DNA ORGANIZATION N STRUCTURE

• Basic Structure of DNA:

o DNA is composed of two polynucleo�de chains forming a double helix.

o Each chain consists of a sugar-phosphate backbone on the outside and nitrogenous

bases on the inside.

o The double helix structure results from the winding of these two strands around
each other.

• Base Posi�oning and Pairing:

o Nitrogenous bases are located on the interior of the helix, protected by the sugar-
phosphate backbone.

o Bases from opposite strands are held together by hydrogen bonds, forming base
pairs.

o The pairing rules follow strict complementarity: adenine (A) pairs with thymine (T),
and guanine (G) pairs with cytosine (C).

o A bulkier purine (two-ring structure, A or G) always pairs with a smaller pyrimidine

(one-ring structure, T or C), ensuring consistent width of the helix.

• Importance of Complementary Base Pairing:

o Complementary base pairing is essen�al for stability, as it enables the base pairs to
be packed in the most energe�cally favorable arrangement.

o This packing of bases ensures each base pair has a similar width, which holds the
sugar-phosphate backbones a constant distance apart.

o As a result, DNA maintains a uniform diameter essen�al for its double helical
structure.

• Energe�c Favorability and Stability:

o The complementary base pairs stack efficiently, maximizing stability within the helix
and protec�ng the gene�c informa�on.

o The specific pairing (A-T and G-C) also enables each DNA strand to be an exact
complement of the other, allowing accurate replica�on of gene�c informa�on.

2. Describe the helical structure of DNA, including the role of the sugar-phosphate backbone,
the an�parallel orienta�on of strands, and the structural consequences of base-pairing
rules.

• Right-Handed Double Helix Forma�on:

o DNA forms a right-handed double helix, where the two sugar-phosphate backbones
twist around each other.

o One full turn occurs every ten base pairs, providing a stable and compact structure.
 o The double helix allows DNA to store large amounts of gene�c informa�on in a
compact space while protec�ng it from environmental damage.

• An�parallel Orienta�on of Strands:

o The two DNA strands are oriented in opposite direc�ons, known as an�parallel
configura�on: one strand runs 5' to 3', while the other runs 3' to 5'.

o This an�parallel arrangement is essen�al for the correct alignment and bonding of
complementary bases within the helix.

o Only with opposite polarity can A pair with T, and G with C, allowing the double helix
to form and func�on properly.

• Impact of Base-Pairing on Helical Structure:

o The specific pairing of a purine with a pyrimidine keeps the helix width consistent,
crucial for the molecule's structural integrity.

o The base pairs stack in a way that stabilizes the structure, crea�ng an environment
that is both energe�cally favorable and protec�ve for the gene�c code.

o The consistent width along the helix prevents distor�on, which helps DNA maintain
its helical shape and allows accurate cellular processes like replica�on.

• Consequence for Gene�c Informa�on:

o Each strand's nucleo�de sequence is complementary to the other, meaning if one

strand’s sequence is known, the other can be deduced.

o This complementary nature ensures that each DNA strand can serve as a template
for replica�on, allowing cells to copy gene�c material precisely.

o The structural configura�on, including an�parallel orienta�on and base-pairing, is

vital for DNA's role in heredity and cellular func�ons.

1. Explain the significance of the double-helical structure of DNA in terms of gene�c

informa�on storage and replica�on.

• The double-helical structure of DNA, discovered by James Watson and Francis

Crick in 1953, is pivotal for understanding how gene�c informa�on is stored and
replicated.
• This structure consists of two long strands of nucleo�des that coil around each
other, forming a helix. Each nucleo�de is composed of a phosphate group, a
deoxyribose sugar, and one of four nitrogenous bases: adenine (A), thymine (T),
guanine (G), or cytosine (C). The specific pairing between bases—A with T and G
with C—creates complementary strands that are crucial for the replica�on
process.
• During DNA replica�on, the two strands of the double helix separate, facilitated
by enzymes such as helicase. Each strand then serves as a template for
synthesizing a new complementary strand through base-pairing rules.
 • This semi-conserva�ve method ensures that each new DNA molecule consists of
one original strand and one newly synthesized strand, thereby preserving the
gene�c informa�on across genera�ons.
• The precision of this process is vital for maintaining genomic integrity and
preven�ng muta�ons.
• Furthermore, the linear sequence of nucleo�des in DNA encodes the
instruc�ons necessary for synthesizing proteins, which are essen�al for cellular
structure and func�on.
• Genes, which are segments of DNA that contain these instruc�ons, can vary in
their nucleo�de sequences among different organisms.
• This varia�on underlies the biological diversity observed in nature. Thus, the
double-helical structure not only facilitates accurate replica�on but also serves
as a repository for the gene�c code that dictates an organism's traits.

2.Discuss how the gene�c code translates nucleo�de sequences into proteins and its implica�ons
for gene�c diversity.

• The gene�c code is a cri�cal mechanism that translates the linear sequences of nucleo�des
in DNA into func�onal proteins. Each gene contains a specific sequence of nucleo�des that
corresponds to a sequence of amino acids in a protein. The code is read in triplets called
codons, where each codon specifies a par�cular amino acid or serves as a stop signal during
protein synthesis.
• For instance, the codon AUG func�ons both as a start signal for transla�on and codes for the
amino acid methionine. This triplet nature of the gene�c code allows for 64 possible
combina�ons (4^3) from four nucleo�des, which correspond to 20 standard amino acids
used in protein synthesis. This redundancy in the code means that mul�ple codons can
specify the same amino acid, providing a buffer against muta�ons.
• The rela�onship between nucleo�de sequences and protein synthesis has profound
implica�ons for gene�c diversity. Varia�ons in nucleo�de sequences—due to muta�ons,
inser�ons, dele�ons, or recombina�on—can lead to changes in protein structure and
func�on. Such varia�ons can result in different phenotypes within a popula�on, influencing
traits such as resistance to diseases or adaptability to environmental changes. Over �me,
these varia�ons contribute to evolu�onary processes through natural selec�on.

Addi�onally, alterna�ve splicing allows a single gene to produce mul�ple protein isoforms by
rearranging exons during mRNA processing. This mechanism further enhances diversity without
requiring addi�onal gene�c material. Understanding this intricate rela�onship between DNA
sequences and protein synthesis is essen�al for fields like gene�cs, molecular biology, and
biotechnology.

3. Analyze the role of non-coding regions in DNA and their poten�al impact on gene
expression and regula�on.

Non-coding regions of DNA cons�tute a significant por�on of the genome—over 98% in

humans—and play vital roles beyond merely serving as templates for protein coding. These
regions include introns within genes and intergenic regions between genes, which were once
thought to be "junk DNA."

However, research has shown that non-coding regions are crucial for regula�ng gene
expression and maintaining genomic stability.
 Many non-coding sequences contain regulatory elements such as promoters, enhancers,
silencers, and insulators that control when and where genes are expressed. For example,
enhancers can be located far from their target genes yet interact with transcrip�on factors to
boost gene expression levels significantly.

Silencers serve to repress gene expression when bound by specific proteins. These regulatory
mechanisms allow cells to respond dynamically to internal signals and external
environmental changes.

Furthermore, non-coding RNAs (ncRNAs), including microRNAs (miRNAs) and long non-
coding RNAs (lncRNAs), have emerged as essen�al players in gene regula�on. miRNAs can
bind to messenger RNA (mRNA) molecules to inhibit their transla�on or promote
degrada�on, thereby fine-tuning protein produc�on levels.

LncRNAs can modulate chroma�n structure or interact with transcrip�on factors to influence
gene expression paterns.

The complexity introduced by non-coding regions underscores their importance in cellular

differen�a�on and development.

Varia�ons or muta�ons within these regions can lead to dysregula�on of gene expression
associated with various diseases, including cancer and gene�c disorders. Understanding the
func�ons of non-coding regions enhances our comprehension of gene�cs and opens
avenues for therapeu�c interven�ons targe�ng these regulatory elements.

• This is a 3D space-filling model showing 1.5 turns of the DNA double helix.

• Each turn of the helix includes about 10.4 nucleo�de pairs, which means that each complete
turn in the double helix has approximately 10-11 base pairs.

• Width of DNA: The distance from one side of the helix to the other is approximately 2
nanometers (nm).

• Grooves: The coiling of the two strands around each other creates two dis�nct grooves:
 o Major Groove: The wider groove in the helix, which is important because it allows
proteins and other molecules easier access to DNA bases for binding.

o Minor Groove: The narrower groove, which also serves as a binding site but has less
accessibility.

Part (B): Detailed View of DNA Strand Connec�ons

• Backbone Structure: The backbone of each DNA strand is composed of alterna�ng sugar and
phosphate groups.

• 5’ and 3’ Ends: Each strand has a direc�onality, indicated by the 5’ (5-prime) and 3’ (3-prime)
ends:

o The 5’ end typically has a phosphate group atached.

o The 3’ end usually has a hydroxyl (-OH) group atached to the sugar.

o This polarity is essen�al because it gives each DNA strand a direc�on, with the two
strands running in opposite (an�parallel) direc�ons.

• Base Pairing:

o Bases are located on the inside of the double helix and are connected by hydrogen
bonds.

o Adenine (A) pairs with Thymine (T) via two hydrogen bonds, while Guanine (G) pairs
with Cytosine (C) via three hydrogen bonds.

o This complementary base-pairing (A-T and G-C) allows the DNA strands to be packed
closely, with each base pair having a similar width to maintain the helical structure.

• Distance Between Nucleo�de Pairs: The center-to-center distance between adjacent base
pairs is 0.34 nanometers, allowing DNA to be densely packed within the helix.

Bases in DNA: In DNA, the purines( double-ring ) are adenine (A) and guanine (G).

Pairing:

• Adenine (A) pairs with thymine (T) through two hydrogen bonds.

• Guanine (G) pairs with cytosine (C) through three hydrogen bonds.

Posi�oning in the Helix: The larger size of purines is balanced by pairing with the smaller
pyrimidines, which helps maintain a consistent width across the DNA double helix.

Bases in DNA: In DNA, the pyrimidines ( single-ringed )are thymine (T) and cytosine (C).

Pairing:

• Thymine (T) pairs with adenine (A) through two hydrogen bonds.

• Cytosine (C) pairs with guanine (G) through three hydrogen bonds.

 Role in Maintaining DNA Structure: The pairing of a smaller pyrimidine with a larger purine keeps
the DNA double helix at a uniform diameter, as seen in the diagram.

DNA ORGANIZATION - EUKARYOTES

Discuss the organiza�on and func�onal significance of the eukaryo�c cell nucleus in DNA storage,
replica�on, and gene expression. In your answer, include the roles of the nuclear envelope, nuclear
pores, and the rela�onship between DNA, genes, and proteins.

Answer:

• Nucleus as DNA Storage Compartment:

o Nearly all DNA in a eukaryo�c cell is stored within the nucleus.

o The nucleus occupies about 10% of the total cell volume in many eukaryo�c cells.

o This compartmentaliza�on helps in organizing DNA and its associated processes,

such as replica�on and transcrip�on, in a controlled environment.

• Nuclear Envelope:

o The nucleus is surrounded by a nuclear envelope, which consists of two concentric

lipid bilayers.

o This envelope serves as a barrier, maintaining the nucleus as a separate

compartment from the cytoplasm.

o The nuclear envelope is directly connected to the endoplasmic re�culum (ER), a

network of membranes extending into the cytoplasm, facilita�ng communica�on
and material transfer.

• Nuclear Pores:

o The nuclear envelope contains nuclear pores, which are large openings allowing the
regulated exchange of molecules between the nucleus and the cytosol.

o These pores permit the entry of proteins necessary for DNA processes and the
export of RNA and ribosomal subunits synthesized within the nucleus.

• Nuclear Lamina:

o Beneath the inner membrane of the nuclear envelope lies the nuclear lamina, a
network of intermediate filaments.

o The lamina provides structural support to the nuclear envelope and plays a role in
organizing chroma�n and stabilizing the nucleus.

• DNA Replica�on:

o The double-stranded DNA serves as a template for replica�on, where each strand
pairs with complementary bases (A with T, and G with C).

o During replica�on, each parental DNA strand is used as a template to form two
iden�cal daughter DNA molecules, preserving the gene�c informa�on for cell
division.

• Gene Expression and Rela�onship to Proteins:

o DNA contains genes, which are specific sequences encoding proteins.

 o For example, the β-globin gene encodes a subunit of hemoglobin, a protein essen�al
for oxygen transport in the blood.

o The gene�c code in DNA is transcribed into messenger RNA (mRNA), which is then
translated into proteins.

o Each gene carries instruc�ons for synthesizing a par�cular protein, as exemplified by

genes A, B, and C, which produce different proteins involved in cellular func�ons.

• Func�onal Compartmentaliza�on:

o The nuclear envelope separates nuclear enzymes (ac�ng on DNA) from cytosolic
enzymes, crucial for proper cell func�oning.

o This organiza�on allows the cell to regulate gene expression and DNA processes
efficiently, maintaining cellular integrity and specialized func�ons of eukaryo�c cells.

Explain the structure and packaging of DNA in eukaryo�c cells. How is it possible for such a large
amount of DNA to fit inside the small nuclear space, and how does the organiza�on allow
accessibility for cellular processes like replica�on and transcrip�on?

Answer:

• Func�on of DNA:

o The primary func�on of DNA is to carry genes, which contain the instruc�ons for
producing RNA molecules and proteins that are crucial for all cellular func�ons.

o DNA also provides informa�on on when, where, and in what quan��es specific RNAs
and proteins are to be produced, helping regulate cellular processes.

• Chromosomal Structure:

o In eukaryo�c cells, nuclear DNA is divided into mul�ple chromosomes.

o Each chromosome contains a unique arrangement of genes, as well as specialized

sequences necessary for DNA replica�on and transmission during cell division.

• Challenges of DNA Packaging:

o In humans, the total length of DNA in a single cell, if stretched out, would reach
about 2 meters, yet it is stored in a nucleus only about 6 μm in diameter.

o This is analogous to compac�ng 40 km (24 miles) of thin thread into a tennis ball,
represen�ng a massive spa�al challenge.

• Role of DNA Packaging Proteins:

o DNA is packaged by various proteins, primarily histones, that bind to DNA and form
nucleosomes, which are the basic units of DNA packaging.

o These proteins help fold the DNA into organized coils and loops, crea�ng mul�ple
levels of compact structure that prevent DNA tangling and damage.

• Hierarchical Levels of DNA Organiza�on:

 o DNA wraps around histone proteins, forming nucleosomes, which further coil into
chroma�n fibers.

o These fibers are looped and folded into higher-order structures, allowing efficient
packaging within the nucleus.

• Accessibility of Compacted DNA:

o Even though DNA is �ghtly packed, it remains accessible to enzymes involved in

replica�on, repair, and transcrip�on.

o The dynamic structure of chroma�n allows certain regions to be unpacked

temporarily, gran�ng access to specific genes when needed.

Ques�on 2:

Discuss the concept of homologous chromosomes and the methods used to dis�nguish individual
chromosomes. How are chromosomes visualized, and what techniques have been developed to
iden�fy specific chromosomes in humans?

Answer:

Chromosomes carry genes—the func�onal units of heredity. A gene is o�en defined as a segment of
DNA that contains the instruc�ons for making a par�cular protein (or a set of closely related
proteins).

• Homologous Chromosomes:

o Eukaryo�c cells contain homologous chromosome pairs, with one chromosome

inherited from each parent.

o In humans, there are 46 chromosomes, organized into 22 pairs of autosomes

(common to both males and females) and one pair of sex chromosomes.

o Sex Chromosomes: Females have two X chromosomes, while males have one X and
one Y chromosome, which are the only non-homologous chromosomes in a male.

• Chromosome Visualiza�on Techniques:

o Chromosome Pain�ng:

 Chromosome pain�ng uses DNA hybridiza�on with fluorescent probes,

which are short strands of nucleic acids tagged with fluorescent dyes.

 These probes bind to specific DNA sequences on chromosomes, allowing

each chromosome to be "painted" a unique color.

 This method is o�en performed during mitosis, when chromosomes are

most compact and easiest to observe.

o Banding Paterns:

 Tradi�onal chromosome iden�fica�on involves staining techniques that

produce dis�nct banding paterns on mito�c chromosomes.
  These paterns are unique to each chromosome and provide a way to
iden�fy and number chromosomes based on their appearance.

 Banding paterns likely reflect differences in chroma�n structure, though the

exact basis is not fully understood.

• Significance of Chromosome Iden�fica�on:

o Chromosome visualiza�on techniques are essen�al for studying gene�c disorders,

karyotyping, and understanding chromosomal abnormali�es.

o Chromosome pain�ng and banding paterns allow researchers to iden�fy structural

differences, such as dele�ons, duplica�ons, or transloca�ons, that may contribute to
diseases.

o The display of the 46 human chromosomes at mitosis is called the human karyotype.
If parts of chromosomes are lost or are switched between chromosomes, these
changes can be detected either by changes in the banding paterns or—with greater
sensi�vity—by changes in the patern of chromosome pain�ng

o Cytogene�cists use these altera�ons to detect inherited chromosome abnormali�es

and to reveal the chromosome rearrangements that occur in cancer cells as they
progress to malignancy

FEATURES OF HUMAN GENOME

• Chromosome Overview: Humans have 22 autosomes and 2 sex chromosomes, along with
mitochondrial DNA. The nuclear genome contains 3.2 billion nucleo�de pairs in total,
distributed across these chromosomes.

• Example: Chromosome 22: This chromosome, one of the smallest, has approximately 48
million nucleo�de pairs, represen�ng only 1.5% of the en�re genome. It includes both
coding (exons) and noncoding (introns) regions, as well as repe��ve DNA sequences.

• Mitochondrial DNA: Each cell also has mitochondrial DNA (~16,569 nucleo�de pairs),
contribu�ng essen�al genes involved in cellular energy produc�on.

2. Gene Structure and Coding Regions:

• Exons and Introns: Only about 1.5% of the human genome consists of exons, which are DNA
sequences that directly code for proteins. The remaining segments within genes are introns,
which are noncoding regions.

• Size and Complexity of Genes: The average human gene is about 27,000 nucleo�de pairs
long, though only around 1,300 pairs are required for coding an average-sized protein. This is
because genes o�en contain long strings of introns, interrup�ng the coding exons.

• Protein-Coding Genes: The human genome has roughly 21,000 genes that code for proteins.
However, the presence of noncoding DNA and regulatory sequences makes each gene larger
and more complex compared to organisms with smaller, more compact genomes.
 • Gene Organiza�on in Different Species:
Unlike many simpler organisms, most human
genes are interspersed with introns, which
makes our genes significantly longer. Genes
in organisms with smaller genomes tend to
lack introns, allowing for more compact,
con�nuous coding regions.

3. Repe��ve and Noncoding DNA:

• Repe��ve DNA Elements: Nearly 50% of the

human genome consists of repe��ve DNA.
Many of these repe��ve sequences are
"transposable elements," which are mobile
DNA pieces that can move to different
genome loca�ons. These elements, over millions of years, have inserted themselves
repeatedly across chromosomes.

• Pseudogenes: The genome contains over 20,000 pseudogenes—segments that resemble

func�onal genes but have accumulated muta�ons that prevent them from producing
func�onal proteins.

• Func�on of Repe��ve Elements: While these repe��ve sequences were once considered
"junk," they are now understood to have roles in genome stability, regula�on, and evolu�on.

4. Noncoding RNA Genes and Regulatory DNA:

• Noncoding RNA Genes: Besides protein-coding genes, the human genome has
approximately 9,000 genes that produce noncoding RNAs. These RNAs are essen�al for
various cellular processes, even though they don’t make proteins. They play roles in gene
regula�on, RNA processing, and maintaining chromosome structure.

• Regulatory DNA Sequences: Each protein-coding gene is associated with regulatory DNA
sequences. These sequences determine when, where, and how strongly a gene is expressed.
In humans, regulatory regions are spread out over tens of thousands of nucleo�de pairs,
unlike more compact organisms where regulatory sequences are closer and more
concentrated.

5. Genomic Complexity and "Junk" DNA:

• “Junk” DNA: Much of the human genome appears to be clutered with non-func�onal or
"junk" DNA, which doesn’t directly code for proteins or regulate gene ac�vity. However, this
“junk” plays a role in maintaining genome integrity and may contribute to evolu�onary
diversity.

• Organiza�onal Complexity: Unlike highly organized genomes in simpler organisms, the

human genome is a patchwork of coding and noncoding sequences. Some researchers have
 compared it to a clutered storage space, containing valuable items (essen�al genes and
regulatory regions) mixed with seemingly disorganized elements.

• Evolu�onary Role: The human genome's complexity, with its regulatory sequences and
noncoding elements, likely evolved through gene duplica�on and the accumula�on of non-
func�onal DNA. This structure allows humans to maintain gene�c diversity and develop
complex traits.

Cell Cycle

1. Interphase

o Gene Expression and Chromosome Replication: Chromosomes are replicated,

forming two sister chroma�ds for each chromosome.

o Chromatin Structure: Chromosomes remain extended and are not easily

dis�nguishable as they exist as long threads within the nucleus.

2. Mitosis Ini�a�on

o Chromosome Condensation: Chromosomes condense, making the sister chroma�ds

visible as dis�nct structures.

3. Mito�c Chromosomes Forma�on

o Distinct Units: Chromosomes operate as individual units, each capable of replica�ng

and segrega�ng correctly.

4. Chromosome Replica�on Requirements

o Specialized Nucleotide Sequences: Three key sequences in DNA ensure proper

replica�on and segrega�on:

 Replica�on Origins: Mul�ple origins per chromosome to enable rapid and

complete replica�on.

 Centromere: Atachment point for sister chroma�ds, essen�al for their

separa�on.

 Telomeres: Repeated sequences at chromosome ends that allow complete

replica�on and protect chromosome integrity.

5. Kinetochore Forma�on

o Centromere Attachment: A kinetochore forms at each centromere, linking

chromosomes to the mito�c spindle.

6. Chroma�d Segrega�on
 o Mitotic Spindle Action: Duplicated chromosomes are pulled apart, with one copy of
each chroma�d moving to opposite daughter cells.

7. Cell Division Comple�on

o Daughter Cell Formation: Each daughter cell receives an iden�cal set of

chromosomes, ensuring gene�c con�nuity.

If the 48 million nucleo�de pairs of DNA in human chromosome 22 could be laid out as one long
perfect double helix, the molecule would extend for about 1.5 cm if stretched out end to end. But
chromosome 22 measures only about 2 μm in length in mitosis

Represen�ng an end to-end compac�on ra�o of over 7000-fold. This remarkable feat of
compression is performed by proteins that successively coil and fold the DNA into higher and higher
levels of organiza�on.

Although much less condensed than mito�c chromosomes, the DNA of human interphase
chromosomes is s�ll �ghtly packed.
Chroma�n and Chromosome Structure

1. Chroma�n Composi�on:

o Eukaryo�c chromosomes consist of DNA bound to

proteins, primarily divided into two classes: histones
and non-histone chromosomal proteins.

o The combined mass of both types of proteins is

roughly equal to the mass of DNA within a
chromosome. Together, these proteins and the
nuclear DNA form chroma�n.

2. Histones and Nucleosomes:

o Histones play a cri�cal role in the ini�al stage of DNA

packaging by forming structures called nucleosomes.

o The nucleosome structure was first iden�fied in 1974

and is the most basic unit of chromosome packing.

o Under an electron microscope, chroma�n appears as

a 30 nm fiber in interphase, represen�ng its
condensed structure. When par�ally unfolded,
chroma�n takes on a “beads-on-a-string”
appearance, where each “bead” is a nucleosome
core par�cle.

3. Nucleosome Structure:

o A nucleosome core par�cle consists of DNA wrapped

around a histone core. Each histone core contains eight histone proteins (two each
of H2A, H2B, H3, and H4).

o The DNA wound around this histone core measures 147 nucleo�de pairs in length.

o Between each nucleosome, there is a linker DNA segment of varying length,

averaging about 200 nucleo�de pairs per nucleosome.

4. Role in Chroma�n Compac�on:

o The forma�on of nucleosomes reduces the DNA’s length to about one-third of its
original size. For example, a diploid human cell has around 6.4 billion nucleo�de
pairs, requiring approximately 30 million nucleosomes.
5. Nucleosome Core Par�cle Structure:

o High-resolu�on studies (e.g., X-ray diffrac�on) revealed a disc-shaped histone core

around which DNA coils 1.7 �mes
in a le�-handed turn.

o Each of the core histones (H2A,

H2B, H3, and H4) contains a
histone fold formed by three α-
helices connected by two loops.
In the nucleosome, histones
interact to form H3–H4 and H2A–
H2B dimers. Two H2A–H2B
dimers and one H3–H4 tetramer
combine to form the histone
octamer around which DNA
wraps.

6. DNA-Histone Interac�ons:

o There are 142 hydrogen bonds

between the DNA and the
histone core, mainly between the
histone’s amino acid backbone
and the DNA’s sugar-phosphate backbone.

o These bonds, along with hydrophobic interac�ons and salt linkages, provide a stable
associa�on between DNA and histones.

o The core histones are rich in lysine and arginine residues, which are posi�vely
charged and help neutralize the nega�vely charged DNA backbone.

7. Histone Conserva�on and Variants:

o Histones are highly conserved across eukaryotes due to their essen�al role in DNA
func�on. For instance, the histone H4 protein differs by only two amino acids
between a pea plant and a cow.
 o Eukaryotes also produce variant histones with small amino acid differences from the
core histones. These variants, combined with histone modifica�ons, create diverse
chroma�n structures.

8. Histone Tails and Modifica�ons:

o Each core histone has an N-terminal tail that extends outward from the nucleosome.
These tails undergo covalent modifica�ons (e.g., acetyla�on, methyla�on),
influencing chroma�n structure and gene regula�on.

Chroma�n Dynamics and Remodeling

1. Nucleosome Stability and Dynamics:

o Contrary to earlier beliefs, nucleosomes are not permanently fixed on DNA. Instead,
they exhibit dynamic behavior, allowing par�al unwrapping for 10–50 milliseconds.
This periodic unwrapping permits temporary DNA exposure for interac�ons with
other proteins.

2. ATP-Dependent Chroma�n Remodeling Complexes:

o Cells contain various ATP-dependent chroma�n remodeling complexes that can

alter nucleosome posi�ons. These complexes use ATP to slide nucleosomes along
DNA, temporarily loosening DNA-histone interac�ons.

o Some remodeling complexes, with the aid of histone chaperones, can replace
histone subunits or completely remove histone octamers from DNA.

3. Chroma�n Remodeling and Gene Accessibility:

o Chroma�n remodeling complexes play an essen�al role in regula�ng gene

expression by reposi�oning nucleosomes, thereby exposing or hiding DNA regions
from transcrip�on machinery.

o The remodeling complexes are o�en localized to specific DNA regions to facilitate
transcrip�onal or replica�on processes, ensuring that chroma�n structure
dynamically adapts to cellular needs.

4. Nucleosome Posi�oning Factors:

o Besides DNA sequence preference, other �ghtly bound proteins on DNA can
influence nucleosome posi�oning by ac�ng as obstacles or recruitment factors.

o As a result, nucleosome posi�ons are o�en flexible and adaptable, allowing for the
con�nuous reorganiza�on of chroma�n in response to cellular signaling and
func�onal requirements.
 1. Chroma�n Remodeling Complex:

o In this image, the ATP-

dependent chroma�n
remodeling complex (shown
in green) interacts with the
nucleosome, which consists of
DNA (red) wrapped around a
histone core (yellow).

2. Role of ATP in Nucleosome Sliding:

o The complex uses the energy

from ATP hydrolysis to
facilitate nucleosome
movement along the DNA
strand.

o ATP hydrolysis is a process

where ATP (adenosine
triphosphate) is broken down
into ADP (adenosine
diphosphate) and inorganic phosphate (PiP_iPi), releasing energy.

o This energy allows the chroma�n remodeling complex to "push" on the DNA,
loosening its atachment to the histone core.

3. Cycle of ATP Binding and Hydrolysis:

o Each cycle begins with the binding of ATP to the remodeling complex, followed by its
hydrolysis, which releases energy.

o A�er ATP is converted to ADP and PiP_iPi, the products are released, and the DNA is
shi�ed slightly with respect to the histone core.

o Mul�ple cycles of ATP binding, hydrolysis, and product release are required to slide
the nucleosome along the DNA in the direc�on indicated by the arrow.

Part B: Structure of the ATPase Subunit

1. ATPase Subunits in Chroma�n Remodeling:

o This panel shows a detailed structure of the ATPase subunits involved in the ISWI
(Imita�on SWItch) family of chroma�n remodeling complexes.

o The ATPase subunits, shown in green, bind to the nucleosome and generate the
force necessary to slide it back and forth along the DNA.

o The ATPase subunit exists as a dimer (two iden�cal units), which provides stability
and ensures efficient sliding of the nucleosome.

2. Func�on of the ISWI Complex:

o The ISWI family is one of several families of chroma�n remodeling complexes,

specifically involved in reposi�oning nucleosomes to regulate DNA accessibility.
 o Through ATP hydrolysis, this dimer structure moves the nucleosome along the DNA
strand, which can help expose or conceal DNA sequences, depending on cellular
needs.

Part C: Structure of a Larger Chroma�n Remodeling Complex

1. Large Remodeling Complex Structure:

o This image shows the structure of a larger chroma�n remodeling complex, known as
the RSC complex (Remodel the Structure of Chroma�n).

o The green model represents the yeast RSC complex, which consists of 15 subunits.

2. Components and Func�ons:

o Among these 15 subunits, the complex includes an ATPase and at least four subunits
with domains that specifically recognize covalently modified histones.

o These modifica�ons (e.g., methyla�on, acetyla�on) serve as signals that help direct
the chroma�n remodeling complex to par�cular regions of the genome.

o The complex is structured in a way that allows it to wrap around a nucleosome,

posi�oning it to exert force and slide the nucleosome along the DNA effec�vely.

3. Scale:

o A scale bar indica�ng 10 nm (nanometers) helps give an idea of the size of the RSC
complex rela�ve to the nucleosome. This scale shows that the remodeling complex is
quite large, able to cover and interact with the nucleosome effec�vely.

Nucleosome removal and histone exchange are catalyzed by ATP-dependent chroma�n remodeling
complexes. These processes are crucial for modifying chroma�n structure, allowing for the
regula�on of gene expression, DNA repair, and other DNA-based processes. The image illustrates two
main ac�ons of chroma�n remodeling complexes: the exchange of histone dimers (top series) and
the removal or replacement of the en�re nucleosome core (botom series).
Top Series: Exchange of H2A-H2B Dimers

1. ATP-Dependent Chroma�n Remodeling Complex:

o The green structure represents the ATP-dependent chroma�n remodeling complex,

which binds to the nucleosome.

o This complex uses ATP to modify the nucleosome structure, allowing for the
exchange of specific histone components.

2. Histone Chaperone:

o Histone chaperones (shown as two-part structures in orange and blue) are proteins
that assist in the removal or replacement of histones without allowing the DNA to
tangle or degrade.

o In this case, the histone chaperone facilitates the removal of H2A-H2B histone
dimers (a type of histone pair in the nucleosome) from the nucleosome structure.

3. ATP Hydrolysis and Dimer Exchange:

o As in nucleosome sliding, the chroma�n remodeling complex requires ATP hydrolysis

(conversion of ATP to ADP and PiP_iPi) to provide energy.

o With this energy, the H2A-H2B dimer is removed, and a new variant histone dimer
can be inserted in its place. For example, a variant like H2A.Z-H2B may replace the
standard H2A-H2B dimer, altering the proper�es of the nucleosome.

o This exchange of dimers can affect the nucleosome’s interac�on with other proteins
and poten�ally influence gene expression.

Botom Series: Exchange of the En�re Nucleosome Core

1. Complete Removal of Nucleosome Core:

o In the botom process, the ATP-dependent chroma�n remodeling complex works

with histone chaperones to remove the en�re nucleosome core, rather than just
individual histone dimers.

o This involves the removal of the histone octamer, which consists of all four core
histones (H2A, H2B, H3, and H4).

2. DNA Lacking Nucleosome:

o Once the histone octamer is removed, the DNA is le� in an open or “nucleosome-
free” state, which makes it more accessible to transcrip�on factors, repair enzymes,
or replica�on machinery.

o This removal can be essen�al for processes that require direct access to DNA, like
transcrip�onal ac�va�on of certain genes.

3. Replacement with a New Nucleosome Core:

 o A�er removal, a new nucleosome core can be reassembled, either with the same
histone types or with modified or variant histones, depending on the needs of the
cell.

o For this to happen, ATP hydrolysis occurs again, allowing the remodeling complex to
cooperate with histone chaperones to assemble a new histone octamer onto the
DNA.

4. Exchange of Nucleosome Core:

o This reassembly can involve variant histones, which have different proper�es from
the original histones and can impact chroma�n structure and gene regula�on.

o The exchange of the nucleosome core allows cells to modify chroma�n in response
to various signals or environmental condi�ons, providing a dynamic means of
regula�ng DNA accessibility.

Summary

In summary, this figure illustrates two ATP-dependent processes facilitated by chroma�n remodeling
complexes:

1. Histone Dimer Exchange (top): Removal and replacement of specific histone pairs like H2A-
H2B with variants, modifying nucleosome composi�on.

2. Nucleosome Core Exchange (botom): Complete removal of the nucleosome core, crea�ng
an open DNA region, and reassembly with possibly new or modified histones.

These mechanisms allow cells to dynamically alter chroma�n structure, influencing DNA accessibility
and enabling precise control over cellular processes like transcrip�on, repair, and replica�on.

What causes nucleosomes to stack so �ghtly on each other?

1. Nucleosome-to- nucleosome linkages that involve histone tails, most notably the H4 tail,
cons�tute one important factor.

2. Another important factor is an addi�onal histone that is o�en present in a 1-to-1 ra�o with
nucleosome cores, known as histone H1. This so-called linker histone is larger than the
individual core histones and it has been considerably less well conserved during evolu�on.

3. A single his tone H1 molecule binds to each nucleosome, contac�ng both DNA and protein,
and changing the path of the DNA as it exits from the nucleosome.

4. This change in the exit path of DNA is thought to help compact nucleosomal DNA
CHROMATIN STRUCTURE AND FUNCTION

Certain types of chroma�n structure can be inherited; that is, the structure can be directly passed
down from a cell to its descendants. Because the cell memory that results is based on an inherited
chroma�n structure rather than on a change in DNA sequence, this is a form of epigene�c
inheritance. The prefix epi is Greek for “on”; this is appropriate, because epigene�cs represents a
form of inheritance that is superimposed on the gene�c inheritance based on DNA.

Chroma�n Types Iden�fied in Light Microscopy (1930s)

• Chroma�n: DNA-protein complex found in eukaryo�c cells; helps in DNA packaging and gene
regula�on.

• Types: Two types of chroma�n were iden�fied in interphase nuclei:

1. Heterochroma�n – Highly condensed form.

2. Euchroma�n – Less condensed form, represen�ng all other chroma�n.

Heterochroma�n

• Defini�on: A compact and densely packed form of chroma�n.

• Appearance and Loca�on:

o Easily dis�nguishable due to its condensed nature.

o Found primarily in specialized chromosome regions:

 Centromeres: Essen�al for chromosome separa�on during cell division.

 Telomeres: Protec�ve ends of chromosomes.

 o Also present in various other loca�ons across chromosomes; loca�on can change
depending on the cell’s physiological state.

• Genome Composi�on:

o In mammalian cells, more than 10% of the genome is packaged as heterochroma�n.

• Gene Content and Expression:

o Typically contains few genes.

o When regions of euchroma�n (less condensed) are transformed into

heterochroma�n, genes in those areas are usually silenced (turned off).

• Func�onal Implica�ons:

o Not "Dead" DNA: Originally thought to be inert or inac�ve DNA, heterochroma�n is

now understood to contain dis�nct modes of chroma�n compac�on.

o Resistance to Gene Expression: The defining feature of heterochroma�n is its

resistance to gene expression, making it less accessible for transcrip�on.

o Different forms of heterochroma�n may have varying effects on gene regula�on and
expression, sugges�ng its role is more complex than just silencing DNA.

Euchroma�n

• Defini�on: A less condensed form of chroma�n compared to heterochroma�n.

• Characteris�cs:

o Euchroma�n is generally associated with ac�ve gene expression because its

structure is more open and accessible.

o It contains most of the ac�vely transcribed genes.

• Dynamic Transi�on:

o Euchroma�n can transi�on into heterochroma�n based on cellular condi�ons and

signals, resul�ng in gene silencing for specific regions when required.

Through chromosome breakage and rejoining, whether brought about by a natural gene�c accident
or by experimental ar�fice, a piece of chromosome that is normally euchroma�c can be translocated
into the neighborhood of heterochroma�n.

Remarkably, this o�en causes silencing—inac�va�on—of the normally ac�ve genes. This
phenomenon is referred to as a posi�on effect.

It reflects a spreading of the heterochroma�c state into the originally euchroma�c region, and it has
provided important clues to the mechanisms that create and maintain heterochroma�n. First
recognized in Drosophila, posi�on effects have now been observed in many eukaryotes, including
yeasts, plants, and humans.
In histones, lysine residues are primarily found on the
N-terminal "tails" that extend outward from the
nucleosome core. These tails are unstructured and
highly accessible, which makes them key targets for
various covalent modifica�ons.

Types of Histone Modifica�ons

• Histone tails undergo various covalent

modifica�ons:

o Acetyla�on: Addi�on of an acetyl

group to lysine residues, removing the
posi�ve charge and reducing atrac�on between nucleosomes, resul�ng in a
loosened chroma�n structure.

o Methyla�on: Lysines can be mono-, di-, or trimethylated. Methyla�on does not

change the charge of the histone but provides binding sites for specific proteins,
impac�ng gene expression.

o Phosphoryla�on: Adds a phosphate group to serines or threonines, introducing a

nega�ve charge that alters chroma�n structure and func�on.

o Other Modifica�ons: Include ADP-ribosyla�on, ubiquityla�on, sumoyla�on, and

bio�nyla�on. Each modifica�on affects chroma�n dynamics and gene expression
differently.

Enzymes Involved in Modifica�ons

• Modifica�ons are reversible:

o Histone Acetyltransferases (HATs): Add acetyl groups to lysines.

o Histone Deacetylases (HDACs): Remove acetyl groups from lysines.

o Histone Methyltransferases: Add methyl groups to lysines.

o Histone Demethylases: Remove methyl groups from lysines.

• Enzymes are highly specific and recruited to specific sites on chroma�n by transcrip�on
factors, which recognize DNA sequences and recruit these enzymes based on the cell's
needs.

Func�ons of Histone Modifica�ons

• Modifica�ons help regulate gene expression and chroma�n structure:

o Acetyla�on of Lysines: Loosens chroma�n structure, facilita�ng gene expression by

reducing nucleosome affinity.

o Trimethyla�on of H3 Lysine 9 (H3K9me3): Atracts the protein HP1, establishing

heterochroma�n, which silences gene expression.

• The combina�on of histone modifica�ons determines chroma�n func�on, influencing gene

expression, DNA repair, and replica�on.
Histone Code Hypothesis

• Histone Code: Hypothesis that specific combina�ons of histone modifica�ons serve as codes
that influence chroma�n behavior, including gene expression and cellular processes.

• Different paterns of modifica�ons on nucleosomes can signal various cellular processes,

such as:

o DNA Replica�on: Marked chroma�n is newly replicated.

o DNA Damage: Certain modifica�ons indicate damaged DNA, triggering repair

mechanisms.

o Gene Expression Regula�on: Modifica�ons either promote or repress transcrip�on

based on specific combina�ons.

 Histone Variants and Chroma�n Structure

• Histone Variants: In addi�on to the standard histones (H2A, H2B, H3, H4), eukaryo�c cells
contain histone variants, each with specific func�ons:

o H3.3: Linked to transcrip�onal ac�va�on.

o CENP-A: Important for centromere func�on and kinetochore assembly.

o H2AX: Plays a role in DNA repair and recombina�on.

o H2AZ: Involved in gene expression and chromosome segrega�on.

o MacroH2A: Linked to transcrip�onal repression and X-chromosome inac�va�on.

• Histone variants are inserted into nucleosomes through ATP-dependent chroma�n

remodeling complexes in a site-specific manner, altering chroma�n structure and func�on.

Dynamic Nature of Histone Modifica�ons

• Modifica�ons are reversible and frequently adjusted based on cellular condi�ons.

• Histone Tails: The unstructured and accessible nature of histone tails makes them suitable
for dynamic modifica�ons, allowing rapid responses to cellular needs.

• Modifica�ons can be maintained even a�er the ini�al recrui�ng factors (like transcrip�on
regulators) have le�, providing a memory of cellular history that can be passed on during cell
division.

 Reader Complexes and the Interpreta�on of Histone Marks

• Reader Proteins: Recognize and bind specific histone modifica�ons, forming reader
complexes that interpret histone marks.

• Examples:

o PHD Domains: Recognize and bind trimethylated lysines on histones, with specific
PHD domains binding to different methyla�on paterns.

• Reader Complexes enable histone marks to recruit addi�onal proteins to execute specific
biological func�ons at the right �me, such as ac�va�ng or repressing gene transcrip�on.
 The enzymes that add or remove modifica�ons to histones in nucleosomes are part of mul� subunit
complexes. They can ini�ally be brought to a par�cular region of chroma�n by one of the sequence-
specific DNA-binding proteins (transcrip�on regulators)

Posi�on Effect Variega�on (PEV)

• PEV describes how chroma�n modifica�ons can spread along a chromosome, affec�ng gene
expression.

• PEV can lead to gene silencing when a gene is near heterochroma�n (dense, inac�ve
chroma�n) due to chroma�n modifica�ons spreading.

Chroma�n Modifica�ons and Spreading

1. Writer and Reader Enzymes:

o Writer enzymes add chemical modifica�ons to histones (e.g., methyla�on,

acetyla�on).

o Reader proteins recognize these modifica�ons and bind to modified nucleosomes,

poten�ally recrui�ng addi�onal proteins.

o Together, reader-writer complexes can spread modifica�ons from one nucleosome

to the next, crea�ng a "spreading wave" along the DNA.

2. Spreading Mechanism:
 o A transcrip�on regulator directs the writer enzyme to a specific DNA site.

o The writer places a mark on histones, which the reader recognizes, binding to the
mark.

o This binding posi�ons the writer near adjacent nucleosomes, allowing the mark to
spread.

o The spreading process can condense chroma�n, making it transcrip�onally inac�ve

(e.g., heterochroma�n).

3. Types of Modifica�ons:

o Methyla�on and acetyla�on are key histone modifica�ons.

o Examples:

 H3K9 trimethyla�on atracts HP1 protein, which leads to heterochroma�n

forma�on and gene silencing.

 H3K4 trimethyla�on and H4K16 acetyla�on are linked to gene expression.

o Combina�on of Marks: Specific combina�ons of marks convey dis�nct "meanings"

for chroma�n func�on.

Barrier Sequences

1. Preven�ng Uncontrolled Spread:

o Barrier DNA sequences block the spread of heterochroma�n to ensure chroma�n

domains remain dis�nct.

o For example, HS4 sequence separates ac�ve chroma�n from silenced regions.
Dele�on of HS4 leads to abnormal spreading and gene silencing, impac�ng gene
expression.

2. Barrier Mechanisms:

o Tethering: Chroma�n tethered to a fixed nuclear structure (e.g., nuclear pore) acts as
a barrier.

o Barrier Proteins: Proteins bound to specific nucleosomes resist heterochroma�n

spreading.

o Recruitment of Modifying Enzymes: Barrier sequences can recruit histone-

modifying enzymes (e.g., acetylases) that erase marks needed for spreading.

3. Example - HS4 Sequence in Red Blood Cells:

o HS4 protects the β-globin gene from silencing by heterochroma�n. Without HS4, β-
globin is silenced, leading to anemia

Nucleosomes with Histone Variants

 • Dis�nc�ve Character of Nucleosomes:

o Nucleosomes containing histone variants, such as CENP-A (Centromere Protein-A),

are essen�al for establishing specific chroma�n marks that are stable and long-
las�ng.

o CENP-A is a variant of histone H3 and plays a pivotal role in forming the centromere
structure.

Centromere Structure and Func�on

• Role of Centromeres:

o Centromeres are crucial for the proper atachment of chromosomes to the mito�c
spindle, ensuring accurate segrega�on of duplicated genomes during cell division.

o Each centromere is surrounded by specialized centromeric chroma�n that remains

intact during interphase, even though spindle atachment and DNA movement occur
only during mitosis.

• Centromere Composi�on:

o In organisms like budding yeast (S. cerevisiae), a specific DNA sequence of

approximately 125 nucleo�des is sufficient to func�on as a centromere. This
sequence forms a nucleosome with the CENP-A variant histone along with three
other core histones.

o In more complex organisms, such as humans and flies, centromeres are much larger,
spanning hundreds of thousands of nucleo�de pairs, and o�en do not contain a
defined centromere-specific DNA sequence.

Alpha Satellite DNA

• Human Centromeres:

o Human centromeres primarily consist of repeated short DNA sequences known as

alpha satellite DNA. These sequences can repeat thousands of �mes and are
surrounded by pericentric heterochroma�n.

o Although alpha satellite DNA is prevalent at centromeres, it is also found in non-

centromeric regions of chromosomes, indica�ng that it alone is insufficient for
centromere specifica�on.

• Neocentromere Forma�on:

o In certain instances, new centromeres (termed neocentromeres) can form on

fragmented chromosomes, even in regions originally lacking alpha satellite DNA. This
phenomenon illustrates the flexibility and adaptability of centromere forma�on.

Evolu�onary Role of Centromeres

• Inac�va�on and Genesis:

o The inac�va�on of certain centromeres and the spontaneous forma�on of new ones
are significant factors in evolu�onary processes.
 o Chromosomal changes, o�en resul�ng from breakage-and-rejoining events, can lead
to altera�ons in centromere numbers, impac�ng stable inheritance paterns.

o Different species, even those closely related, can exhibit significant differences in
chromosome numbers, highligh�ng the role of centromeres in chromosomal
evolu�on.

Epigene�c Inheritance of Chroma�n Structures

Mechanism of Inheritance

• Epigene�c Changes:

o The stability of changes in centromere ac�vity requires mechanisms that allow these
changes to be passed down through subsequent genera�ons.

o The ini�al forma�on of a centromere involves a specialized DNA-protein structure

containing nucleosomes made with the CENP-A variant of histone H3.

• Direct Inheritance:

o During DNA replica�on, H3-H4 tetramers from parent nucleosomes are inherited by
the newly formed daughter DNA strands at the replica�on fork, promo�ng the
con�nuity of centromere iden�ty across cell divisions.

o Once a stretch of DNA has assembled CENP-A-containing nucleosomes, it becomes

easier to form a new centromere in the same loca�on on both daughter
chromosomes.

Coopera�ve Forma�on of Chroma�n

• All-or-None Crea�on:

o The assembly of centromeric chroma�n appears to be a coopera�ve and all-or-none

process, sugges�ng that the forma�on of this structure is highly integrated and
requires mul�ple components to work together.

o The phenomenon resembles posi�on effect variega�on, where specific chroma�n

structures are propagated along chromosomes and inherited across cell genera�ons.

• Role of Reader-Writer Complexes:

o The coopera�ve recruitment of proteins, along with the ac�vity of reader-writer

complexes, plays a crucial role in the spreading of specific chroma�n forms. This
process ensures that chroma�n structures are not only established but also
maintained and propagated through cell divisions.

o In centromeric regions, proteins marking chroma�n with specific modifica�ons (e.g.,

dimethyl lysine 4 on H3) are involved in this coopera�ve process, allowing for the
establishment of dis�nct chroma�n domains.
CHROMOSOMES

Structure of Chromosomes in Interphase Cells

Lampbrush Chromosomes

• Defini�on and Characteris�cs:

o Lampbrush chromosomes are unique to certain amphibians and represent the

largest known chromosomes.

o They are found in growing oocytes (immature eggs) that are preparing for meiosis.

o These chromosomes are visible under a light microscope due to their unusual
structure and extensive length.

• Structure:

o Lampbrush chromosomes consist of a linear chromosomal axis from which large

chroma�n loops extend.

o Each loop contains the same specific DNA sequence that remains extended as the
oocyte grows, indica�ng that these loops are sites of ac�ve transcrip�on.

• Gene Expression:

o The genes located within these DNA loops are ac�vely expressed, contribu�ng to the
produc�on of large amounts of RNA required for oocyte development.

o In contrast, the majority of the DNA remains highly condensed along the
chromosome axis, where genes are generally not expressed.

• Loop Structure:

o A typical set of lampbrush chromosomes in amphibians has around 10,000 loops.

o Each chromosome consists of paired sister chroma�ds, resul�ng in four copies of

each loop within the cell.
 o This four-stranded structure is characteris�c of the diplotene stage of meiosis, where
the oocyte is arrested.

Interphase Chromosome Organiza�on

• General Arrangement:

o It is believed that interphase chromosomes in all eukaryotes are similarly organized

into loops, although these loops are typically too small and fragile to be directly
observed under a light microscope.

• Detec�on of Loops:

o Modern DNA technologies enable the assessment of the proximity of different loci
on an interphase chromosome, providing insights into the loop structures.

o These methods help infer the presence and organiza�on of chroma�n loops, even if
they cannot be directly visualized.

• Size of Loops:

o The loops of DNA within interphase chromosomes can vary in size, with typical loops
containing between 50,000 and 200,000 nucleo�de pairs.

o In some cases, loops of up to one million nucleo�de pairs have also been suggested,
indica�ng a significant variability in loop structure and organiza�on.

Polytene Chromosomes

Defini�on and Characteris�cs

• Forma�on:

o Polytene chromosomes are formed from the alignment of all copies of each
chromosome side by side in exact register, resembling drinking straws in a box.

o This arrangement allows for the detec�on of features that may be shared with
ordinary interphase chromosomes but are typically difficult to visualize.

• Source:

o Polytene chromosomes are notably found in the salivary glands of fruit flies
(Drosophila) and are much larger than typical chromosomes, making them easier to
study under a light microscope.

Structure and Banding Patern

• Appearance:

o When viewed under a light microscope, polytene chromosomes exhibit dis�nct

alterna�ng dark bands and light interbands.

o Approximately 95% of the DNA in polytene chromosomes is present in these bands,

while about 5% exists in the interbands.

• Band Composi�on:

o Band Thickness:
  Thin bands can contain around 3,000 nucleo�de pairs, while thick bands may
comprise up to 200,000 nucleo�de pairs in each chroma�n strand.

o Dark Bands:

 The chroma�n in bands appears darker due to higher condensa�on

compared to interbands, which may also reflect a greater concentra�on of
associated proteins.

• Chromosome Mapping:

o Drosophila polytene chromosomes contain approximately 3,700 bands and an equal

number of interbands.

o Each band is iden�fied by its thickness and spacing, allowing for the genera�on of a
chromosome map indexed to the fly's complete genome sequence.

Importance in Chroma�n Studies

• Large-Scale Organiza�on:

o Polytene chromosomes serve as an effec�ve model for examining the large-scale

organiza�on of chroma�n.

o They provide insights into how chroma�n is structured and func�onally organized
across the genome.

• Chroma�n Types:

o Various forms of chroma�n exist, each characterized by nucleosomes containing

different combina�ons of modified histones.

o Specific non-histone proteins associate with these nucleosomes, influencing

biological func�ons in diverse ways.

• Spreading of Protein Effects:

o The recruitment of certain non-histone proteins can extend over long distances
along the DNA, impar�ng a similar chroma�n structure across broad genomic
regions.

Mito�c Chromosomes

Overview of Chromosome Structure During Mitosis

• Visibility in Mitosis:

o Chromosomes in nearly all eukaryo�c cells become highly visible through light
microscopy during mitosis due to condensa�on.

o The length of a typical interphase chromosome is reduced about tenfold during this
process, leading to a drama�c change in appearance.

Structure of Mito�c Chromosomes

• Metaphase Chromosomes:
 o A typical mito�c chromosome at the metaphase stage consists of two sister
chroma�ds, each containing one of the two iden�cal DNA molecules produced by
DNA replica�on during interphase.

o Sister chroma�ds are held together at their centromeres.

• Chroma�d Organiza�on:

o Chroma�ds are covered with

various molecules, including RNA-
protein complexes.

o Once these coverings are stripped

away, electron microscopy reveals
that each chroma�d is organized
into loops of chroma�n emana�ng
from a central scaffolding.

• Gene Order and Chroma�n Structure:

o Experiments using DNA

hybridiza�on show that the visible
features along a mito�c
chromosome reflect the order of
genes along the DNA molecule,
indica�ng a level of organiza�on in
the chroma�n structure.

Func�ons of Chromosome Condensa�on

• Separa�on of Sister Chroma�ds:

o The compac�on ensures that sister chroma�ds are disentangled and posi�oned side
by side, facilita�ng their separa�on by the mito�c apparatus during cell division.

• Protec�on of DNA:

o Condensa�on protects fragile DNA molecules from breaking as they are pulled apart
into separate daughter cells during mitosis.

Dynamics of Chromosome Condensa�on

• Timing and Connec�on to Cell Cycle:

o The condensa�on of interphase chromosomes into mito�c chromosomes begins in

early M phase and is closely linked to cell cycle progression.

o During M phase, gene expression is largely shut down, and specific modifica�ons to
histones occur to facilitate chroma�n reorganiza�on.

• Role of Cohesins and Condensins:

o Two classes of ring-shaped proteins, cohesins and condensins, play cri�cal roles in
the compac�on of chromosomes, helping to produce the two separately folded
chroma�ds of a mito�c chromosome.