Molecular biology

Eukaryotic chromatin and chromosome

Vedemy | Virendra Singh

 Overview

1. Replication Large Change

Small change
5.Transcription 4. Repair
Gene expression
Immature transcript Repaired DNA

6. Processing 9.Gene regulation

3’AAAAAA
5’CH3 functional
protein

7.Translation 8. PTM
mature transcript
Gene expression

Non functional protein

Chromosome
 Eukaryotic chromatin and chromosome

• Chromatin: DNA and protein structure.

• Chromatin vs. Chromosomes: Condensation difference.

• Origin of "Chromosome": Greek, color and body.

• Euchromatin vs. Heterochromatin: Light vs. dark.

• MAR (Matrix Associated Regions): A.T-rich DNA sequences.

Vedemy | Virendra Singh

 Eukaryotic chromatin and chromosome

• - E. coli chromosome: 4.64 million base pairs.

• - E. coli DNA: 1000x cell length.

• - Human cells: 6 billion base pairs.

• - Human DNA: 1.8 meters long.

• - Small chromosome: 14,000x nucleus length.

• - DNA tightly packed for compactness.

• - Genetic information transmission: centromere, spindle fibers.

• - DNA replication initiation: replication origins.

• - Chromosome end protection: telomeres.

Vedemy | Virendra Singh

 Eukaryotic chromatin and chromosome

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 Eukaryotic chromatin and chromosome

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 Eukaryotic chromatin and chromosome

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Euchromatin vs heterochromatin
Characteristic Euchromatin Heterochromatin
Chromatin condensation Less condensed More condensed
Location On chromosome arms At centromeres, telomeres, and
other specific places
staining Light Dark
Type of sequences Unique sequences Repeated sequences*
Presence of genes Many active genes Few active genes*
When replicated Throughout S phase (early) Late S phase
Transcription Often ( frequent) Infrequent (rare)
Crossing over Common Uncommon
Appearance Exist in extended state, dispersed 1. Constitutive ; always inactive
through the nucleus and staining and condensed.
diffusely. 2. Facultative; - Genetically
active(decondensed) and
inactive (condensed),
Ric region GC rich AT rich
Gene expression May or may not Poorly expressed
 Eukaryotic chromatin and chromosome

• Eukaryotic Chromatin Components: Centromere

• Telomere

• Origin of Replication

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 Centromere • Kinetochores assemble, holding sister chromatids.

• Kinetochore complex: spindle microtubules attach.

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 Centromere

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 Centromere

• Chromosome classification based on centromere position.

• Median centromere: arms nearly equal, V-shaped appearance.

• Submedian centromere: one arm shorter, L-shaped appearance.

• Subterminal centromere: one very short, one very long arm, rod-shaped appearance.

• Terminal centromere: centromere at the tip.

• Chromosome arms represented by p and q.

• Most chromosomes have a single kinetochore assembly site.

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Chromosome morphology
Telomere
• Telomeres: Cap ends of eukaryotic chromosomes.

• Functions: Maintain structural integrity, ensure complete replication.

• Composed of short, tandemly repeated sequences.

• 100–1000 repeats depending on the organism.

• G-rich single-strand 3' overhang, 50–300 nucleotides.

• Telomeric sequence highly conserved in evolution.

• Examples:

• TTGGGG (Paramecium),

• TAGGG (Trypanosoma),

• TTTAGGG (Arabidopsis),

• TTAGGG (Homo sapiens).

Vedemy | Virendra Singh
 Origin of replication

• Chromatins are multirepliconic.

• More than one replicon.

• Replicons typically 40 kb to 100 kb.

• Each replicon has its origin.

• One chromatin has multiple origins.

• Human chromatins: 30,000 origins.

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 Human karyotype

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Human karyotype
Autosomes are numbered from largest to smallest, except that chromosome 21 is smaller than chromosome 22.
Group Chromosomes
A 1–3
B 4,5
C 6–12,X
D 13–15
E 16–18
F 19,20
G 21,22,Y
Description
Largest; 1 and 3 are metacentric but 2 is submetacentric
Large; submetacentric with two arms very different in size
Medium size; submetacentric
Medium size; acrocentric with satellites
Small; 16 is metacentric but 17 and 18 are submetacentric
Small; metacentric
Small; acrocentric, with satellites on 21 and 22 but not on the Y
Chromosome Nomenclature
• - ISCN: International System for Human Cytogenetic Nomenclature.
• - Paris nomenclature: basic terminology for banded chromosomes.
• - Short arm: labeled p (petit).
• - Long arm: labeled q (queue).
• - Each arm divided into regions: p1, p2, p3, q1, q2, q3, etc.
• - Regions further divided into bands: p11, p12, p13, etc.
• - Sub-bands: p11.1, p11.2, etc.
• - Sub-sub-bands: e.g., p11.21, p11.22, etc.
 Regions bands and sub-bands

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 Metacentric Submetacentric Acrocentric
(Chromosome 9) (Chromosome 14)
(Chromosome 1)
 Chromosome banding
Technique Procedure Banding pattern

G-banding Mild proteolysis with trypsin followed by staining with Giemsa Dark bands are AT-rich (low gene density)
CG' stands for Giemsa). Light bands are GC-rich (high gene
density)

R-banding Thermal denaturation followed by staining with Giemsa. Dark bands are GC-rich
Reverse of G-banding and 'R' stands for Reverse. Light bands are At-rich

Q-banding Stain with Quinacrine mustard (a fluorescent stain). 'Q' stands Dark bands are At-rich
for Quinacrine. Light bands are GC- rich

C-banding Denaturation with barium hydroxide and then staining With Dark bands contain constitutive
Giemsa. 'C' stands for Constitutive heterochromatin. Heterochromatin

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 Nucleosome

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 Nucleosome
• Nucleosomes:
• Majority of eukaryotic DNA is packaged into nucleosomes.
• Each nucleosome consists of an octamer of histone proteins with DNA wrapped around it.
• The DNA between nucleosomes resembles a “string” in the “beads on a string” model.
• Nucleosomes compact DNA approximately sixfold.
• Tightly Associated DNA:
• The DNA most tightly associated with the nucleosome is called the “core particle”.
• It winds around the histone octamer like thread around a spool.
• Length: 147 base pairs (bp) (invariant across eukaryotic cells).
• Linker DNA:
• Between nucleosomes, there is variable linker DNA.
• Typically spans 20–60 bp.
• Each eukaryote has a characteristic average linker DNA length.

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 Nucleosome

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 Packaging of DNA into chromosomes

• Histones: Histone Molecular mass Lysine/Arginine

• Replication-dependent type (Dalton) content

H1 23,000 Lysine-rich
• H1 ( Linker) , H2A, H2B, H3, H4 (Core) H2A 14,000 Slightly lysine-rich

• synthesized during S-phase. H2B 13,800 Slightly lysine-rich

H3 15,300 Arginine-rich
• H3 and H4 are highly conserved.
H4 11,300 Arginine-rich

• Histone H1 shows variation between species.

• Replication-independent histones.

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 Packaging of DNA into chromosomes

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 Packaging of DNA into chromosomes

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Packaging of DNA into chromosomes

1. DNA Helix → 2nm

2. Beads on string → 11nm

3. Solenoid o Zig-Zag Fiber Model → 30nm

4. Loop Model → 300nm

5. Scaffold Model → 700nm

6. Chromosome → 1400nm
 11nm fiber and nucleosome organization

• Histones pack DNA into nucleosomes.

• Nucleosome is basic chromatin unit.

• Forms beads-on-a-string structure.

• Invariant component in chromatins.

• Nucleosome results in 11 nm fiber.

• Nucleosome shape: 11 nm and 6 nm.

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 11nm fiber and nucleosome organization

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 11nm fiber and nucleosome organization

• Nucleosome core has histone octamer.

• Octamer: 2 each of H2A, H2B, H3, H4.

• Core histones neutralize DNA charge.

• Core histones form barrel-shaped octamer.

• DNA wraps around octamer exterior.

• DNA wraps 1.65 turns in left-handed coil.

• Core histones have histone-fold domain.

• Each core histone has flexible N-terminal tail.

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 11nm Beads on string Model

• Nucleosome Structure:
• The nucleosome is not perfectly symmetrical but has an approximate twofold axis of symmetry called the dyad axis.
• Imagine the nucleosome face as a clock, with the midpoint of the 147 bp of DNA at the 12 o’clock position.
• The DNA ends are just short of 11 and 1 o’clock.
• Rotation of the nucleosome by 180° around the dyad axis reveals a nearly identical view.
11nm Beads on string Model

Nucleosome core
Histone octamer Linker Histone Linker DNA
1.65 turn wrap H1 Naked ( No histone)
146-147 bp 20 bp InterDNA between two octamer
33-34 bp
Chromatosome
147+20=167 bp
Nucleosome = 200 bp wrap
Beads on string Model
• Nucleosome contains about 200 base pairs of DNA.

• Two copies of each histone (H2A, H2B, H3, H4).

• Histone octamers not conserved during/after replication; but H32-H42 tetramers are conserved.

• DNA structure altered in the middle of nucleosome core particles, with increased base pairs per
turn.

• DNA forms a flat super helix with 1.65 turns around the histone octamer.

• Six nucleosomes per turn organize into two start helix for 30 nm fibers.

• N-terminal histone tails in a core particle are randomly ordered and protrude from the nucleosomes'
minor groove channels of DNA.

• Chromatin remodeling is ATP-dependent, regulated.

Beads on string Model
• H3.H4 tetramers

• interact with the central 60 bp of DNA within the nucleosome.

• The amino-terminal region of H3 forms a fourth alpha helix that interacts with the final 13
bp at each end of the bound DNA.

• occupies the top half of the histone octamer,

• binding the middle and both ends of the DNA.

• H2A.H2B dimers

• associate with approximately 30 bp of DNA

• on either side of the central 60 bp bound by H3 and H4.

• Together, the two H2A.H2B dimers form the bottom part of the histone octamer across the
disc from the DNA ends.
Beads on string Model
30nm Fiber Model
30nm Fiber Model

Regular, spiral
configuration
containing six
nucleosomes per
turn

Irregular configuration where

nucleosomes have little face-
to-face contact
30nm Fiber Model