Transposon and Epigenetic Regulation Liang Wu, Haiyun Zhang, Weizhen Cai

Transposon/ Transposable Elements All mobile DNA segments in the genome, regardless of their mechanism of transposition. Consists of ~50% of human genome. Transpose is the movement of a genetic element from one location of the genome to another. Retrotransposon and DNA transposons, autonomous and non-autonomous.

Retrotransposons Derived from viral genome. R. Keith Slotkin & Robert Martienssen. Nature Reviews Genetics 8, 272-285 (April 2007)  Derived from viral genome. LTR: Long Terminal Repeats. Needs to be reverse transcribed. SINE (Short Interspersed Elements) and LINE (Long Interspersed Elements) are non-LTR retrotransposons.

DNA Transposons Transposase is responsible for transposition. Transposase mutated Miniature Inverted-Repeat Elements New member of transposon R. Keith Slotkin & Robert Martienssen. Nature Reviews Genetics 8, 272-285 (April 2007) 

Transposition Henry L. Levin & John V. Moran. Nature Reviews Genetics 12, 615-627 (September 2011)

Consequences Richard Cordaux & Mark A. Batzer. Nature Reviews Genetics 10, 691-703 (October 2009) Transposons are highly mutagenic, cause insertion, chromosome breakage, recombination and genome rearrangement.

Transposon Needs Regulation Number of transposable elements negatively correlated with fitness of the host. Silenced by different mechanisms.

Silencing Mechanisms that Suppress TEs Post-transcriptional silencing of TEs by RNAi Chromatin modification RNAi-mediated chromatin modification

Post-transcriptional Silencing of TEs by RNAi dsRNA cleaved by Dicer siRNAs RISC, Argonaute siRNA-guided transcript -cleavage complex, RISC; Argonaute proteins is the cataclytic component. RdRP can additionally produce dsRNA from the cleaved RNA products.

Post-transcriptional Silencing of TEs by RNAi RNAi is the main mechanism of TE silencing C. elegans, silencing Tc1 DNA transponson in germ line Mutations in both argonaute- and dicer- family proteins cause the reactivation of TEs in eukaryotic species TEs give rise to numerous siRNAs. How TEs are specifically targeted by RNAi? stuctures help to distinguish transcripts 3, in most species, including human.

Chromatin Modification Modification of histone tails Alert protein factors binding, send information to transcription factors Methylation of histone H3K9 is a signal for transcriptionally repressive and inactive chromatin. DNA methylation Cytosine residue, in a symmetrical context (CpG) Inheritable (mice, DNMT1 DNA methyltransferase) Alterations in chromatin packing and condensation SWI/SNF chromatin-remodelling proteins are required for TE silencing. (plant) DDM1 in A. thaliana CpG methylation is copied to new DNA strand upon DNA replication, providing a mechanism for inheritance of TE silencing. DNMT1 DNA methyltransferase is reponsible for this maintenance of DNA methylation. Elevated IAP retrotransposon transcript levles in Dnmt-1-deficienct embryos. SWI/SNF chromatin-remoldelling proteins use ATP hydrolysis to remodel nucleosomes, which are required for TE silencing. DDM1 is a SWI/SNF2-like protein, which is specifically required for TE silencing and chromatin condesation. If passed from a ddm1 mutant to wild-type background, TEs remains actived and do not regain DNA and H3K9 methylation. Remodelling activity is specifically targeted to TEs: only TE transcripts are elevated in ddm1 mutants, most genes are unaffected.

RNAi-mediated Chromatin Modification The maintenance of TE silencing at chromatin level involves both RNAi-independent and RNAi dependent pathway. In S. pombe (fission yeast) RNAi complex works on nascent RNAs Argonaute proteins (Ago1) cleave nascent RNAs; the cleavage recruites the histone H3K9 methyltransferase Clr4 to the chromosome, mechanism unknown. RNAi is specifically required for spreading H3K9 methylation from repeats into reporter genes. d nascent RNAs are still bound to the chromosome and are processed co-transcriptionally into siRNAs.

RNAi-mediated Chromatin Modification How do TEs that are silenced at the chromatin level produce transcripts that are to be cleaved into siRNAs? In A. thaliana, plant-specific RNA polymerase Ⅳ Loss of PolⅣ can transcriptionally reactive silenced TEs. PolⅣa: generate siRNAs PolⅣb: transfer siRNA signal Symmetrical DNA methylation is heritable; Asymetrical methylation must be replaced at each cell division (less important). Targetting mechanism perhaps involves RNAi. polⅣb is probably guided to the correct location by its C-terminal domain, which is significantly longer than that of PolVa. C-terminal domain interacts directly with AGO4 in specific place( nuclear bodies) in an RNA-dependent manner, and might serve as a platform for different protein factors.

TEs influence the Genomes TEs contribute to chromosome form and function TE-induced epigenetic regulation of specific genes Evolutionary implications

TEs contribute to chromosome form and function Centromeres, telomeres and knobs PEV and chromatin insulators X inactivation

Centromeres and Telomeres The centromeres are function as trafficking chromosomes at cell division. The DNA composition of centromeres differs widely in both length and primary nucleotide sequence between species, and it is the specific epigenetic context that confers their function. Thus the presence of TEs, and the epigenetic silencing apparatus that they recruit play a role in centromere function. The telomeres work for preventing chromosome shortening following replication. Although whether TEs contribute to telomere- length regulation in mammals remains unknown, recent studies show that the epigenetic status (predominantly DNA methylation) of telomeric and sub-telomeric repeats has a role in regulating telomere elongation in mammalian cells.

KNOB A chromosomal knob is a fragment of constitutive heterochromatin that has no known function. Analysis of an A. thaliana knob, composed almost, entirely of full- length and fragmented TEs (many of which are inserted into each other), showed that heterochromatin was dependent on the epigenetic mechanisms of TE silencing.

Position Effect Variegation (PEV) and chromatin insulators PEV: Heterochromatin can insert into adjacent genes and causes gene-silencing phenotype. TEs are nucleation centers for the formation of the type of facultative heterochromatin. TEs themselves can function as insulators.

X inactivation Female mammals must compensate for their double dosage of X- chromosome genes compared with XY males. This compensation is achieved by the inactivation and heterochromatinization of one of the two X chromosomes. Inactivation is initiated at the X-chromosome inactivation center (XIC), and then spreads outwards from the XIC to the rest of the chromosome. TEs function in both the initiation and spread of X inactivation. This is indirectly supported by a study on mice cells.

TE-induced epigenetic regulation of specific genes Epiallele production and phenotypic variation Paramutation Imprinting V(D)J Recombination

Epiallele production and phenotypic variation TE promoters can influence the transcriptio of nearby genes. In mice, the intracisternal A-particle (IAP) retrotransposon produces an outward-reading transcript that extends into the agouti coat-color gene. The level of agouti transcript, and the color of the coat, is subject to the epigenetic status of the retrotransposon and inheritable.

Paramutation Paramutation is an interaction between alleles that results in a heritable expression change of one allele. In maize, several genes in the anthocyanin pigmentation pathway have paramutable alleles, which are associated with and pendent on TEs.

Imprinting On the mammalian X chromosome, TEs are specifically associated with monoallelic expressed genes that are located on autosomes, raising the possibility that TEs are involved in the regulation of imprinted genes. The effect of TE-mediated imprinting in mammals might be mediated through the action of DNMT3L

Evolutionary implications In the same specie, TEs have the ability to mutate genes, alter gene regulation and generate new genes, each providing fuel for evolution. the stress-induced reactivation of TEs and the invasion of a new genome by TEs. In different species, TEs can generate intraspecies variation. This happens in virus, plants, and animals.

Conclusions TEs are all mobile DNA segments in the genome. Classified as retrotransposon, DNA transposon, autonomous and non-autonomous. TEs silencing mechanisms: Post-transcriptional silencing by RNAi, chromatin modification(H3K9, CpG, SWI/SNF), RNAi-mediated chromatin modification. TEs are involved in many epigenetic mechanisms, from chromosome form and function to specific genes, playing significant role in evolutionary history.