Chapter Two
Chromsome Structure & Function
1. Ploidy levels and the cell cycle
• The chromosome set is the number of different
chromosomes in a nucleated cells and is designated n
and the associated DNA content is designated C.
• In humans, n=23 and C=ca. 3.5 pg (3.5 x 10-12 g). The
DNA content of a cell during the cell cycle is 2C from
anaphase/telophase of mitosis (M) until right before
entering S phase while between S phase and mitosis the
DNA content is 4C.
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• In the germline, the DNA content is 2C
before S phase then 4C during S until the
anaphase/telophase of meiosis I. At the end
of meiosis I, each reduced cell has 2C DNA
content. By the end of meiosis II, each
haploid gamete (egg or sperm is C).
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2. Chrosmosome structure and function:
• In human chromosomes, DNA is packaged in
multiple hierarchies of DNA folding.
• DNA wraps around a histone octamer to form the
10 nm string of beads. The latter further coil to
form the 30 nm chromatin fiber (interphase). The
30 nm fibers compacts further forming a rosette
shape and attaches to a scaffold of acidic proteins
(topoisomersaes II).
• The packaging ratio for DNA in human
chromosomes is: 1:6 for nucleosomes, 1:36 for the
30 nm fiber, and >1:10,000 for the metaphase
chromosome.
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• Chromosome organization in a human interphase
is not random. Besides the nucleolus (site of
rRNA synthesis) attaching to the nucleolar
Organization Region (NOR), chromosomes
occupy certain chromosome territories. For
example, the most gene-rich chromosomes tend to
concentrate at the center of the nucleus whereas
the more gene-poor chromosomes tend to locate
towards the nuclear envelope.
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• Centromere: is the primary constriction in the
chromosome and the site at which the the large
multiprotein complexes (kinetochore) attach to
each of the centromere at late prophase.
• In yeast, CEN (centromere element) is 110 bp long
containing two highly conserved elements (9 and
11 bp) flanking a central AT rich region.
• In mammals, centromeres consist of 100s of
kilobases of repetitive DNA (some is chromosome
specific). In humans, alpha-satellite is a complex
family of tandemly repeated 171 bp core
sequence. Several proteins such as CANP-B bind
to the alpha satellite region.
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• Origins of replication: In simple eukaryotes
(yeast) it is known as autonomously
replicating sequences (ARS). In mammals,
due to the lack of a genetic assay, origins of
repliaction are less well defined.
Mammalian artificial chromosomes seem to
work without specific ARS sequences.
• Telomeres: function in
- Maintaining structural integrity by preventing the
fusion of chromosome ends or protect the ends of
chromosomes from degradation by binding to
specific telomere-binding proteins.
- Ensure complete DNA replication at the tips of
chromosmes.
- Chromosome positioning.
• In a human telomere, the hexanucleotide
TTAGGG is repeated to span about 3-20 kb and
upstream of this repeat and heading towards the
centromere exist 100-300 kb of telomereassociated repeats before any unique sequence
occurs.
• Telomearse is an RNA-protein enzyme that
extends the 3’ overhang at the end of
chromosomes.
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• Heterochromatin (darkly stained) and euchromatin
(lightly stained):
- Genes is euchromatin may or may not be
expressed while genes in heterochromatin are
unlikely to be expressed.
- Constitutive heterochromatin.
- Facultative heterochromatin (e.g. X-inactivation in
females or X & Y silencing during male meiosis
for a period of 15 days).
3. Mitosis and meiosis:
• Mitosis is the somatic cell division.
• Meiosis produces sperms and eggs cells
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• Meiosis contributes to genetic diversity in two ways:
- by the independent assortment of maternal and
paternal chromosomes during anaphase I. There are
223 or 8.4 million possible different combinations of
parental chromosomes to be produced by one person
per gamete.
- Recombination (crossing over between paternal and
maternal chromosomes in prophase I).
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• X-Y pairing and the pseudoautosomal region:
- The pairing of X & Y in males is made possible by
a 2.6 Mb region at the tips of their short arms.
This region is known as the major
pseudoautosomal region and genes in this area are
not subject to inactivation and they obligatory
crossover resembling normal autosomal genes (i.e.
do not follow the X-linked mode of inheritance).
- A 320 Kb region of homolgy between X & Y exist
at the tip of their long arms but pairing and
crossing over is not obligatory.
4. Visualizing human chromosomes
(Cytogenetics):
• Mitotic chromosomes could be stained and
visualized but meiotic chromosomes are hard to
visualize. White blood cells are cultured in a
medium with phytohemagglutinin to induce cell
division, synchronized by adding a thymidine
analog, and colcemid to disrupt spindle fibre
formation.
• Karyotyping and chromosome banding. 46,XX or
46,XY. Human chromosomes have a total of 850
bands at high resolution. G bands (dark condensed
chromatin) replicate in late S phase while G bands
(light less condensed chromatin) replicate in early
S phase. G bands have lower %GC content than R
bands.
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• Molecular cytogenetics using FISH
(fluorescence in situ hybridization). Probe is
labeled by incorporation of fluorescentlabeled nucleotide precursors or by
incorporation of a nucleotide carrying a
reporter molecule (biotin or digoxigenin)
which is detected that is detected by binding
to a fluorescently labeled affinity molecule.
• Resolution of metaphase FISH is several
Mb while that of prometaphase FISH is 1
MB. For higher resolution interphase FISH
is used.
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• Chromosome painting is achieved by using probes
composed of a large collection of different DNA
fragments from a single chromosome (such probes
are obtained from chromosome-specific DNA
libraries or by using Alu-PCR of DNA extracted
from monochromosomal hybrid cells ). This
technique is used to identify chromosomal
rearrangements in cancer patients.
• Molecular karyotyping using mixed fluorophores
(one per chromosome) also known as multiplex
FISH (M-FISH) allowed all 24 human
chromosomes to be identified by color.
5. Chromosome abnormalities:
• Result from missrepair of broken
chromosomes, by improper recombination,
or by malsegregation of chromosomes
during mitosis or meiosis.
• Two types:
1. Constitutional where all cells of the body
have the abnormality. This results from a
defective gamete or abnormal fertilization.
2. Somatic: occur only in certain cells or
tissues of the body. This results in a mosaic
individual.
• Numerical chromosome abnormalities: loss
or gain of complete chromosomes.
- Polyploidy: where two sperms fertilize one
egg (dispermy) or a diploid gamete
resulting in a triploid 3n (lethal). Tetrapoild
(4n) is extremely rare and results from
failure to complete the first zygotic division.
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- Aneuploidy: one or more chromosome is
missing or present in an extra copy or two.
Trisomy (2n + 1) e.g. trisomy 21
(47,XX,+21 or 47,XY,+21) also known as
Down Syndrome; Klinefelter (47, XXY)
Monosomy (2n-1) e.g. Turner (45,X).
Causes of aneuploidy are:
1. nondisjunction
2. anaphase lag.
• Mixploidy: two or more genetically different cell
lineages within one individual. Genetically
different cell populations can arise from one
zygote (mosaicism) or more rarely can originate
from different zygotes (chimerism)
- Aneuploidy mosaics (e.g. 2n/2n+1 are common)
due to non-disjunction or chromosome lag in early
mitotic divisions of the zygote.
- Polyploidy mosaics (e.g. 2n/3n are occasionally
found) mostly arise by fusion of the second polar
body with one of the cleavage nuclei of a normal
diploid zygote.
• Clinical consequences of numerical
abnormalities: (Table 2.4)
- Autosomal monosomies are more
devastating than trisomics. Trisomic
embryos survive longer than monosomic
ones.
- sex chromosome aneuploids is less
devastating than in autosomal aneupoilds.
This is because of X-inactivation
mechanisms and the fact that Y carries very
few genes that determine male sex.
• Structural chromosome aberrations:
- Result from misrepair or or mal-recombination.
- chromatid breaks (occur after replication in S
phase) .
- chromosome breaks occur in G1 phase.
- misrepair of breaks could result in acentric
chromosomes, dicentric chromosomes,
isochromosomes.
- structural abnormalities are balanced (no net gain
or loss of genes) or unbalanced (there is net gain
or loss of genes).
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• Robertsonian translocation are considered
balanced even though some material is lost
(the lost material is highly redundant rRNA
genes).
• Unbalanced abnormalities can arise through
deletion, duplication, or malsegregation of
chromosomes during meiosis of a balanced
abnormality.
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