Chromatin Hierarchy

Question 0

Euchromatin vs. Heterochromatin

Answer 0

Euchromatin: less tightly packaged and transcriptionally active genes
Heterochromatin: much more condensed, contains transcriptional silent genes, found closer to nucleus membrane

Question 1

Nucleosome

Answer 1

• Packages of 200 bp DNA, plus protein.
• Fundamental unit of chromatin structure
• Beads on a string, beads = octamer of histone proteins
  -

Question 2

Histone Families

Answer 2

Histone H1 = linker between beads

H2A, H2B, H3 and H4 = core histones = the bead

Question 3

Histone Function

Answer 3

1. Structural: scaffold of chromosomes
2. Inhibit Transcription: bind DNA
   
   • modifications alter chromatin structure –> regulating transcription

Question 4

Histone Modifications - Inhibition

Answer 4

Rapid short term change involving:
Inhibition of Transcription
Phosphorylation of all histones –> condenses chromatin into chromosomes
Methylations of histone H3 –> heterochromatin caused by binding to non-histone protein HP1 (not transcribable)

Question 5

Histone Modifications - Activations

Answer 5

Activation of Transcription

- Acetylation via histone acetyl transferases (HATs)
	- K+ and R+ residues neutralized which repells histone ionic attraction from DNA
		- Forms euchromatin (transcribable)

Question 6

Location of Histone Modifications

Answer 6

specific aa residues located within the amino terminal tails of the core histones –> protrude from nucleosome surface

Question 7

Histone Modifying Enzymes effect on Chromosome Function

Answer 7

Impact chromosome function by

1. Altering electrostatic charge of core histones (acetylation)
2. Provide binding sites for transcription regulatory proteins (methylation: HP-1)

Question 8

Epigenetic Modifications

Answer 8

Produce heritable changes in gene function without a change in DNA sequence
- reversible changes
Example: DNA methylation blocks binding of transcription factors that normally activate gene expression –> repress gene expression

Question 9

Synergy of repressive histone modifications

Answer 9

Methyl binding proteins (MBPs)

1. recruit histone deacetylases (HDACs) - remove acetyl group so histones package DNA more tightly
2. recruit histone methyltransferases (HMTs) –> binding by chromodomain of HP-1 protein –> silence gene expression

Question 10

Other parts of the Nucleosome

Answer 10

Non-Histone Proteins - DNA/RNA polymerases, repair enzymes, nucleases
DNA: 140 bp DNA (2 loops of 70 bp each) wrap around each histone octamer, 60 bp DNA between each nucleosome (H1)

Question 11

Transcribing Chromatin

Answer 11

Contains nucleosomes even though histones inhibit transcription

no H1, 1-2 nucleosomes are absent but only from the gene’s promoter, nuclease assessable, histones must be acetylated

Question 12

Histone Acetyl Transferases (HATs)

Answer 12

Bind to transcription factors that activate specific genes

HAT proteins may integrate growth factor signals to differentiate 210 different cell types

Question 13

2nd Order of Chromatin Structure

Answer 13

30 nm Fiber
- solenoid
- Nucleosome aggregate as the 30 nm fiber
- H1 required to from structure
- genes are transcriptionally silent
- 6 nucleosome in fiber
	200 bp DNA x 6 nucleosomes = 1,200 bp

Question 14

3rd Order of Chromatin Structure: The Loop

Answer 14

• Fifty 1,200 bp 30 nm fibers form loops of chromatin containing ~ 60,000 bp DNA
  each loop can contain 1-several genes
• ## the base of each loop attaches to the nuclear matrix, which enables interaction of DNA domains between loops, which also facilitates gene expression (enhancers)

Question 15

Nuclear Matrix

Answer 15

Contains machinery for DNA replication and transcription, such as transcription factors and DNA and RNA polymerases

Question 16

4th Order of Chromatin Structure: The Miniband

Answer 16

Loops of chromatin wrap around the long axis of this mitotic chromosome. One loop = miniband

- Miniband contains 18 loops @ 60,000 bp/loop
- 1 million bp DNA in each mini band
- stack one atop another to form the chromosome

Question 17

5th Order of Chromatin Structure: The Chromosome

Answer 17

• Each chromosome has 75 million bp DNA (75 minibands)
• condensation of chromatin mediated by hyper-phosphorylation of chromatin proteins
• centromere: locus of repetitive DNA in the middle of the chromosome
• Telomere: loci of GGGTTA repeats that cap chromsome ends
  
  • function in protection, replication and stabilization of ends
  • some telomere is lost after each cell divison
  • when short enough, cell becomes senescent
  • germ line cells TELOMERASE maintains chromosome length = immortality
  • Some cancers exhibit cells that became immortal with gain of function mutation in telomerase

Question 18

Amount of DNA in Cell

Answer 18

75 million bp / chromosome
46 chromosomes/cell
=
3 billion bp/ cell!