Chromatin remodeling and plant stress

3/2/2020 1
Presented by
ABINASH KAR
PGS18AGR7797
M.Sc (MBB)

Chromatin and its different types
What is chromatin remodeling?
Why chromatin remodeling is needed?
Mechanisms of chromatin remodeling
Relationship between chromatin modification and stress
Conclusion
02-03-2020 Department of Biotechnology 2

What is chromatin?
Chromatin is a complex which is formed to achieve a highly
condensed form by winding of DNA around proteins called
histones.
(Phillips T., 2008)

Chromatin Types
(Stralfors and KarlEkwall, 2011)
a

How DNA is packed?

What is Chromatin remodeling?
� The process of making DNA more or less accessible in the eukaryotic
genome using a series of specialized proteins.
� Chromatin remodelling is the dynamic modification of chromatin
architecture to allow access of condensed genomic DNA to the regulatory
transcription machinery proteins, and thereby control gene expression.
� Chromatin remodelling is the enzyme-assisted process to facilitate access of
nucleosomal DNA by remodelling the structure, composition and positioning
of nucleosomes.
(Phillips T., 2008)

Why Chromatin remodeling?
(Phillips T., 2008)

(b) (c)(a)
How Chromatin remodeling occurs?
(Clapier et al., 2017)

.
Mechanism of chromatin
remodeling
ATP-Dependent
Chromatin
remodeling
Covalent modification
of histone

ATP-Dependent
Chromatin remodelers
(CR) subfamily
ISWI
SWI/
SNF
CHD
INO80/S
WR1

ISWI (Imitation SWItch)
Structure:
N C
AutoN (Auto-inhibitory N-terminal) & NegC (Negative regulator of
coupling) :
Two domains that flank the ATPase lobes and regulate the activity of
the ATPase domain.
ATPase Domain: Contains two Rec A-like lobes i.e. Lobe 1 (DExx) and
Lobe 2 (HELICs), which are separated by a small insertion sequence.
HSS (HAND SANT SLIDE) : Binds the unmodified histone H3 tail and
the linker DNA flanking the nucleosome. (Clapier et al., 2017)

Functions:
Assemble and regularly space nucleosomes to limit chromatin
accessibility and gene expression.
A subset, Nucleosome remodelling factor (NURF) complex, have
accessory subunits that confer access and that promote transcription.

SWI/SNF (SWItch/Sucrose non-fermentable)
Structure:
 HSA (Helicase SANT associated) Domain: Binds actin and/or actin-related
proteins (ARPs).
 ATPase Domain: Contains two Rec A-like lobes i.e. Lobe 1 (DExx) and Lobe
2(HELICs), which are separated by a small insertion sequence.
 Two AT Hooks, SnAC (Snf2 ATP coupling): It is a histone binding domain (HBD),
maintain octamer attachment during forcible DNA translocation, a property
that is necessary for nucleosome ejection.
 C-terminal Bromodomain.
Functions
:Typically facilitates chromatin access, as they slide and eject nucleosomes, and
are used for either gene activation or gene repression.
N C

CHD (Chromodomain Helicase DNA binding
Structure:
 Resemble that of ISWI sub-family but differ in its two signature amino
terminus of tandemly arranged Chromodomains.
 ATPase Domain: Contains two Rec A-like lobes i.e. Lobe 1 (DExx) and Lobe
2(HELICs), which are separated by a small insertion sequence.
 NegC Domain.
 DBD (DNA binding domain): Comprised of only SANT and SLIDE domains.
Functions
:Nucleosome assembly (spacing nucleosome), Chromatin access (exposing
promoters), Nucleosome editing (substitute histones or histone variants).
N C

INOsitol requiring (INO80) complexes
Structure:
 HSA (Helicase SANT associated) Domain: Binds actin and/or actin-related
proteins (ARPs).
 ATPase Domain:
� Contains two Rec A-like lobes i.e. Lobe 1 (DExx) and Lobe 2(HELICs), which
are separated by a long insertion sequence (Binds to a hetero-hexameric
ring of the helicase related (AAA + ATPase).
� It scaffold three modules: N-terminus, Rvb 1 and Rvb 2, and C-terminus.

INO80 subfamily remodellers have unique editing functions.
Although INO80C also conducts chromatin access and nucleosome
spacing functions.
The SWR1C, p400 and Snf2-related CBP activator protein (SRCAP)
complex subtypes replace canonical H2A–H2B dimers with H2A.Z histone
variant-containing H2A.Z–H2B dimers, whereas INO80C can catalyse the
reciprocal reaction.
The vertebrate p400 subtype may also replace H3.1 with the variant H3.3.
Yeast INO80C removes the variant H2A.X, which probably underlies its
DNA repair functions, as well as its chromatin access and transcription
activation functions.
Functions:

Covalent modification of histone
�Histone Acetylation.
�Histone Methylation.
�Histone Phosphorylation.
�Other modifications:
Deimination.
Ubiquitylation.
(Bannister and Kouzarides, 2011)

Histone Acetylation/deacetylation

Histone Methylation/demethylation
Histone arginine methylation

Histone arginine demethylation
Histone lysine demethylation

Histone Phosphorylation
Serine/Threonine kinase

� There may be competitive antagonism between modifications if more than one
modification pathway is targeting the same site(s). This is particularly true for
lysine that can be acetylated, methylated or ubiquitylated. (Mutually exclusive)
� One modification may be dependent upon another. A good example of this trans-
regulation comes from the work in Saccharomyces cerevisiae; methylation of
H3K4 and of H3K79 is totally dependent upon the ubiquitylation of H2BK123.
Importantly, this mechanism is conserved in mammals, including
humans. (Dependent)
� The binding of a protein to a particular modification can be disrupted by an
adjacent modification. For example, as discussed above, HP1 binds to
H3K9me2/3, but during mitosis, the binding is disrupted due to phosphorylation
of H3S10. This action has been described as a ‘phospho switch’. (Inhibit
protein binding)
� There may be cooperation between modifications in order to efficiently recruit
specific factors. For example, PHF8 specifically binds to H3K4me3 via its PHD
finger, and this interaction is stronger when H3K9 and H3K14 are also acetylated
on the same tail of H3. (Supportive modification)
Histone modification crosstalk
02-03-2020 Department of Biotechnology 24(Bannister and Kouzarides, 2011)

Stress induced chromatin modification
(Fabado et al., 2017)

Abiotic stress induced modification
(Fabado et al., 2017)

Stress-Mediated Alterations in Chromatin
Architecture Correlate with Down-Regulation of
a Gene Encoding 60S rpL32 in Rice
(Mukhopadhyay et al., 2013)
 Down-regulation of rpL32_8.1 under salt stress is not mediated by
DNA methylation.
 Down-regulation of rpL32_8.1 is accompanied by a shift in the
first nucleosome downstream of the TSS.
 Alteration in histone modifications within the 5’-UTR correlate
with the down-regulation of rpL32_8.1.

Down-regulation of rpL32_8.1 under salt stress is not mediated by DNA
methylation.

Down-regulation of rpL32_8.1 is accompanied by a shift in the first nucleosome
downstream of the TSS

Alteration in histone modifications within the 5'-UTR correlate with the down-
regulation of rpL32_8.1. (By ChIP-PCR)

Generation of Arabidopsis transgenic lines expressing dCas9
Molecular and phenotypic characterization of dCas9HAT-sgA
dCas9HAT-sgA plants have enhanced drought stress tolerance

Generation of Arabidopsis transgenic lines expressing dCas9

Molecular and phenotypic characterization of dCas9HAT-sgA

sgA1
sgA2
Control

dCas9HAT-sgA plants have enhanced drought stress tolerance

Rehydration
MSDS 48hr
Stomatal Aperture

A schematic model for dCas9HAT function in transcriptional
activation

CONCLUSION

THANK YOU

Chromatin remodeling and plant stress

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Chromatin remodeling and plant stress