collectively, our data suggest that rRNA gene activation can occur in a gene-autonomous fashion, independent of chromosomal location, whereas rRNA gene silencing in nucleolar dominance is locus-dependent.     

Cytosine methylation and nucleolar dominance in cereal hybrids

In wheat-rye hybrids the nucleolus organizer regions (NORs), the sites of ribosomal RNA genes, from rye are suppressed. Wheat and wheat-rye hybrid genetic stocks containing different numbers of wheat and rye nucleolus organizers, as well as addition lines and rye-barley hybrids, were used in Southern hybridization experiments to determine the cause of nucleolar dominance and suppression in cereal hybrids. Based on the use of restriction endonucleases that cleave near the ends of the spacer unit and an additional, methylation-sensitive enzyme, HpaII, which does not recognize the CCGG restriction site if the internal C is methylated, an indirect method of assaying NOR expression was established. The results indicated that cleavage by the HpaII enzyme of the rye NOR sequences, is reduced when major NORs from other cereals were present. The reduction in the number of rye rRNA genes containing an unmethylated CCGG site in the promoter was associated with the suppression of the rye nucleolus. These results are consistent with a model in which promoter and upstream regulatory repeats of ribosomal RNA genes compete for limited concentrations of regulatory proteins, and genes that are methylated at key binding sites fail to engage these regulatory proteins and thus remain inactive. Received: 15 November 1996 / Accepted: 19 March 1997     

Postembryonic establishment of megabase-scale gene silencing in nucleolar dominance

Nucleolar dominance is an epigenetic phenomenon in plant and animal genetic hybrids that describes the expression of 45S ribosomal RNA genes (rRNA genes) inherited from only one progenitor due to the silencing of the other progenitor's rRNA genes. rRNA genes are tandemly arrayed at nucleolus organizer regions (NORs) that span millions of basepairs, thus gene silencing in nucleolar dominance occurs on a scale second only to X-chromosome inactivation in female mammals. In Arabidopsis suecica, the allotetraploid hybrid of A. thaliana and A. arenosa, the A. thaliana -derived rRNA genes are subjected to nucleolar dominance and are silenced via repressive chromatin modifications. However, the developmental stage at which nucleolar dominance is established in A. suecica is currently unknown. We show that nucleolar dominance is not apparent in seedling cotyledons formed during embryogenesis but becomes progressively established during early postembryonic development in tissues derived from both the shoot and root apical meristems. The progressive silencing of A. thaliana rRNA genes correlates with the transition of A. thaliana NORs from a decondensed euchromatic state associated with histone H3 that is trimethylated on lysine 4 (H3K4me3) to a highly condensed heterochromatic state in which the NORs are associated with H3K9me2 and 5-methylcytosine-enriched chromocenters. In RNAi-lines in which the histone deacetylases HDA6 and HDT1 are knocked down, the developmentally regulated condensation and inactivation of A. thaliana NORs is disrupted. Collectively, these data demonstrate that HDA6 and HDT1 function in the postembryonic establishment of nucleolar dominance, a process which recurs in each generation.     

The epigenetics of estrogen

Epigenetic processes have been impli- zcated in everything from cell proliferation to maternal behavior. Epigenetic alterations, including histone alterations and DNA methylation, have also been shown to play critical roles in the formation of some types of memory, and in the modulatory effects that factors, such as stress, drugs of abuse and environmental stimulation, have on the brain and memory function. Recently, we demonstrated that the ability of the sex-steroid hormone 17β-estradiol (E₂) to enhance memory formation is dependent on histone acetylation and DNA methylation, a finding that has important implications for understanding how hormones influence cognition in adulthood and aging. In this article, we provide an overview of the literature demonstrating that epigenetic processes and E₂ influence memory, describe our findings indicating that epigenetic alterations regulate E₂-induced memory enhancement, and discuss directions for future work on the epigenetics of estrogen.     

New evidence for nucleolar dominance in hybrids of Drosophila arizonae and Drosophila mulleri

Drosophila mulleri (MU) and D. arizonae (AR) are cryptic species of the mulleri complex, mulleri subgroup, repleta group. Earlier cytogenetic studies revealed that these species have different regulatory mechanisms of nucleolar organizing activity. In these species, nucleolar organizing regions are found in both the X chromosome and the microchromosome. In the salivary glands of hybrids between MU females and AR males, there is an interspecific dominance of the regulatory system of the D. arizonae nucleolar organizer involving, in males, amplification and activation of the nucleolar organizer from the microchromosome. The authors who reported these findings obtained hybrids only in that cross-direction. More recently, hybrids in the opposite direction, i.e., between MU males and AR females, have been obtained. The purpose of the present study was to evaluate, in these hybrids, the association of the nucleoli with the chromosomes inherited from parental species in order to cytogenetically confirm the dominance patterns previously described. Our results support the proposed dominance of the AR nucleolar organizer activity over that of MU, regardless of cross-direction.     

The epigenetics of CHARGE syndrome

In biology, we continue to appreciate the fact that the DNA sequence alone falls short when attempting to explain the intricate inheritance patterns for complex traits. This is particularly true for human disorders that appear to have simple genetic causes. The study of epigenetics, and the increased access to the epigenetic profiles of different tissues has begun to shed light on the genetic complexity of many basic biological processes, both physiological and pathological. Epigenetics refers to heritable changes in gene expression that are not due to alterations in the DNA sequence. Various mechanisms of epigenetic regulation exist, including DNA methylation and histone modification. The identification, and increased understanding of key players and mechanisms of epigenetic regulation have begun to provide significant insight into the underlying origins of various human genetic disorders. One such disorder is CHARGE syndrome (OMIM #214800), which is a leading cause of deaf-blindness worldwide. A majority of CHARGE syndrome cases are caused by haploinsufficiency for the CHD7 gene, which encodes an ATP-dependent chromatin remodeling protein involved in the epigenetic regulation of gene expression. The CHD7 protein has been highly conserved throughout evolution, and research into the function of CHD7 homologs in multiple model systems has increased our understanding of this family of proteins, and epigenetic mechanisms in general. Here we provide a review of CHARGE syndrome, and discuss the epigenetic functions of CHD7 in humans and CHD7 homologs in model organisms.     

The epigenetics of the cell

A report on the 42nd Annual Meeting of the American Society for Cell Biology, San Francisco, 14-18 December 2002.     

The nucleolar detention pathway

Molecular dynamics ensures that proteins and other factors reach their site of action in a timely and efficient manner. This is essential to the formation of molecular complexes, as they require an ever-changing framework of specific interactions to facilitate a model of self-assembly. Therefore, the absence or reduced availability of any key component would significantly impair complex formation and disrupt all downstream molecular networks. Recently, we identified a regulatory mechanism that modulates protein mobility through the inducible expression of a novel family of long noncoding RNA. In response to diverse environmental stimuli, the nucleolar detention pathway (NoDP) captures and immobilizes essential cellular factors within the nucleolus away from their effector molecules. The vast array of putative NoDP targets, including DNA (cytosine-5)-methyltransferase 1 (DNMT1) and the delta catalytic subunit of DNA polymerase (POLD1), suggests that this may be a common and significant regulatory mechanism. Here, we discuss the implications of this new posttranslational strategy for regulating molecular networks.     

A concerted DNA methylation/histone methylation switch regulates rRNA gene dosage control and nucleolar dominance

Eukaryotes regulate the effective dosage of their ribosomal RNA (rRNA) genes, expressing fewer than half of the genes at any one time. Likewise, genetic hybrids displaying nucleolar dominance transcribe rRNA genes inherited from one parent but silence the other parental set. We show that rRNA gene dosage control and nucleolar dominance utilize a common mechanism. Central to the mechanism is an epigenetic switch in which concerted changes in promoter cytosine methylation density and specific histone modifications dictate the on and off states of the rRNA genes. A key component of the off switch is HDT1, a plant-specific histone deacetylase that localizes to the nucleolus and is required for H3 lysine 9 deacetylation and subsequent H3 lysine 9 methylation. Collectively, the data support a model in which cytosine methylation and histone deacetylation are each upstream of one another in a self-reinforcing repression cycle.