Download Eukaryotic and Prokaryotic Cells

DNA Architecture and
Physical Properties;
Organellar DNA
AHMP 5406
Objectives:
Describe the molecular structure of DNA
Describe the packing and organization of
DNA into eukaryotic chromosomes
Discuss the function, structure and
components of nucleosomes
Understand the difference between
nuclear and organellar codon usage
DNA Double Helical Structure
Eukaryotic Chromosomes
Consist of highly coiled DNA,
RNA, histones and nonhistone proteins
Chromosomes are numbered
from largest to smallest 1
through 22, plus X and Y
Eukaryotic Chromosomes
Chromosomes are distinguished by:
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size
centromere location
differential staining
DNA probes
Eukaryotic chromosomes
In metaphase of mitosis,
chromosomes can be seen under
microscope
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they have a compact rod-like structure
The ends of chromosome are called
telomeres, function is to protect the
ends of the DNA
Eukaryotic chromosomes
Near the middle is the
centromere
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Function is to attach to spindles
during cell division
Ensure correct segregation
Telomeres and centromeres
contain special DNA sequences
and associated proteins
Eukaryotic chromosomes
Telomeres are replicated differently from the rest of the genome
Different regions of the chromosome can be stained with dyes
(e.g. Giemsa) giving a characteristic banding pattern
Eukaryotic
chromosome
structure
(heterochromatin)
Genes, repeated sequences,
replication origins
(mostly euchromatin)
Telomeres (heterochromatin)
Chromatin
packing
?
Histones – What are they???
DNA packing material
Can be acetylated and interact
in gene transcription
Methylation of lysines can
inhibit or enhance transcription
Histones – What are they???
Positively charged proteins
– lots of Arg and Lys
Octameric
5 different kinds – 4 are
found in nucleosomes, one
kind joins linker regions of
DNA
Histones – What are they???
Small proteins
 102-135 AA
Similar structural motif
 Three a-helices
 Connected by two loops
 Histone-fold
H3-H4 dimer
H2A-H2B dimer
The nucleosome
DNA wrapped in 1.6 lefthanded turns
DNA 146 bp
11 nm in width
Bending of DNA around nucleosome
A-T rich sequences are
easier to bend
Explains precise
positioning of
nucleosomes along DNA
Proteins also affect
binding
Euchromatin vs. Heterochromatin
Heterochromatin
Is found in parts of the chromosome where
there are few or no genes, such as
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centromeres
telomeres
transposons and other "junk" DNA
Heterochromatin
Is densely-packed
Has reduced crossing over in meiosis
Those genes present in heterochromatin
are generally inactive; that is, not
transcribed…How?
Heterochromatin
Decreased acetylation of histones
Decreased methylation of lysine-9 in
histone H3,
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which now provides a binding site for
heterochromatin protein 1 (HP1),
blocks access to transcription factors needed
for gene transcription
Euchromatin
Found in parts of the
chromosome that contain
many genes
Loosely-packed in loops of
30-nm fibers
Euchromatin
Loops are separated from
adjacent heterochromatin by
insulators
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Boundary sequences
Loops are often found near
the nuclear pore complexes
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Makes sense for the gene
transcripts to get to the cytosol
Euchromatin
The genes in euchromatin are active and
thus show
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decreased methylation of the cytosines in
CpG islands of the DNA
increased acetylation of histones and
decreased methylation of lysine-9 in histone
H3.
Signaling by Modification of
Nucleosome Histone Proteins
Modification of Histone tails
Histone tails are subject to covalent
modification
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Lysine
methylation (M)
acetylation (Ac)
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Serine
phosphorylated (P)
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Ubiquitin (U)
76 AA protein
Modification of Histone tails
Histone acetyl transferases (HATs)
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Add acetyl groups
Histone deacetylases (HDACs)
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Remove acetyl groups
Affects stability of 30 nm fiber
Can attract specific proteins to modified chromatin
Some positions can be modified more than one way
Modification of Histone tails
Convey messages to the cell
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E.g Chromatin is newly replicated
Or gene expression should not take place
Different modifications can attract
different protein complexes
Allows chromatin to be dynamic
Modification of Histone tails
Only a few effects
of specific
modifications
known
Examples of Histone tail mods
Gene expression
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Ac of lysine (K) 14
on H3
Ac of K 8 and 16
on H4
P (S 10) and Ac (K
14) on H3
Examples of Histone tail mods
Gene silencing
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Unmodified H3
and H4
M of K 9 on H3
Other parts of the chromosome in
detail
Centromeres
Essential for correct
segregation of chromosomes
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During cell division
By attachment to spindles
Consists of a small, core DNA
sequence (AT rich) and specific
proteins
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a satellite DNA
In many species (e.g. humans)
flanked by 100s of copies of a
tandemly-repeated sequences
Centromeres
Packaged as heterochromatin
Has specialized packaging
structures
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Centromere-specific proteins
Centromere-specific nucleosomes
H3 variant = CENP-A
Facilitate construction of other
centromere binding proteins
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These attach to kinetochore
Telomeres
Repetitive sequence at the ends of chromosomes
TTAGGG
Many thousands of repeats
Lose a few repeat units with each replication cycle
Protects the genes contained in the chromosomes
The problem of telomere
replication
More on genome organization
Compared to prokaryotes
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Eukaryotic genomes are completely different
in their organization and much larger
Their genes are mostly “split” into exons
and introns
Exons may allow evolution of proteins in a
“modular” way
Eukaryotic gene organization
5’UTR
Exon1
Intron 1
Exon2
Intron 2
Exon 3
Promoter
Primary RNA transcript
splicing
mRNA
Protein
3’UTR
The human genome is complex:
Repetitive/Mobile
Elements
Origins and function of DNA
classes
Highly repetitive:
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Bits of old virus genomes
Simple sequence repeats e.g. CACACA….
Special sequences such as centromeres
Moderately repetitive:
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Other old virus genomes
Multi-gene families, e.g. ribosomal RNA
Single-copy:
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Most “normal” genes
Types of repeated DNA
Tandemly repeated
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Satellite DNA
Centromeric DNA
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Minisatellites (10-100 bp)
E.g. (CAGACAGTATGA)n
DNA finger printing
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Microsatellites (2-4 bp)
Can be used to study
microevolution
Biparentally inherited
High rates of mutation
Alleles = size differences
(CA)6
CACA CA CA CA CA
Types of repeated DNA
Interspersed
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SINES
Short interspersed nuclear
elements
Approx. 500bp
Reversed transcribed RNA
sequences
Non-coding
Ex. In humans Alu repeats
(CA)6
CACA CA CA CA CA
Types of repeated DNA
Interspersed
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LINES
Long int. nucl. elem.
Approx. 5Kb
Code for two proteins
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Reverse transcriptase and
endonuclease activity
RNA binding ability
Increase genome size by
copying themselves
(CA)6
CACA CA CA CA CA
Transposons
Move by “cut-n-paste”
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Transposon is cut out of
its location by an
enzyme
Transposase is encoded
within the transposon
Then inserted into a new
location
Transposons and disease
Transposons are mutagens
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Functional gene insertion
Can result in damage
Alter gene function
Inactivate protein
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Faulty repair of the gap left at the old site
The string of identical repeats can cause problem in
base pairing during mitosis
unequal crossing over
THE GENETIC CODE
The Genetic Code
Rules by which DNA
encode AAs
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Each codon = an AA
Thus determine
polypeptide sequence
Protein function
The mitochondrial genome
Study of human
mtDNA
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Different code found
Other variants have
been found
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Bacteria
Nuclear vs. mtDNA