Dynamics of assembly and disassembly of nucleosomes along a stretched DNA

1
Dynamics of assembly and disassembly of
nucleosomes along a stretched DNA

• Ranjith Padinhateeri
• work done with
• Jie Yan and John F. Marko

University of Illinois at Chicago/ Northwestern
University
2
Nucleosome -- basic unit of Chromatin
DNA
Nucleosomes
Transcription, replication and other in-vivo DNA
processing in eukaryotes take place in the
context of chromatin.
Molecular Bilogy Of the Cell Alberts et al
3
Nucleosome
50nm of DNA is wrapped around histone octamer
(Alberts et al, MBC)
4
Nucleosome assembly/disassembly kinetics, under
external force
f
DNA

f
nucleosome

l150bp
f

Stalling force 3.5pN

• Single molecule experiments measure change in
• end-to-end distance (approximately 50
  nm/nucleosome)
• We try to understand experiments in xenopus egg
• extracts (chaperones present) with no ATP

5
Experiments show
lt Assembly constant force (magnetic
tweezer) Yan et al Mol Bio Cell, 2007 Fast decay
followed by slow tail
Assembly/disassembly Constant force (Yan et al)
Disassembly -- constant velocity Bennink et al
(nat. str. bio 2001)
6
Assembly of hard-core particles
Single molecule experiments measure change in
end-to-end distance (approximately 50
nm/nucleosome) Nucleosomes are like hard-core
particles(steric interaction)
f
nucleosome


l150bp (50nm)

• New nucleosomes can occupy only the empty space
  on
• the DNA. Empty space decreases as nucleosomes
  assemble.
• So is the end to end distance. There is a
  one-to-one relation
• between the empty space and the end-to-end
  distance

7
Simple model Random sequential adsorption
Randomly place hard-core particles on a
lattice(DNA is like a Lattice)
and measure the empty space on the lattice at
t0 the whole line is empty
As each particle is adsorbed number of empty
sites are reduced by a certain amount (in the cae
of nucleosome it is about 50nm)
We can measure the empty space and compute the
end-to-end length
8
Model Adsorption desorption and diffusion of
nucleosomes
Hard-core particles of length l (50 nm) Measure
the the total gap length (empty space on the
line) Total gap length is related to the
end-to-end-distance measured in the experiments
(we compute exptly measured end-to-end extension
from this gap length)
9
Nucleosome Diffusion

• Thermal fluctuations can form
• loops on the nucleosomes
• and reptation of these loops
• could lead to repositioning of the
• nucleosome (Schiessel et al, PRL,. 2001, 2002))

In egg extract conditions, at f1pN, D(f) 3
10-15 cm2/sec (using free energy 42kBT)
NAP-1 assists Nucleosome sliding Y-J Park et
al, J. Bio. Chem. (2005)
(Schiessel et al)
10
Sequence dependence of nucleosome positioning
It is recently proposed that there is sequence
dependence in nucleosome positioning (Segal,
Widom et al, Nature 2006)
Their model computes the sequence dependent
potential Vi in which nucleosomes move around
(rates now depend on this potential)
We determine ltVigt -42 kBT based on experiments
(corresponding to stalling of 3.5 pN) We intend
to test the role of sequence in explaining the
experimental results
11
Model for adsorption, desorption and diffusion
in this potential
l
on
off
diffuse
Given the potential and rates, we use Monte-Carlo
simulation to obtain the dynamics of assembly
and disassembly
Energies (like V) are expressed in units of kBT
12
Randomly place hard-core dimers on a lattice
what happens If there is no sliding and no
desorption?
Jammed! no more dimers can be added
When 147-mers(nucleosomes) are placed on a DNA
randomly, the average amount of empty space is
aproximately 25 of thetotal length of the DNA.
One need to have desorption/diffusion to get to
higher density of nucleosomes.
13
Dynamics of assembly -- constant force
(f1pN)

• NO ATP
• Egg extract

Experiment Yan et al, Mol Bio Cell (2007) Model
with D5 10-15 cm2/sec Model with No diffusion
(jamming gap density 0.25) Inset zoomed in
view -- final filling facilitated by diffusion
Early time exponentially fast filling Late
time reorganization via diffusion --gt slow
filling
14
Assembly and disassembly -- constant force
Velocity of one end of the DNA
Model Assembly stalls around 3.5 pN Barrier to
unwrap
Experiment Yan et al Mol. Bio. Cell(2007)
determines the contribution of the force into
the on-rate (75 ) determine barrier height.
Dynamics display memory of nucleosome
configuration
15
Dynamics of disassembly -- constant velocity
Experiment Bennink et al (2001) Nature Str. Biol.
Model
DNA of Lambda phage virus DNA with homogeneous
sequence
Sequence is essential to explain the slope in the
curve
16
World-line of nucleosomes -- disassembly - lambda
DNA
Each line corresponds to a nucleosome. Line ends
when the nucleosome is desorbed from the DNA
Nucleosomes on the left part are short lived --
this is due to a specific nature of lambda DNA
sequence. This prediction can be tested
experimentally.
17
Conclusion

• A model with adsorption, desorption and diffusion
  of nucleosomes can explain some of the in vitro
  experiments
• We can estimate the adsorption-rate, desorption
  rate and the amount of nucleosome sliding.
• We estimate the dependence of force in the on-
  and off- rates.
• Sequence effects are important in explaining
  experiments.