The role of PACT in the RNA silencing pathway

Small RNA-mediated gene silencing was recognized as one of the core mechanisms involved in the epigenetic modification of vital process of cell. First discovered in C. elegans in 1995 and reported on Nature magazine in 1998, RNAi was now commonly used as a convenient way of specific gene silence in labs. Further studies indicated the complexity of RNA-mediated regulation system on gene transcription. Researcher nowadays found two major kinds of short RNA, small interfering RNA (siRNA) and microRNA (miRNA), both involved in translational repression by selected mRNA degradation.

But within the functional differences of siRNA and miRNA, multi-domain ribonuclease Dicer and the effector complex, RNA-induced silencing complex, serves as basal participators. In year 2005, Scientists found two factor, hAgo2 and TRBP, participated in the action of RISC on recognition and degradation of mRNA.

Scientists from Korea reported on The EMBO Journal early this year that they found a new dsRNA-binding protein, PATC, which used to be recognized as an activator of protein kinase R and shared similar domain structure with TRBP, interacted with Dicer and hAgo2 in vitro. Further studies using mutants and coimmunoprecipitation show that the third dsRBD of PACT may be responsible for its interaction with Dicer. Similar protocol was used to find the interaction of hAgo2 and PACT. Results indicated that the binding function of PACT with hAgo2 by different domain independently from that with Dicer. In vitro experiment revealed that PACT, TRBP and Dicer associated in the mRNA slicer activity. Results of RNA interference against PACT and TRBP both indicated their participation in RNA silencing pathway, while PACT may perform less than TRBP. Finally, researchers indicated that PACT was involved in accumulation of mature miRNA in HeLa cell line.

From all the results above the author of this article present the fact that human Dicer may associate with two dsRNA-binding proteins, PACT and TRBP. PACT may play a role at RISC assembly, but have no function on pre-miRNA cleavage.

Within this article, the present of PACT with certain roles in a ~500kDa complex provide more knowledge about RNA-associated silencing. This achievement helps us to explore further cross-talking signals and relation of functional factors involved in RNAi, even provide a new field for epigenetic researches and prospects for epigenetic therapy.

Epigenetic influences may explain dental differences in monozygotic twin pairs

As I am monozygotic twins, I am interested in any article about twins.
Specially, I have a missing teeth since I was born. And suprigingly, My twin sister got a teeth on her palate when she was twelve years old. At that time we thought, that teeth was mine and she took a my teeth.

This paper is about dental differences in monozygotic twin pairs and relating to epigenetic.
The aim of this paper is to determine the prevalence of discordant expression for simple hypodontia (missing teeth) and supernumerary teeth (extra teeth) in monozygotic twin pairs and to explain how phenotype differences might occur despite their similar genotypes.

Hope you guys enjoy to reading this article.

Cheers!

Patricia

The Polycomb group protein EZH2 directly controls DNA methylation

The mystery of tumor formation and progression has long plagued scientists and medical experts alike. However a new study into epigenetic gene silencing has uncovered a significant correlation between all stages of tumor formation and progression with Chromatin alterations.

The study demonstrated that epigenetically mediated transcriptional-silencing events that are associated with increases in DNA methylation — particularly at promoter regions of genes that regulate important cell function were generally present in situations of tumor formation and growth.

Genes encoding Ploycomb group(PcG) proteins were first identified in Drosophila as essential factors for the regulation of homeotic genes during development. PcG proteins are divided into two families in accordance with their physical associations in distinct multiprotein complexes termed Polycomb repressor complexes (PRCs). Enhancer of Zeste homolog (EZH2) is a particular protein within the Polycomb group. It is classified as a histone lysine methyltransferase (HKMT) and has been associated with tran-scriptional repression.

In this article, the authors certificated the connection of corresponding silencing pathways. They obtained the result that the PcG protein EZH2 interacts with DNA methyltransferases(DNMTs) and associates with DNA methyltransferase activity in vivo.

Furthermore, the proximate relationship identified between EZH2 and DNMTs might also be relevant to X-chromosome inactivation in female mammals. Consequently the research into the mechanistic link between the two essential epigenetic systems may be of substantial scientific significance.

Thanks for reading :)

Yiwen Cui

Structural Basis for Double-Stranded RNA Processing by Dicer

RNA interference is one of the hotspots in biological studies these days. People keep trying to apply it as a new research tool and a new drug of defeating diseases. However, its molecular mechanism has still not been fully understood, which prevents its whole practical potential from being developed. This article reveals the crystal structure of dicer from Giardia intestinalis; makes us one step further to fully understanding of the initiation step of RNAi.

Firstly, the authors decided to use dicer from Giardia intestinalis, a flagellated protozoan parasite, rather than human dicer. Since the former one is smaller and have the intact enzyme activity, the whole work can be much simpler. The Giardia Dicer has six domains (as labeled in the right figure) and looks like a hatchet from this direction. The two RNase III domains form the blade, the PAZ domain makes up the base of the handle and the connector helix connects the two parts, while platform domain wraps the connector helix.

According to the authors' study, the two RNase III domains have two metal binding sites each (purple spheres in the box), which are considered as two-metal-ion cleavage sites for dsRNA. PAZ domain can bind the end of dsRNA with its 3' two-nucleotide RNA binding pocket. The distance from two-metal-ion cleavage sites of RNase III to the two-nucleotide RNA binding pocket of PAZ domain is ~65 Å, which matches the length of 25 dsRNA base pairs. Here the connector helix seems to play an important role in determining the length of siRNA to be cleaved. Variant lengths of connector helix thus cause different lengths of siRNA being produced.

With this article, several gaps of understanding of how dicer works in RNAi were filled, several previous observations were able to be explained.

Wanna know more about it? Wait for my literature review! It's coming soon!

Chen

Histone modifications silence the GATA transcription factor genes in ovarian cancer

Article: Histone modifications silence the GATA transcription factor genes in ovarian cancer
C Caslini, C D Capo-chichi, I H Roland, E Nicolas, A T Yeung and X-X Xu (2006) 1-16.



GATA transcription factor genes are important for lineage determination during development. Loss/ absence of GATA 4 and GATA 6 lead to loss of the expression of its transcriptional target such as tumor suppressor gene Disable-2, Dab 2, which has been found in ovarian cancer.
Caslini et al. believed that the mechanism pathways of other cancers are different than in ovarian cancer, although the mechanism is still not clear.

In this study, they indicated that the alternation of chromatin conformation, including the hypoacetylation and histone H3 and H4 and the decrease of reduction in di- and tri methylation of H3 lysine 4 play a critical role in silencing the GATA 4 and GATA 6 in five ovarian epithelial and carinoma cell lines HIO-117, HIO-114, A2780, SKOV3 and ES2).

Takyan

Epigenetic differences arise during the lifetime of monozygotic twins

This article examines a possible epigenetic cause for differences in the phenotypes of monozygotic twins. It has been described by one commentator as "the most comprehensive and detailed study of age-associated epigenetic changes so far reported, certainly for the case of human subjects. It is a technical tour de force with the utilisation of a battery of powerful molecular genetic methodologies coupled with competitive chromosomal hybridisations."(Martin 2005)

Phenotypical discordance between monozygotic twins is readily observed but poorly understood. Previous studies have indicated that differences do arise in the placenta, amniotic sac, and vascularisation of separate cell masses , however there has been a lack of published experimental data. As a result, the underlying causes of phenotypical discordance in traits and for common diseases between monozygotic twins remain unknown.

Eighty monozygotic twins from Spain of varying ages were used in the study. Despite sharing a common genotype, substantial epigenetic disparity was indicated between the subject twins in examinations of the global and locus-specific differences in DNA methylation and histone acetylation of the paired subjects. It was shown that older monozygotic twins exhibited substantial differences in overall content and genomic distribution of 5-methylcytosine DNA and histone acetylation which was indicated in their respective gene expression portraits. Epigenetic differences in younger twins were almost indistinguishable.

The observable tendency of older twins to exhibit phenotypical differences as a factor of aging appeared to be caused by both external (diet, lifestyle, health) and internal (epigenetic) factors. The epigenetic differences which were amplified over the lifetime of the subjects have been coined "epigenetic drift" and may be the result of defective or degraded signalling of epigenetic information through successive cell divisions.

The writers consider that epigenetic defects may occur at a much faster rate than genetic mutations as mechanisms for their correction do not appear to be as prevalent or well developed as those corresponding to genetic mutations, nor do the results of epigenetic defects appear to be so dramatic. The test results indicated that there was a role that epigenetic differences played in the discordant frequency/onset of diseases in monozygotic twins.

The study therefore led to the postulation that there may be an epigenetic cause for phenotypical discordance between monozygotic twins and that further investigation of epigenetic mechanisms is warranted.

Matt Sulman

Regulation of adult olfactory neurogenesis by insulin-like growth factor-I

The human nervous system is a stupendous piece of biological machinery. The treat of neurological impairment evokes a special dread. Damage to the spinal cord can create equal misery in just an instant.

For many years spinal cord injury has been seen as clinically irreversible, although many attempts have been made in animals to regeneration the spinal cord using a variety of transplanted cell types. Fortunately, significant process has been made recently with olfactory ensheathing cell's transplantation in injured spinal cord.

The article "Regulation of adult olfactory neurogenesis by insulin-like growth factor-I" (European Journal of Neuroscience, Vol. 22, pp. 1581–1588, 2005) investigate the roles of insulin-like growth factor-I (IGF-I) in regulating proliferation and differentiation in the olfactory mucosa. The action of IGF-I was assayed in serum-free culture combined with bromodeoxyuridine-labelling of proliferating cells and immunochemistry for specific cell types, as well as IGF-I and its receptor were expressed by the neuronal precursor and by olfactory neurons. It showed that IGF-I reduced the numbers of proliferating neuronal precursors, induced their differentiation into neurons and promoted morphological differentiation of neurons. The evidence suggests that IGF-I is an autocrine and ⁄ or paracrine signal that induces neuronal precursors to differentiate into olfactory sensory neurons.

It is hoped that one day, the transplanting of special olfactory glial ensheathing cells from the patient's own nose into the damaged spinal enable damaged spinal tissue to repair itself, and I'm sure of that day's coming.

Ying Wang

Prevention of early flowering by expression of FLOWERING LOCUS C requires methylation of histone H3 K36.

The study is interesting because this is the first demonstration on the role of H3K36 methylation in eukaryote development. It has been known from the literature that methylation of histone H3 K9 and K27 represses the expression of Flowering Locus C (FLC), which encodes a repressor of flowering in Arabidopsis; while methylation of H3K4 activates the expression of FLC. The result of this study shows that methylation of histone H3 K36 stimulates the expression of FLC lead to prevent early flowering. The methylation of H3K36 is a novel epigenetic memory code required for the expression of FLC in preventing early flowering. This study suggests for a further research on determination whether H3K36 methylation also has a critical function in other eukaryotes.

Trinh Tran

Human PAD4 Regulates Histone Arginine Methylation Levels via Demethylimination: SUMMARY

Dear people here is a summary of the article:

Histone proteins bind to the long DNA molecules and help compacts them into the euakryotic cell nucleus. Enzymes that covalently modify histone come in pairs that have opposing effects on gene regulation. For example acetylases and deactylases or kinases and phosphatases. Among these enzymes are the arginine methyltransferases and lysine methyltransferases that add methyl groups to these two amino acid residues of histones during gene regulation.
Unlike the the other enzymes mentioned above that work in opposite to each other that requires one to catalyze the addition of reactive groups and the other to remove the reactive groups; in this article, these athors describe a new enzyme (human pepetidylarginine deiminase 4 (HumanPAD4) that can perform two functions:
1. It can convert the undmodified arginine residue to citrulline in histone
2. It can also convert the methylated arginine residues in histone to citrulline by "demethyliminating" (i.e removal of the methyl group).
The significance of this is that PAD4 can modulate the expression of genes that are regulated by arginine methylases. This enzyme is thought to be the first histone demethylase enzyme identified.

Nason Pue

Epigenetic Mechanism of rRNA Gene Silencing

Here is the abstract for my literature review as promised in the

literature review topics blog.

Research over the past decade has revealed that the chromatin structure is far more than a static carrier of the genetic information encoded in the DNA as it actively and dynamically mediates regulation of processes requiring protein access to DNA, i.e. transcription, recombination and replication. Emerging evidence indicates that epigenetic factors control and regulate most nuclear processes. These factors alter chromatin structure by covalent DNA modifications, covalent histone modifications and nucleosome reorganization. A prominent case where nucleosome remodelling and histone acetylation contribute within one pathway is the silencing of rDNA promoters transcribed by RNA polymerase with targeting of the nucleolar remodelling complex (NoRC) to the rDNA promoter as initial step. NoRC is a multi-functional entity that catalyses the precise positioning of a nucleosome at the promoter but also recruits histone deacetylase (HDAC) and DNA methylase complexes which establish silent chromatin.
Raffaella Santoro and Ingrid Grummt have investigated the functional interrelationship and temporal order of NoRC-mediated chromatin remodelling, histone modification, and DNA methylation leading to rRNA gene silencing. The results of their study reveal a clear hierarchy of epigenetic events.

Have a nice and relaxing easter break. I see you all for class next tuesday.

Katharina

Methylation/phosphorylation neighboring switch

Heterochromatin contains predominantly transcriptionally silent genes and includes specialized chromosome structures such as centromers and telomeres. It has been recognized during most of the cell cycle, heterochromatin protein 1 (HP1) is concentrated in heterochromatin, but with the onset of chromosome condensation at metaphase, much of HP1 leaves the chromatin.

It has been also known that tri-methylation of the lysine at the ninth position in the tails of histone H3 (H3K9me3) is important for recruiting HP1 to discrete regions of the chromosome, thereby regulating gene expression and heterochromatin formation. Fischle et al. (2005) found that phosphorylation of the serine at the tenth position of histone H3 (H3S10ph) can reverse the recruitment of HP1, causing the dissociation of HP1 from the chromatin at metaphase.

This binary 'methyl/phos switching' permits dynamic control of the HP1-H3K9me3 interaction, resulting the formation of heterochromatin. Unlike the dissociation of HP1 from the chromatin by the demetylation of lysine 9, this switch allow H3 to maintain methylation pattern. Fischle et al. reason:
"Stable transmission of the heterochromatin-defining H3K9me3 mark is needed accurately convey, from one cell generation to the next, which regions of the genome are supposed to be permanently silenced. If removal of HP1 from M-phase chromatin were accomplished by H3K9me3-erasing demethylase activity, the epigenetic information underlying this mark-and effector-system would have to be accurately re-established at the end of every cell cycle."


Hiroshi Izumi

Clinical implications of aberrant DNA methylation patterns in acute myelogenous leukemia

This artical is abt epigenetics and leukaemia and is taken by David.

Homeotic gene repression homolog in humans

Hi,

After Tuesdays lecture Paul said to post here. So...

This one seemed interesting.

It concerns the protein hPCL3 (part of the PcG polycomb group of proteins). Which are
responsible for the 'stable epigenetic repression of homeotic genes'. The study has
found that the levels of the two proteins (large and small), which make up the complex,
are over expressed in many types of cancer. Including; colon, skin, lung, rectal,
cervical, uterus, and liver cancers.

Ok, now its in the right place. :)

Ashley.

Junk DNA May Not Be So Junky After All

A very interesting article on the so called "Junk DNA". Researchers at the McKusick-Nathans Institute of Genetic Medicine at Johns Hopkins have invented a cost-effective and highly efficient way of analyzing what many have termed "junk" DNA and identified regions critical for controlling gene function.

Mike

To deacetylise or not to deacetylise?

Breast Cancer Research and Treatment (2005) 90: 15–2
this paper discusses acetylation and deacetylation of histones (in particular in H3). If you recall histone deacetylase (HDAC) is involved in deacetylation of histones and this study found the expression of HDAC proteins turns out to be a good indicator of cancer survival rates (in specific types of breast cancer). Whats really interesting is that based on previous findings, the authors were expecting quite the opposite result!
Padideh Zareh

Epigenetic therapeutic trials in monkeys

An important test on the way toward developing therapeutic uses of RNA mediated gene suppression for humans is to trial the technologies on other primates. If you are interested in therapeutic uses of epigenetics, you may wish to follow the link to the original news article.