2. Organization of DNA Within a Cell
2 meters of DNA is packed
into a 10 mm diameter cell

4. Appearance of Chromatin Depends on Salt Concentration
Physiological ionic strength
30 nm fiber
Low ionic strength
Beads on a string
from Lodish et al., Molecular Cell Biology, 6th ed. Fig 6-28

6. Nucleosomes are Packaged into a 30 nm Fiber
from Lodish et al., Molecular Cell Biology, 6th ed. Fig 6-30

7. Chromatin Organization
Each layer of chromatin organization
reflects aspects of gene regulation
Scaffold-associated regions
can act as boundaries
Condensed chromosomes
are visible during metaphase
from Zhou et al., Nature Rev.Mol.Cell Biol. 12, 7 (2011)

10. Genes Can be Localized on Drosophila Polytene Chromosomes
Polytene chromosomes exhibit
a characteristic banding pattern
Localization of a gene by in situ hybridization
Biotinylated probe was detected by
avidin conjugated to alkaline phosphatase
AP substrate results in the formation of an
insoluble precipitate at the site of hybridization
from Lodish et al., Molecular Cell Biology, 6th ed. Fig 6-44

11. Chromosome Puffs Ecdysone produces a characteristic pattern of
puffs in polytene chromosomes of salivary glands
Puffs correspond to actively transcribed genes
from Alberts et al., 3rd ed., Fig. 8-23

13. Actively Transcribed Genes are Present in Decondensed Chromatin
Loss of 4.6 kb Bam HI fragment
when the b-globin gene is active
and histones are acetylated
The 4.6 kb Bam HI fragment is
present when the b-globin gene is
inactive and histones are deacetylated
from Lodish et al., Molecular Cell Biology, 6th ed. Fig 6-32

14. Nucleosome Structure Nucleosomes contain 2 copies
of H2A, H2B, H3 and H4
147 bp of DNA is wrapped
around nucleosome
Histone tails emanate from core
Some nucleosomes
contain histone variants
H1 is a linker histone
from Jiang and Pugh, Nature Rev.Genet. 10, 161 (2009)

15. Assembly of Nucleosomes
Histone chaperones assemble
histones into nucleosomes
Histone chaperones prevent
non-specific associations
of histones with DNA
Histone chaperones prevent
formation of deleterious
off-pathway intermediates
from Das et al., Trends Biochem.Sci. 35, 476 (2010)

16. Histone Tails Histones contain flexible termini
that extend from the globular
structure of the nucleosome
from Lodish et al., Molecular Cell Biology, 6th ed. Fig 6-31

17. Modification of Histone Tails
Chromatin structure is a
source of epigenetic information
Posttranslational modifications and
histone variants contribute to structural
and functional characteristics of chromatin
The combination of histone
modifications constitutes the histone code
The histone code influences chromatin
condensation and function and defines
actual or potential transcription states
from Lodish et al., Molecular Cell Biology, 6th ed. Fig 6-31

18. Histone Modifications Affect Chromatin Structure
H3K4 methylation and H3K9 acetylation
are hallmarks of active chromatin
H3K27 methylation and H3K9 methylation
are hallmarks of silent chromatin
from Johnstone and Baylin, Nature Rev.Genet. 11, 806 (2010)

19. Histone Modifications Define Functional Elements
Each histone modification
has a unique biological role
Histone modifications
are interdependent
from Zhou et al., Nature Rev.Mol.Cell Biol. 12, 7 (2011)

20. Chromatin Immunoprecipitation (ChIP)
Fix cells with formaldehyde
Isolate and shear chromatin
Immunoprecipitate with antibodies
to modified histones or binding proteins
Identify binding sites by qPCR or Southern blot
from Orlando, Trends Biochem.Sci. 25, 99 (2000)

21. Nucleosome Position Can Be Mapped
be mapped by ChIP-seq
Cross-link histones to
DNA and digest linker
Immunoprecipitate
Sequence DNA
from Jiang and Pugh, Nature Rev.Genet. 10, 161 (2009)

22. Positioning of Nucleosomes at Promoters
Nucleosome-free region at the
beginning and end of genes
Nucleosomes have defined
locations near the promoter
Nucleosome position is determined
by ATP-dependent trans-acting factors
from Jiang and Pugh, Nature Rev.Genet. 10, 161 (2009)

23. Properties of Acetylated Histones
Less positively charged
Chromatin is less condensed
H4K16Ac prevents formation of 30 nm fiber

24. Control of Gene Expression by Acetylation
Repressor recruits a complex
that contains a histone deacetylase
Neighboring histones are deacetylated
Activator recruits a complex
that contains a histone acetylase
Neighboring histones are acetylated
from Lodish et al., Molecular Cell Biology, 6th ed. Fig 7-38

25. Effect of Histone H3 K9 Methylation
SUV39 methylates K9
Methylated K9 recruits HP1
Heterochromatin formation
HP1 binds to SUV39 to
propagate methylation
Methylated K9 or phosphorylated S10
inhibits methylation of K9
from Turner, Cell 111, 285 (2002)

26. Effect of Histone H3K4 Methylation
Set9 methylates K4
Inhibits association of NuRD
remodeling and deacetylase complex
Inhibits association of SUV39
H3K4me is associated
with active genes
from Turner, Cell 111, 285 (2002)

27. Nucleosomal Histones and Their Variants
from Sarma and Reinberg, Nature Rev.Mol.Cell Biol. 6, 139 (2005)

28. Role of H2A.Z H2A.Z is found on either side
of a nucleosome-free region
H2A.Z may protect promoters
from being methylated
from Talbert and Henikoff, Nature Rev.Mol.Cell Biol. 11, 264 (2010)

29. Action of HMG-box Proteins
HMG-box proteins bend DNA
DNA bending can affect transcription
and site-specific recombination
from Thomas and Travers, Trends Biochem.Sci. 26, 167 (2001)

30. Spatial Assembly of Expression Units
from Dekker, Science 319, 1793 (2008)

31. Regulation of DNA Accessibility
Nucleosome sliding exposes binding sites
Chromatin remodelling complexes
extract DNA from the nucleosome surface
Nucleosome eviction may be
necessary for transcription initiation
from Jiang and Pugh, Nature Rev.Genet. 10, 161 (2009)
Histone chaperones
incorporate histone variants