lezione 22 epigenetics iii · four amino acids guide the assembly or disassembly of arabidopsis...

Lezione 22Epigenetica III

Different types of chromatin:

Active chromatin - Chromatin can become ‘opened’ at gene regulatory regions, which allows RNA polymerase to function.accompanied by ‘active’ covalent modifications of the core histones, including histone H3 methylation on lysine 4 (H3K4me) and H3K9 and H3K27 acetylation At promoters, these modifications can flank a nucleosome-free region immediately upstream of the transcription start site.

‘Low signal state’ chromatin Chromatin that does not accumulate either active or repressive histone modifications and thus can be considered as unprogrammed.

Actively repressed chromatin this chromatin is inhibitory to the binding of most transcription factors, including pioneer factors, and thus strongly repress transcription. Such closed domains are typically accompanied by repressive covalent modifications of the core histones, including H3K9 or H3K27 methylation, although usually it does not possess both marks at the same time, thus reflecting the different mechanisms of chromatin repression:

-facultative heterochromatin often forms at developmentally regulated genes, and its level of compaction changes in response to developmental cues and/or environmental signals

-constitutive heterochromatin preferentially assembles at repetitive elements such as satellite DNA and transposons, and maintains high compaction level.Iwafuchi-Doi and Zaret, Development, 2016Wang et al., Trends in Genetics 2016

© 2016 American Society of Plant Biologists

H3K27me3 methylation is conferred by Polycomb Repressive Complex 2

Active genes Silenced genes

Polycomb Repressive Complex 2

H3K27me3

Polycomb repressive complex proteins are also known as Polycomb group proteins (PcG)


E(Z) (methylase)

SU(Z)12ESC NURF55

Drosophila PRC2

Arabidopsis PRC2 CURLY LEAF (CLF)

MEDEA (MEA)

SWINGER (SWN)

FERTILIZATION-INDEPENDENT SEED 2

(FIS2)

EMBRYONIC FLOWER 2 (EMF2)

VERNALIZATION 2 (VRN2)

FERTILIZATION INDEPENDENT ENDOSPERM

(FIE)

MULTICOPY SUPPRESSOR

OF IRA1 (MSI1,2,3,4,5)

Arabidopsis has multiple forms of three of the core PRC2 proteins


Plants make multiple PRC2 complexes with different targets

MEA + FIS2 complex

CLF/SWN + VRN2 complex

CLF/SWN + EMF2 complex

Seed development

Transition to flowering

Floral organogenesis


In animals, H3K27me3 silencing is maintained by PRC1

Active genes Silenced genes Stably silenced genes



H3K27me3


In plants, PRC1-like functions similarly

Active genes Silenced genes Stably silenced genes


H3K27me3

PRC1-like

LHP1

LHP1 binds specifically to H3K27me3


LHP1 co-localizes with H3K27me3

Turck F, Roudier F, Farrona S, Martin-Magniette M-L, Guillaume E, et al. 2007 Arabidopsis TFL2/LHP1 specifically associates with genes marked by trimethylation of histone H3 lysine 27. PLoS Genet 3(6): e86.

Note the close correlation of LHP1 and H3K27me3 distributions

http://dx.doi.org/doi:10.1371/journal.pgen.0030086


LHP1 co-localizes with H3K27me3

Here’s a transposon marked with H3K9me2!

Turck F, Roudier F, Farrona S, Martin-Magniette M-L, Guillaume E, et al. 2007 Arabidopsis TFL2/LHP1 specifically associates with genes marked by trimethylation of histone H3 lysine 27. PLoS Genet 3(6): e86.

http://dx.doi.org/doi:10.1371/journal.pgen.0030086


The histone variant H2A.Z promotes transcription and is swapped into the nucleosome by the SWR1/SRCAP complex.

SWR1/ SRCAP complex

H2AH2A.Z

Inactive geneActive gene

An H2 variant, H2A.Z is associated with some active genes


There are three histone H3 variants: H3.1, H3.3, and CENH3

Redrawn from Jiang, J., Birchler, J.A., Parrott, W.A., and Dawe, R.K. (2003) A molecular view of plant centromeres. Trends Plant Sci. 8: 570-575 with permission from Elsevier. Stroud, H., Otero, S., Desvoyes, B., Ramírez-Parra, E., Jacobsen, S.E. and Gutierrez, C. (2012). Genome-wide analysis of histone H3.1 and H3.3 variants in Arabidopsis thaliana. Proc. Natl. Acad. Sci. USA 109: 5370-5375. See also Shi, L., Wang, J., Hong, F., Spector, D.L. and Fang, Y. (2011). Four amino acids guide the assembly or disassembly of Arabidopsis histone H3.3-containing nucleosomes. Proc. Natl. Acad. Sci. USA 108: 10574-10578.

Nucleosomes at the centromere incorporate an H3 variant called CENH3 that is necessary for centromere maintenance

CENH3H3 H3.1 and H3.3

differ in only four amino acids, but their distribution is very different. H3.1 is associated with heterochromatin, and H3.3 with transcribed genes

http://www.cell.com/trends/plant-science/abstract/S1360-1385(03)00274-7

http://www.pnas.org/content/109/14/5370.abstract

http://www.pnas.org/content/108/26/10574.short


Chromatin remodelers use energy to move or alter histone octamers

Reprinted by permission of Annual Reviews from Clapier, C.R. and Cairns, B.R. (2009). The Biology of Chromatin Remodeling Complexes. Annu. Rev. Biochem. 78: 273-304.

http://www.annualreviews.org/doi/abs/10.1146/annurev.biochem.77.062706.153223?journalCode=biochem


Example: Model for reversal of polycomb repression

Wu, M.-F., Sang, Y., Bezhani, S., Yamaguchi, N., Han, S.-K., Li, Z., Su, Y., Slewinski, T.L. and Wagner, D. (2012). SWI2/SNF2 chromatin remodeling ATPases overcome polycomb repression and control floral organ identity with the LEAFY and SEPALLATA3 transcription factors. Proc. Natl. Acad. Sci. USA 109: 3576-3581.

Polycomb Repressive Complex (PRC) Repressive histone marks (Red stars)

Loss of PRC Activating histone marks (Green stars)

In this example, the chromatin remodelers SPLAYED (SYD) and BRAHMA (BRM) facilitate switching of a gene from OFF to ON

http://www.pnas.org/content/109/9/3576.abstract

Eterocromatina facoltativa: funzione dei complessi regolativi Polycomb nei geni omeotici


Epigenetic controls in whole-plant processes

• Transposon silencing • Control of flowering time • Developmental switches and stress

responses • Control of imprinted genes • Gene silencing in trans; paramutation • Resetting the epigenome

Eterocromatina facoltativa: esempio del Flowering locus C di Arabidopsis

Michaels and Amasino, Plant Cell 1999


Epigenetic programming controls developmental transitions

Embryonic development

Vegetative development

Reproductive development

Vegetative to reproductive transition

Embryonic to vegetative transition


Epigenetic control of flowering time

Some plants require a prolonged cold period (vernalization) - as experienced during winter, before they will flower.

Vegetative Development

Reproductive Development

Prolonged cold treatment

Winter SpringAutumn


FLOWERING LOCUS C (FLC) mutants flower early

Winter SpringAutumn

FLC is an inhibitor of flowering; removing FLC removes the vernalization requirement

flc mutant


FLC inhibits FT, an activator of flowering

FLCTranscription of FT gene repressed by FLC binding

FT gene

FT gene

FT

Wild-type plant

flc mutant plant


FLC is silenced by vernalization

FLC gene transcribed FLC gene silenced

Winter SpringAutumn

After 40 days at 4ºC, FLC is not expressed. Ten days after return to 22ºC, FLC expression is still off.

Reprinted by permission from Macmillan Publishers, Ltd: NATURE Sung, S., and Amasino, R.M. (2004) Vernalization in Arabidopsis thaliana is mediated by the PHD finger protein VIN3. Nature 427: 159-164. Copyright 2004.

During vegetative growth, FLC is transcribed at a high level. Epigenetic marks maintain this transcriptional activation

http://www.nature.com/nature/journal/v427/n6970/abs/nature02195.html


FLC is regulated by epigenetic modifications

H2A.Z incorporation H3K4me, H3K36me H3K9Ac, H3K14Ac

H3K9me2, H3K27me2 H3K27me3

FLC gene transcribed FLC gene silenced

cold

Winter SpringAutumn


VIN3 and the PRC2 complex epigenetically silence FLC

PRC2 (including

VIN3)FLC gene transcribed FLC gene silenced

VIN3

Winter SpringAutumn

vernalization: the acceleration of flowering through exposure to prolonged cold

FLC expression is repressed by prolonged cold exposure, and this repression is then epigenetically maintained until embryo development after flowering

Berry and Dean, Plant Journal 2015

sense FLC and antisense COOLAIR transcripts

The periods of growth before and after cold exposure are periods of stable FLC expression, whereas vernalization and embryogenesis down-regulate and up-regulate FLC expression, respectively.

FLC: floral repressor locus

A series of antisense RNAs named COOLAIR and the lncRNA called COLDAIR are encoded by the FLC locus and upregulated during vernalization. COLDAIR physically interacts with the PRC2 complex to repress FLC gene expression during vernalization

Cell-autonomous Switching Underlies Quantitative Silencing During Cold Exposure

cold exposure instead increases the number of cells that have switched from an ON state to an OFF state

Polycomb Repressive complex 2 (PRC2)-mediated H3K27 methylation forms facultative heterochromatin.

It is not yet clear how PRC2 is recruited, although the antisense COOLAIR might play a role.


Winter SpringAutumn


FLC expression is also regulated by chromatin remodeling proteins

Reprinted from Jégu, T., Latrasse, D., Delarue, M., Hirt, H., Domenichini, S., Ariel, F., Crespi, M., Bergounioux, C., Raynaud, C. and Benhamed, M. (2014). The BAF60 subunit of the SWI/SNF chromatin-remodeling complex directly controls the formation of a gene loop at FLOWERING LOCUS C in Arabidopsis. Plant Cell. 26: 538-551.

BAF60 is a SWI/SNF subunit. By modulation of histone density, composition, and posttranslational modification, BAF60 activity creates a repressive chromatin configuration at the FLC locus.

http://www.plantcell.org/content/26/2/538


Summary: Mechanisms underlying FLC activation and repression

Reprinted from Hepworth, J. and Dean, C. (2015). Flowering Locus C’s lessons: Conserved chromatin switches underpinning developmental timing and adaptation. Plant Physiol. 168: 1237-1245.

http://www.plantphysiol.org/content/168/4/1237.long


Natural variation of flowering time can be correlated to FLC responses

Reprinted from Hepworth, J. and Dean, C. (2015). Flowering Locus C’s lessons: Conserved chromatin switches underpinning developmental timing and adaptation. Plant Physiol. 168: 1237-1245.

http://www.plantphysiol.org/content/168/4/1237.long


Epigenetic control of flowering time• During vernalization, silencing of FLC allows expression of FT

and other flowering promoters

• VIN3 and PRC2 complex proteins epigenetically modify FLC to silence it

• Maintaining the silenced state requires PRC1-like complex which includes LHP1

• Trithorax protein SDG26 induces expression of flowering activator SOC1 by methylating H3 Lys residues at SOC1 promoter

• Long non-coding RNAs contribute to epigenetic control of gene expression


Winter SpringAutumn

FLC ON FLC OFF

When does FLC switch on again between generations?

Resetting the epigenome (or not…)

Reprogramming FLC Expression in the Seed

Unlike the slow quantitative switching from the ‘ON’ to ‘OFF’ state during cold exposure, the reprogramming occurs relatively synchronously in the developing seeds. FLC expression increases throughout embryogenesis, and reaches a maximum when the seed has fully formed.

ELF6 is a jumonji domain-containing protein with H3K27me3 demethylase:The elf6-5 mutant doens not have an increase in FLC expression and fails to fully remove the H3K27me3 modification.



Holec, S. and Berger, F. (2012). Polycomb group complexes mediate developmental transitions in plants. Plant Physiol. 158: 35-43.

Many developmental genes and stress-responsive are epigenetically regulated

http://www.plantphysiol.org/content/158/1/35.full


Reprinted from Probst, A.V. and Mittelsten Scheid, O. (2015). Stress-induced structural changes in plant chromatin. Curr. Opin. Plant Biol. 27: 8-16.

Stress can cause diverse epigenetic changes including…

Unstressed

Stressed

Heterochromatin decondensation

Transposon activation

Gene activation or silencing

http://www.sciencedirect.com/science/article/pii/S136952661500062X


Winter SpringAutumn

FLC ON FLC OFF

When does FLC switch on again between generations?

Resetting the epigenome (or not…)


Epigenetic reprogramming in plants

Generative cell

Vegetative nucleus

3n endosperm

Large-scale epigenetic changes have been observed in the endosperm and in the vegetative nucleus of the male gametophyte.


Summary

• Expression of DNA is controlled by epigenetic marks including DNA methylation and histone modifications.

• siRNAs contribute to epigenetic programming

• Epigenetic programming silences transposons and controls the timing of many genes that control plant development and stress responses.

lezione 22 epigenetics iii · four amino acids guide the assembly or disassembly of arabidopsis...

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