CHAPTER 10

CHROMOSOME ORGANIZATION AND MOLECULAR STRUCTURE

The length of DNA

46 chromosomes 6,000,000,000 6 000 000 000 (6 x 109) base pairs. pairs The linear distance between nucleotide pairs in ( ) DNA is 0.34 nanometers (0.34 x 10-9 meters) Total length= 6 x 109 X 0.34 x 10-9 = 2 m The diameter of the nucleus from a typical human somatic cell is about 5 x 10-6 meters.

The problem that had to be solved

with respect to the cell nucleus and the DNA it contains.

Introduction

1.Viral Genomes 2.Bacterial Chromosomes 3.Eukaryotic Chromosomes

10.1 VIRAL GENOMES

General Characteristics of Viruses
Small infectious particles containing nucleic acid surrounded by a capsid of proteins
Eith RNA or DNA, not both Either DNA t b th

Replication in host cells (obligate parasite) Most viruses exhibit a limited host range

General structure of viruses

Lipid bilayer Picked up when virus leaves host cell

Shrimp White Spot Syndrome Virus

Grouper iridovirus

Viral Genomes
Single Stranded DNA Double Stranded

Nucleic Acid
RNA

Double Stranded Positive Single Stranded Negative RNA DNA

Viral assembly

1.Self-assembly
(e.g.Tobacco (e g Tobacco mosaic virus)
Capsid protein

2. Directed assembly
(e.g. Phage T4)
Capsid composed of 2,130 2 130 identical protein subunits

T4 Bacteriophage DNA compaction

Cell 135, 12511262, December 26, 2008

10.2 BACTERIAL CHROMOSOMES

The looped structure compacts the chromosome about 10-fold Supercoiling within loops creates a more compact DNA

Looped chromosomal DNA

Supercoiled and looped DNA

Coil and Supercoil S il

Regulation of supercoiling
Competing action of these two enzymes 1. DNA gyrase (topoisomerase II)

2. DNA topoisomerase I

DNA containing one positive supercoil il

DNA gyrase

Composed of two A and two B subunits

DNA containing one negative supercoil p

The B subunits use ATP to catalyze this p process by the A subunits

Application Inhibition by antibiotics

1. 1 The coumarins (including novobiocin). novobiocin) 2. 2 The quinolones (including nalidixic acid and ciprofloxacin). ciprofloxacin)

Animation A i ti
Bacterial chormosome compaction Action of DNA gyrase
http://highered.mcgraw-hill.com/sites/0072835125/student_view0/index.html

10.3 EUKARYOTIC CHROMOSOMES

Has a genome that is more than twice as large as that of

Genome size (nucleotide base pairs per haploid genome

Organization of Eukaryotic Chromosomes

1. 1 Telomere 2. 2 Origin of Replication 3. Centromere

Eukaryotic Chromatin Compaction

There are three levels of chromatin organization: 1. DNA wrapping around nucleosomes: The "beads on a string structure beads string" structure. 2. A 30 nm condensed chromatin fiber: Nucleosome arrays in their most compact form 3. Higher level DNA packaging into the metaphase chromosome

Nucleosomes_shortens 7 fold
Vary in length between 20 to 100 bp, depending on species and cell type Diameter of the nucleosome

 Histone proteins are basic They contain many positively charged amino acids positively-charged Lysine and arginine These bind with the phosphates along the DNA backbone There are five types of histones H2A, H2B, H3 and H4 are the core histones Two of each make up the octamer H1 is the linker histone Binds to linker DNA Also binds to nucleosomes But not as tightly as are the core histones g y

Play a role in the organization and compaction of the ti f th chromosome

Figure 10.14

Testing the Hypothesis_

Nucleosome Structure
The experiment tests the beads-on-a-string model of chromatin structure
It the model is correct, DNase I should cut in the linker region
Thereby p oduc g DNA p eces t at a e about 200 bp long e eby producing pieces that are 00 o g

The rationale is that the linker DNA is more accessible than the core DNA to the DNase I

Figure 10.15

The Data

30 units ml-1

Interpreting the Data

These longer pieces were all in multiples of 200 bp At low concentrations, DNase I did not cut at all the linker DNA

And this, three

This fragment contains two nucleosomes

All chromosomal DNA digested into fragments that are ~ 200 bp in length

30 units ml-1

Nucleosomes Join to Form a 30 nm Fiber

Nucleosomes associate with each other to form a more compact structure termed the 30 nm fiber Histone H1 plays a role in this compaction p y p
At moderate salt concentrations, H1 is removed
The result is the classic beads-on-a-string morphology

At low salt concentrations, H1 remains bound

Beads associate together into a more compact morphology

Refer to Figure 10.16

Nucleosomes join to form a 30 nm fiber _shortens another 7 fold shortens

Regular, Regular spiral configuration containing six nucleosomes p per turn

Irregular configuration where nucleosomes have little face-to-face contact

Amino terminal tails of histone proteins

http://www.umanitoba.ca/institutes/manitoba_institute_cell_biology/MICB/Images/Davie_Figs/Intro_fig2.jpg

Further Compaction of the Chromosome Ch

The two events we have discussed so far have shortened the DNA about 50-fold A third level of compaction involves interaction between the 30 nm fiber and the nuclear matrix The nuclear matrix is composed of two parts
Nuclear lamina Internal matrix proteins
10 nm fiber and associated proteins fib d i t d t i

 The third mechanism of DNA compaction involves the formation of radial loop domains

MatrixAttachment Regions or Scaffoldattachment regions (SARs)

25,000 to 200,000 bp

MARs are anchored to the nuclear , g matrix, thus creating radial loops

Heterochromatin vs Euchromatin
The compaction level of interphase chromosomes p p is not completely uniform
Euchromatin
Less condensed regions of chromosomes Transcriptionally active Regions where 30 nm fiber forms radial loop domains g p

Heterochromatin
Ti htl compacted regions of chromosomes Tightly t d i f h Transcriptionally inactive (in general) Radial loop domains compacted even further

Heterochromatin vs Euchromatin.

http://www.bio.umass.edu/biology/conn.river/axarus.html

Figure 10.20

There are two types of heterochromatin

Constitutive heterochromatin
Regions that are always heterochromatic Permanently inactive with regard to transcription

Facultative heterochromatin
Regions that can interconvert between euchromatin and heterochromatin Example: Barr body

Figure 10.21

Compaction level in euchromatin During interphase most chromosomal regions are euchromatic Compaction level in heterochromatin

Figure 10.21

Metaphase Chromosomes
As cells enter M phase, the level of compaction p , p changes dramatically
By the end of prophase, sister chromatids are entirely heterochromatic In metaphase chromosomes the radial loops are highly compacted and stay anchored to a scaffold Histones are needed for the compaction of radial loops

These highly condensed metaphase chromosomes undergo little g g gene transcription p

 Two multiprotein complexes help to form and organize metaphase chromosomes Condensin
Plays a critical role in chromosome condensation

Cohesin
Plays a critical role in sister chromatid alignment

Both contain a category of proteins called SMC proteins

Acronym = Structural Maintenance of Chromosomes SMC proteins use energy from ATP and catalyze changes i chromosome structure h in h t t

The condensation of a metaphase chromosome by condensin

During interphase, condensin is in the cytoplasm

Condesin binds to chromosomes and compacts the radial loops

Condesin travels into the nucleus

The number of loops has not changed However, the diameter of each loop is smaller

Cohesins along chromosome arms are released

Cohesin at centromer is degraded

Cohesin remains at centromere

The alignment of sister chromatids via cohesin

Animation
Action of cohesin
http://highered.mcgraw-hill.com/sites/0072835125/student_view0/index.html http://highered mcgraw hill com/sites/0072835125/student view0/index html

Summary

1st l level l Beads on the string g

2nd level 30 nm fibers

During interphase most chromosomal regions are euchromatic Compaction level in euchromatin Compaction level in heterochromatin

3rd level Chromatin Ch ti fiber

Additional level

Metaphase chromosome

Application_ Premature Chromosome Condensation

Am J Hum Genet. 2002 April; 70(4): 10151022.

Chromosome territory

mouse fibroblast nucleus

24 chromosome territories in human cells

Vedio
DNA packaging
http://www.youtube.com/watch?v=AF2wwMReTf8&feature=related http://www.youtube.com/watch?v=OStI5pniHPA&feature=related

Supplementary Reading
Chromosome M h Ch Morphogenesis: C d i Condensin-Dependent C h i i D d t Cohesin Removal during Meiosis Cell, Volume 123, Issue 3, 4 November 2005, Pages 397-407 Hong-Guo Yu and Douglas Koshland

Mitochondrial disease, Lancet 2006; 368: 7082 Anthony H V Schapira