Cytogenetics

Foundations
 GENETICS
1. Human genetics
2. Medical genetics
a. Clinical genetics
b. Genetic Counseling
c. Cytogenetics
d. Molecular genetics
e. Biochemical genetics
 TERMINOLOGIES
v Chromosome- a highly ordered
structure composed of DNA and
proteins that carries the genetic
information
v HOMOLOGOUS
CHROMOSOMES OR
HOMOLOGS- sister
chromosomes, the members of a
pair of chromosomes in which one
is inherited from the mother and
the other from the father.
 TERMINOLOGIES
v LOCUS- the position of a gene on a
chromosome

v ALLELE- an alternative form of a gene

occupying the same locus. An allele
may be the result of a mutation. There
is a maximum of two alleles per diploid
chromosome complement (one allele
per chromosome), but multiple alleles
may exist within a population
 TERMINOLOGIES
v DIPLOID- the presence of two copies of each unique
chromosome per cell. In humans, the chromosomes occur in
pairs and the diploid (2N) number is 46.

v HAPLOID- one copy of each unique chromosome. In

humans, the gametes are haploid (N=23)
 TERMINOLOGIES
v HOMOZYGOUS- both alleles at a locus are the same

v HETEROZYGOUS- the two alleles at a locus are different.

v HEMIZYGOUS- the presence of only one chromosome or

chromosome segment rather than the usual two; applies to
males with a single X chromosome
 TERMINOLOGIES
v GENOTYPE- the genetic
constitution of an individual
or organism (ex. What alleles
are present)

v PHENOTYPE- the
appearance of an individual
that results from the
interaction of environment
and genotype
 TERMINOLOGIES
v DOMINANT ALLELE- an allele
that is expressed when present
in only a single dose
v RECESSIVE ALLELE- in a
diploid organism, an allele that is
only expressed when homozygous
v CODOMINANT ALLELE- in a
diploid organism, allele that show
no dominance or recessivity to
each other but, when present
together, are both fully
expressed
 TERMINOLOGIES
v MUTATION – a permanent heritable change in the
sequence of genomic DNA.
v Constitutional- a mutation present in every cell of the
body
v Acquired- may arise in a single somatic cell, notably
cancer
TERMINOLOGIES
 01

Cytogenetics 02

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Defined as the science of
combines the methods and 04

findings of cytology and 05

genetics to allow the
investigation of heredity at 06

the cellular level

 01

Cytogenetics 02

03
The study of chromosome
number, structure, function, 04

and behavior in relation to 05

gene inheritance, organization
and expression 06
 Background
Chromosomes were first observed by Walther Fleming in
1882, only 16 years after Mendel established genetics as
a new field of science, making cytogenetics one of the
oldest fields of genetics.

Just after the turn of the century, the importance of the

sex chromosome was established and in 1959, cytogenetic
studies were first utilized in clinical laboratory studies
 01

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The Cell 04

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The Cell
 The Cell
v Cell serves as the structural building block to form tissues and
organ
v Each cell is functionally independent- it can live on its own
under the right conditions:
v it can define its boundaries and protect itself from external
changes causing internal changes
v it can use sugars to derive energy for different processes
which keep it alive
v it contains all the information required for replicating itself
and interacting with other cells in order to produce a
multicellular organisms
v It is even possible to reproduce the entire plant from
almost any single cell of the plant
 The Cell
v Cell wall
v protects and supports cell
v made from carbohydrates- cellulose and pectin-
polysaccharides
v strong but leaky- lets water and chemicals pass through-
analogous to a cardboard box

v Cell membrane
v membrane is made up from lipids - made from fatty acids
water-repelling nature of fatty acids makes the diglycerides
form a sheet or film which keeps water from moving past
sheet (think of a film of oil on water)
 The Cell
v Organelles
v sub-compartments within the cell which provide different
functions. Each organelle is surrounded by a membrane that
makes it separate from the cytosol. These include nucleus,
mitochondrion, vacuole, ribosome, endoplasmic reticulum,
and golgi apparatus.
 The Cell
v Cell wall
v protects and supports cell
v made from carbohydrates- cellulose and pectin-
polysaccharides
v strong but leaky- lets water and chemicals pass through-
analogous to a cardboard box

v Cell membrane
v membrane is made up from lipids - made from fatty acids
water-repelling nature of fatty acids makes the diglycerides
form a sheet or film which keeps water from moving past
sheet (think of a film of oil on water)
The Cell Membrane
 CYTOSKELETON
v Adopt a particular shape
v Maintain polarity
v Organization of intracellular organelles
v Movement
v Major Cytoskeleton:
v Actin microfilaments= most abundant
v Intermediate filaments
v Microtubules= connecting cables
 CELL INTERACTIONS
v Provide mechanical links
v Recognize ligands on other cells

v 3 BASIC TYPES:
v Occluding junctions (tight junctions)
v Anchoring junctions (desmosomes)
v Communicating junctions (gap junctions)
 CELL INTERACTIONS
v Provide mechanical links
v Recognize ligands on other cells

v 3 BASIC TYPES:
v Occluding junctions (tight junctions)
v Anchoring junctions (desmosomes)
v Communicating junctions (gap junctions)
 CELL SIGNALLING
Different groups of signal:
Pathways:
v Damage to neighboring cells
and pathogens v Paracrine
Contact with neighboring
Autocrine
v
cells(gap junction signaling) v

v Contact with ECM v Synaptic

Secreted molecules (growth
v
factors, cytokines, hormones)
v endocrine
 CELL SIGNALLING
Protein Receptors: Signaling molecules are called ligands
LIGANDS on cell surface receptors: receptor cross- linking

Ligand Bonding can :

v Open ion channels
v Activate an assoc. Growth Factor signaling pathways
driving cell proliferation (G protein)
v Activate an endogenous/associated enzyme(Tyrosine
Kinase
v Trigger a proteolytic event or a change in protein
binding (Notch , Wnt and Hedgehog) - Regulate
normal dev’t
 CELL SIGNALLING
based on signaling mechanism they use and intracellular
pathways they activate
1. Receptors associated with kinase activity
v Tyrosine kinases (integral membrane proteins)
2. G-protein coupled receptors
v Guanosine diphosphate (GDP)
3. Nuclear receptors
4. Other classes of receptors (Multiple receptor pathways
regulating normal development)
v Notch family
v Wnt protein
CELL DIFFERENTIATION
 APOPTOSIS
v Form of cell death designed to eliminate unwanted host cells through activation
of a coordinated, internally programmed series of events effected by a dedicated
set of gene products
v Responsible for numerous physiologic, adaptive, and pathologic events including:
v Programmed destruction of cells during embryogenesis
v In aging—hormone-dependent involution of tissues in adults
v Cell deletion in proliferating cells, as a homeostatic mechanism to maintain
cell populations.
v As a defense mechanism such as:
v Cell death in tumors
v Death of neutrophils during acute inflammatory response
v Death of immune cells during immune reactions
v Cell death induced by cytotoxic T cells
v Pathologic atrophy in parenchymal organs after duct obstruction when cells
are damaged by disease (viral diseases), noxious agents and injurious
stimuli
 APOPTOSIS
v Morphologic features:
v Cell shrinkage
v Chromatin condensation
and fragmentation
v Formation of cytoplasmic
blebs and apoptotic
bodies,
v Phagocytosis of
apoptotic bodies by
adjacent healthy cells or
macrophages
 APOPTOSIS
v Mechanisms include:
v Chromatin condensation and fragmentation – mediated by
activation of Ca2+-sensitive endonucleases, due to increased
free cytosolic Ca2+
v Phagocytosis of apoptotic bodies is mediated by receptors on
macrophages for surface components on apoptotic cells
v Regulated by a number of apoptosis-associated genes:
v BCL2 – Inhibits apoptosis and extends cell survival
v BAX – stimulates apoptosis with excessive cell death
v p53 – stimulates apoptosis but when mutated or absent
favors cell survival
v c-MYC—either stimulates or inhibits apoptosis, depending
on the presence of other signals
 APOPTOSIS
v Apoptosis is the end point of an energy-dependent cascade of
molecular events, initiated by certain stimuli, and consisting of 4
components. Signaling pathways that initiate apoptosis include:
v Control and integration – intracellular positive and negative
regulatory molecules inhibit, stimulate, or forestall
apoptosis and thus determine the outcome
v Initiation phase – during which some cysteine proteases
called caspases become catalytically active
v Execution phase – during which other caspases trigger the
degradation of critical cellular components
v Removal of dead cells by phagocytosis
 APOPTOSIS
v Two pathways converge on caspase activation:
v Intrinsic (mitochondrial) pathway
v triggered by growth factor deprivation, DNA damage
and protein misfolding
v Associated with leakage of pro-apoptotic proteins
from the mitochondrial membrane into the cytoplasm
and activation caspases
v Extrinsic (death receptor) pathway
v responsible for elimination of self-reactive T cells and
damage by CTL
v Initiated by binding of death receptors (Fas and TNF)
by ligands on adjacent cell and activation of caspases
 01

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The Cell
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Cycle 05

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The Cell
 Control of the Cell Cycle

vCyclins
v a G1 cyclin (cyclin D)
v S-phase cyclins (cyclins E and A)
v mitotic cyclins (cyclins B and A)

vCyclin-dependent kinases (Cdks)

v a G1 Cdk (Cdk4)
v an S-phase Cdk ((Cdk2)
v an M-phase Cdk (Cdk1)

v The anaphase-promoting complex (APC)

v Aka. cyclosome (APC/C.)
 Steps of the Cell Cycle

v A rising level of G1-cyclins bind to their Cdks and signal

the cell to prepare the chromosomes for replication.
v A rising level of S-phase promoting factor (SPF) — which
includes cyclin A bound to Cdk2 — enters the nucleus and
prepares the cell to duplicate its DNA (and its
centrosomes)
v As DNA replication continues, cyclin E is destroyed, and
the level of mitotic cyclins begins to rise (in G2).
v M-phase promoting factor (the complex of mitotic
cyclins with the M-phase Cdk) initiates assembly of the
mitotic spindle
 Steps of the Cell Cycle
v At this point, the M-phase promoting factor activates the
anaphase-promoting complex (APC/C) which o allows
the sister chromatids at the metaphase plate to separate
and move to the poles (= anaphase), completing mitosis;
v destroys cyclin B. It does this by attaching it to the
protein ubiquitin which targets it for destruction by
proteasomes.
v turns on synthesis of G1 cyclin for the next turn of the
cycle;
v degrades geminin, a protein that has kept the freshly-
synthesized DNA in S phase from being re-replicated
before mitosis.
 Meiosis and the Cell Cycle
v The special behavior of the chromosomes in meiosis I
requires some special controls. Nonetheless, passage
through the cell cycle in meiosis I (as well as meiosis II,
which is essentially a mitotic division) uses many of the
same players, e.g., MPF and APC. (In fact, MPF is also
called maturation-promoting factor for its role in meiosis
I and II of developing oocytes.
 Quality Control of the Cell Cycle
v A check on completion of S phase. The cell seems to
monitor the presence of the Okazaki fragments on the
lagging strand during DNA replication. The cell is not
permitted to proceed in the cell cycle until these have
disappeared.
v  DNA damage checkpoints. These sense DNA damage
v before the cell enters S phase (a G1 checkpoint);
v during S phase, and
v after DNA replication (a G2 checkpoint).
 Quality Control of the Cell Cycle
v Spindle checkpoints. Some of these that have been
discovered
v detect any failure of spindle fibers to attach to
kinetochores and arrest the cell in metaphase
(M checkpoint);
v detect improper alignment of the spindle itself
and block cytokinesis;
v trigger apoptosis if the damage is irreparable.
 Regulation of the Cell Cycle
1. The cell cycle control system consists of a molecular clock
and a set of checkpoints that ensure that appropriate
conditions have been met before the cycle advances.

2. For instance, cells must be in contact with adjacent cells

before proper division can occur. Also, cells must reach a
certain size and volume before they can properly divide. All
of the DNA must be properly replicated before the cell
divides.
 Regulation of the Cell Cycle
3. Checkpoints are present in the G1, G2, and M phases of the
cell cycle. The G1 checkpoint is the most critical one for many
cells.

4. If the proper signals are not received, the cell may stay in
a stage known as G0; or the non-dividing state.

5. Protein Kinases are enzymes that help synchronize the cell

cycle events. Protein Kinases catalyze the transfer of a
phosphate group from ATP to a target protein.

6. Phosphorylation induces a conformational change that

either activates or inactivates a target protein.
 Regulation of the Cell Cycle
7. Changes in these target proteins affect the progression
through the cell cycle.

8. Cyclical changes in kinase activity, in turn, are control

9. Protein kinases that regulate cell cycles are active only

when attached to a particular Cyclin molecule.

10. Cyclin concentrations, in turn, vary throughout the cell

cycle (they are highest as the cells prepare to divide). By the
end of cytokinesis, cyclins are present in much smaller
concentrations. The cyclins are broken down as the cells
progress through the M-phase of cell division.
 Regulation of the Cell Cycle
11. Cyclins bind with protein kinases early in the cell cycle and
produce Mitosis Promoting Factor (MPF). MPF promotes
chromosome condensation and nuclear membrane
absorption.

12. Later in the cell cycle, MPF activates proteolytic enzymes

(these enzymes break down proteins) which destroy the
cyclin.

13. Thus, new Cyclin proteins must be produced during

interphase, until appropriate levels build up and promote cell
division.
Regulation of the Cell Cycle
MITOSIS
MEIOSIS
MEIOSIS
 Meiotic Errors

v Nondisjunction- homologues don't separate in meiosis 1

vresults in aneuploidy
vusually embryo lethal
vTrisomy 21, exception leading to Downs syndrome
vSex chromosomes
vTurner syndrome: monosomy X
vKlinefelter syndroms: XXY
v Translocation and deletion: transfer of a piece of one
chromosome to another or loss of fragment of a
chromosome.
Meiotic Errors
Meiotic Errors
Meiosis and Recombination
 01

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